JP2016128396A - Pharmaceutical compositions containing ips cell culture supernatant and production methods thereof, cosmetics and production methods thereof, antiaging compositions, methods for inhibiting disease onset, methods for treating diseases, methods for treating tissue disorders, and cosmetic methods - Google Patents
Pharmaceutical compositions containing ips cell culture supernatant and production methods thereof, cosmetics and production methods thereof, antiaging compositions, methods for inhibiting disease onset, methods for treating diseases, methods for treating tissue disorders, and cosmetic methods Download PDFInfo
- Publication number
- JP2016128396A JP2016128396A JP2015003493A JP2015003493A JP2016128396A JP 2016128396 A JP2016128396 A JP 2016128396A JP 2015003493 A JP2015003493 A JP 2015003493A JP 2015003493 A JP2015003493 A JP 2015003493A JP 2016128396 A JP2016128396 A JP 2016128396A
- Authority
- JP
- Japan
- Prior art keywords
- pharmaceutical composition
- disease
- culture supernatant
- cells
- ips
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 239000012228 culture supernatant Substances 0.000 title claims abstract description 149
- 239000008194 pharmaceutical composition Substances 0.000 title claims abstract description 148
- 238000000034 method Methods 0.000 title claims abstract description 130
- 201000010099 disease Diseases 0.000 title claims abstract description 122
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 title claims abstract description 122
- 239000002537 cosmetic Substances 0.000 title claims abstract description 72
- 238000004113 cell culture Methods 0.000 title claims abstract description 69
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 40
- 239000000203 mixture Substances 0.000 title claims abstract description 29
- 230000003712 anti-aging effect Effects 0.000 title claims abstract description 23
- 230000002401 inhibitory effect Effects 0.000 title abstract description 10
- 208000026062 Tissue disease Diseases 0.000 title abstract 2
- 238000012258 culturing Methods 0.000 claims abstract description 45
- 210000004027 cell Anatomy 0.000 claims description 220
- 210000001519 tissue Anatomy 0.000 claims description 144
- 230000005856 abnormality Effects 0.000 claims description 101
- 238000011282 treatment Methods 0.000 claims description 55
- 206010008118 cerebral infarction Diseases 0.000 claims description 42
- 230000032683 aging Effects 0.000 claims description 32
- 210000000130 stem cell Anatomy 0.000 claims description 27
- 208000019425 cirrhosis of liver Diseases 0.000 claims description 23
- 208000026106 cerebrovascular disease Diseases 0.000 claims description 22
- 238000004108 freeze drying Methods 0.000 claims description 22
- 239000002904 solvent Substances 0.000 claims description 21
- 201000006474 Brain Ischemia Diseases 0.000 claims description 20
- 206010008120 Cerebral ischaemia Diseases 0.000 claims description 20
- 238000005119 centrifugation Methods 0.000 claims description 19
- 238000000502 dialysis Methods 0.000 claims description 19
- 238000007710 freezing Methods 0.000 claims description 19
- 230000008014 freezing Effects 0.000 claims description 19
- 210000001082 somatic cell Anatomy 0.000 claims description 19
- 238000010790 dilution Methods 0.000 claims description 18
- 239000012895 dilution Substances 0.000 claims description 18
- 238000001035 drying Methods 0.000 claims description 18
- 206010003694 Atrophy Diseases 0.000 claims description 17
- 230000037444 atrophy Effects 0.000 claims description 17
- 238000011033 desalting Methods 0.000 claims description 17
- 210000002569 neuron Anatomy 0.000 claims description 17
- 210000002966 serum Anatomy 0.000 claims description 17
- 210000004153 islets of langerhan Anatomy 0.000 claims description 16
- 238000003860 storage Methods 0.000 claims description 16
- 230000004770 neurodegeneration Effects 0.000 claims description 15
- 230000008672 reprogramming Effects 0.000 claims description 15
- 208000005069 pulmonary fibrosis Diseases 0.000 claims description 14
- 206010016654 Fibrosis Diseases 0.000 claims description 13
- 210000001185 bone marrow Anatomy 0.000 claims description 12
- 210000004556 brain Anatomy 0.000 claims description 12
- 239000000047 product Substances 0.000 claims description 12
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims description 11
- 206010003210 Arteriosclerosis Diseases 0.000 claims description 10
- 206010006458 Bronchitis chronic Diseases 0.000 claims description 10
- 206010008111 Cerebral haemorrhage Diseases 0.000 claims description 10
- 206010012289 Dementia Diseases 0.000 claims description 10
- 206010014561 Emphysema Diseases 0.000 claims description 10
- 206010019280 Heart failures Diseases 0.000 claims description 10
- 206010019663 Hepatic failure Diseases 0.000 claims description 10
- 206010020772 Hypertension Diseases 0.000 claims description 10
- 206010023126 Jaundice Diseases 0.000 claims description 10
- 208000001132 Osteoporosis Diseases 0.000 claims description 10
- 206010035664 Pneumonia Diseases 0.000 claims description 10
- 208000017442 Retinal disease Diseases 0.000 claims description 10
- 206010040943 Skin Ulcer Diseases 0.000 claims description 10
- 208000007107 Stomach Ulcer Diseases 0.000 claims description 10
- 208000006011 Stroke Diseases 0.000 claims description 10
- 230000006793 arrhythmia Effects 0.000 claims description 10
- 206010003119 arrhythmia Diseases 0.000 claims description 10
- 208000011775 arteriosclerosis disease Diseases 0.000 claims description 10
- 206010006451 bronchitis Diseases 0.000 claims description 10
- 208000003167 cholangitis Diseases 0.000 claims description 10
- 208000007451 chronic bronchitis Diseases 0.000 claims description 10
- 230000007882 cirrhosis Effects 0.000 claims description 10
- 206010015037 epilepsy Diseases 0.000 claims description 10
- 208000001130 gallstones Diseases 0.000 claims description 10
- 201000005917 gastric ulcer Diseases 0.000 claims description 10
- 208000020658 intracerebral hemorrhage Diseases 0.000 claims description 10
- 208000007903 liver failure Diseases 0.000 claims description 10
- 231100000835 liver failure Toxicity 0.000 claims description 10
- 208000031225 myocardial ischemia Diseases 0.000 claims description 10
- 208000015122 neurodegenerative disease Diseases 0.000 claims description 10
- 208000028169 periodontal disease Diseases 0.000 claims description 10
- 231100000019 skin ulcer Toxicity 0.000 claims description 10
- 210000005036 nerve Anatomy 0.000 claims description 8
- 201000005936 periventricular leukomalacia Diseases 0.000 claims description 8
- 230000003961 neuronal insult Effects 0.000 claims description 7
- 230000037303 wrinkles Effects 0.000 claims description 6
- 208000018578 heart valve disease Diseases 0.000 claims description 5
- 238000007665 sagging Methods 0.000 claims description 5
- 230000005779 cell damage Effects 0.000 claims description 3
- 208000037887 cell injury Diseases 0.000 claims description 3
- 230000006872 improvement Effects 0.000 claims description 2
- 230000000694 effects Effects 0.000 abstract description 21
- 241000282414 Homo sapiens Species 0.000 description 33
- 239000003795 chemical substances by application Substances 0.000 description 20
- 239000003595 mist Substances 0.000 description 19
- 241000699666 Mus <mouse, genus> Species 0.000 description 18
- 230000003247 decreasing effect Effects 0.000 description 18
- 230000007423 decrease Effects 0.000 description 17
- 241001465754 Metazoa Species 0.000 description 16
- 239000002609 medium Substances 0.000 description 16
- 108090000623 proteins and genes Proteins 0.000 description 14
- 210000003491 skin Anatomy 0.000 description 14
- 239000000243 solution Substances 0.000 description 14
- 108010035532 Collagen Proteins 0.000 description 13
- 102000008186 Collagen Human genes 0.000 description 13
- 229920001436 collagen Polymers 0.000 description 13
- 239000003814 drug Substances 0.000 description 12
- 210000001988 somatic stem cell Anatomy 0.000 description 12
- 230000008439 repair process Effects 0.000 description 11
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 10
- 230000008569 process Effects 0.000 description 10
- 239000001963 growth medium Substances 0.000 description 9
- 102000004127 Cytokines Human genes 0.000 description 8
- 108090000695 Cytokines Proteins 0.000 description 8
- 241000288906 Primates Species 0.000 description 8
- 239000006228 supernatant Substances 0.000 description 8
- 230000001629 suppression Effects 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 238000002474 experimental method Methods 0.000 description 7
- 239000000463 material Substances 0.000 description 7
- 238000005507 spraying Methods 0.000 description 7
- NWIBSHFKIJFRCO-WUDYKRTCSA-N Mytomycin Chemical compound C1N2C(C(C(C)=C(N)C3=O)=O)=C3[C@@H](COC(N)=O)[C@@]2(OC)[C@@H]2[C@H]1N2 NWIBSHFKIJFRCO-WUDYKRTCSA-N 0.000 description 6
- 241000700159 Rattus Species 0.000 description 6
- 230000008859 change Effects 0.000 description 6
- OSASVXMJTNOKOY-UHFFFAOYSA-N chlorobutanol Chemical compound CC(C)(O)C(Cl)(Cl)Cl OSASVXMJTNOKOY-UHFFFAOYSA-N 0.000 description 6
- 210000002919 epithelial cell Anatomy 0.000 description 6
- 210000004072 lung Anatomy 0.000 description 6
- 210000003928 nasal cavity Anatomy 0.000 description 6
- 239000012679 serum free medium Substances 0.000 description 6
- -1 Fbx15 Proteins 0.000 description 5
- 101150086694 SLC22A3 gene Proteins 0.000 description 5
- 239000004480 active ingredient Substances 0.000 description 5
- 230000006870 function Effects 0.000 description 5
- 210000004185 liver Anatomy 0.000 description 5
- 230000007087 memory ability Effects 0.000 description 5
- 210000003205 muscle Anatomy 0.000 description 5
- 238000002360 preparation method Methods 0.000 description 5
- 241000282693 Cercopithecidae Species 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 108700021430 Kruppel-Like Factor 4 Proteins 0.000 description 4
- 101710135898 Myc proto-oncogene protein Proteins 0.000 description 4
- 102100038895 Myc proto-oncogene protein Human genes 0.000 description 4
- 241001494479 Pecora Species 0.000 description 4
- 101100247004 Rattus norvegicus Qsox1 gene Proteins 0.000 description 4
- 101710150448 Transcriptional regulator Myc Proteins 0.000 description 4
- NIJJYAXOARWZEE-UHFFFAOYSA-N Valproic acid Chemical compound CCCC(C(O)=O)CCC NIJJYAXOARWZEE-UHFFFAOYSA-N 0.000 description 4
- 238000010521 absorption reaction Methods 0.000 description 4
- 239000003242 anti bacterial agent Substances 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- 239000006143 cell culture medium Substances 0.000 description 4
- 210000003850 cellular structure Anatomy 0.000 description 4
- 230000007547 defect Effects 0.000 description 4
- 210000002242 embryoid body Anatomy 0.000 description 4
- 239000003102 growth factor Substances 0.000 description 4
- 239000007924 injection Substances 0.000 description 4
- 238000002347 injection Methods 0.000 description 4
- 208000014674 injury Diseases 0.000 description 4
- 238000001361 intraarterial administration Methods 0.000 description 4
- 238000007918 intramuscular administration Methods 0.000 description 4
- 238000007912 intraperitoneal administration Methods 0.000 description 4
- 238000001990 intravenous administration Methods 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 210000001778 pluripotent stem cell Anatomy 0.000 description 4
- 239000003755 preservative agent Substances 0.000 description 4
- RXWNCPJZOCPEPQ-NVWDDTSBSA-N puromycin Chemical compound C1=CC(OC)=CC=C1C[C@H](N)C(=O)N[C@H]1[C@@H](O)[C@H](N2C3=NC=NC(=C3N=C2)N(C)C)O[C@@H]1CO RXWNCPJZOCPEPQ-NVWDDTSBSA-N 0.000 description 4
- 230000001172 regenerating effect Effects 0.000 description 4
- 230000009758 senescence Effects 0.000 description 4
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 4
- 238000007920 subcutaneous administration Methods 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 description 3
- 241000283690 Bos taurus Species 0.000 description 3
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 3
- 208000014644 Brain disease Diseases 0.000 description 3
- 241000282472 Canis lupus familiaris Species 0.000 description 3
- 241000283707 Capra Species 0.000 description 3
- 241000700198 Cavia Species 0.000 description 3
- 241000699800 Cricetinae Species 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 241000282326 Felis catus Species 0.000 description 3
- 241000282412 Homo Species 0.000 description 3
- 241000699670 Mus sp. Species 0.000 description 3
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 3
- 241000282887 Suidae Species 0.000 description 3
- 230000001154 acute effect Effects 0.000 description 3
- 101150063416 add gene Proteins 0.000 description 3
- 238000010171 animal model Methods 0.000 description 3
- 229940121375 antifungal agent Drugs 0.000 description 3
- 239000003429 antifungal agent Substances 0.000 description 3
- 230000003796 beauty Effects 0.000 description 3
- 230000008499 blood brain barrier function Effects 0.000 description 3
- 210000004204 blood vessel Anatomy 0.000 description 3
- 210000001218 blood-brain barrier Anatomy 0.000 description 3
- 210000000988 bone and bone Anatomy 0.000 description 3
- 229960004926 chlorobutanol Drugs 0.000 description 3
- 230000003750 conditioning effect Effects 0.000 description 3
- 230000004069 differentiation Effects 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- 210000001198 duodenum Anatomy 0.000 description 3
- 239000003995 emulsifying agent Substances 0.000 description 3
- 210000003238 esophagus Anatomy 0.000 description 3
- 210000001508 eye Anatomy 0.000 description 3
- 210000002950 fibroblast Anatomy 0.000 description 3
- 230000004761 fibrosis Effects 0.000 description 3
- 210000003128 head Anatomy 0.000 description 3
- 229920002674 hyaluronan Polymers 0.000 description 3
- 229960003160 hyaluronic acid Drugs 0.000 description 3
- 230000006698 induction Effects 0.000 description 3
- 239000003112 inhibitor Substances 0.000 description 3
- 210000003734 kidney Anatomy 0.000 description 3
- 210000002429 large intestine Anatomy 0.000 description 3
- 230000013016 learning Effects 0.000 description 3
- 229960004857 mitomycin Drugs 0.000 description 3
- 201000001119 neuropathy Diseases 0.000 description 3
- 230000007823 neuropathy Effects 0.000 description 3
- 210000000496 pancreas Anatomy 0.000 description 3
- 238000007911 parenteral administration Methods 0.000 description 3
- 210000000578 peripheral nerve Anatomy 0.000 description 3
- 208000033808 peripheral neuropathy Diseases 0.000 description 3
- 210000003800 pharynx Anatomy 0.000 description 3
- 230000019612 pigmentation Effects 0.000 description 3
- 239000013612 plasmid Substances 0.000 description 3
- 230000002265 prevention Effects 0.000 description 3
- 230000002250 progressing effect Effects 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 239000008213 purified water Substances 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 210000000813 small intestine Anatomy 0.000 description 3
- 210000000278 spinal cord Anatomy 0.000 description 3
- 210000000952 spleen Anatomy 0.000 description 3
- 239000003381 stabilizer Substances 0.000 description 3
- 210000002784 stomach Anatomy 0.000 description 3
- 230000004083 survival effect Effects 0.000 description 3
- 208000024891 symptom Diseases 0.000 description 3
- 238000007910 systemic administration Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 230000001225 therapeutic effect Effects 0.000 description 3
- 238000011200 topical administration Methods 0.000 description 3
- 210000003437 trachea Anatomy 0.000 description 3
- 230000008733 trauma Effects 0.000 description 3
- 208000001072 type 2 diabetes mellitus Diseases 0.000 description 3
- 241001430294 unidentified retrovirus Species 0.000 description 3
- 239000013598 vector Substances 0.000 description 3
- VBICKXHEKHSIBG-UHFFFAOYSA-N 1-monostearoylglycerol Chemical compound CCCCCCCCCCCCCCCCCC(=O)OCC(O)CO VBICKXHEKHSIBG-UHFFFAOYSA-N 0.000 description 2
- 208000024827 Alzheimer disease Diseases 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
- 208000001992 Autosomal Dominant Optic Atrophy Diseases 0.000 description 2
- 206010005176 Blindness congenital Diseases 0.000 description 2
- 208000010392 Bone Fractures Diseases 0.000 description 2
- 102000004219 Brain-derived neurotrophic factor Human genes 0.000 description 2
- 108090000715 Brain-derived neurotrophic factor Proteins 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 2
- 208000016615 Central areolar choroidal dystrophy Diseases 0.000 description 2
- 206010008025 Cerebellar ataxia Diseases 0.000 description 2
- 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 2
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 2
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 2
- 206010012689 Diabetic retinopathy Diseases 0.000 description 2
- 208000037312 Familial drusen Diseases 0.000 description 2
- 206010017076 Fracture Diseases 0.000 description 2
- 208000036893 GUCY2D-related dominant retinopathy Diseases 0.000 description 2
- 208000010412 Glaucoma Diseases 0.000 description 2
- 101001094700 Homo sapiens POU domain, class 5, transcription factor 1 Proteins 0.000 description 2
- 208000023105 Huntington disease Diseases 0.000 description 2
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 description 2
- 229930182816 L-glutamine Natural products 0.000 description 2
- 208000001089 Multiple system atrophy Diseases 0.000 description 2
- 108010025020 Nerve Growth Factor Proteins 0.000 description 2
- 102000015336 Nerve Growth Factor Human genes 0.000 description 2
- 206010056677 Nerve degeneration Diseases 0.000 description 2
- 208000012902 Nervous system disease Diseases 0.000 description 2
- 208000025966 Neurological disease Diseases 0.000 description 2
- 102100035423 POU domain, class 5, transcription factor 1 Human genes 0.000 description 2
- 208000018737 Parkinson disease Diseases 0.000 description 2
- 229930182555 Penicillin Natural products 0.000 description 2
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 2
- 208000002367 Retinal Perforations Diseases 0.000 description 2
- 206010038848 Retinal detachment Diseases 0.000 description 2
- 206010038897 Retinal tear Diseases 0.000 description 2
- 208000007014 Retinitis pigmentosa Diseases 0.000 description 2
- 208000010112 Spinocerebellar Degenerations Diseases 0.000 description 2
- 229920002472 Starch Polymers 0.000 description 2
- 230000002159 abnormal effect Effects 0.000 description 2
- 230000009471 action Effects 0.000 description 2
- 239000000853 adhesive Substances 0.000 description 2
- 230000001070 adhesive effect Effects 0.000 description 2
- 206010064930 age-related macular degeneration Diseases 0.000 description 2
- 239000003708 ampul Substances 0.000 description 2
- 206010002026 amyotrophic lateral sclerosis Diseases 0.000 description 2
- 229940088710 antibiotic agent Drugs 0.000 description 2
- 229960000686 benzalkonium chloride Drugs 0.000 description 2
- CADWTSSKOVRVJC-UHFFFAOYSA-N benzyl(dimethyl)azanium;chloride Chemical compound [Cl-].C[NH+](C)CC1=CC=CC=C1 CADWTSSKOVRVJC-UHFFFAOYSA-N 0.000 description 2
- 230000037182 bone density Effects 0.000 description 2
- 229940098773 bovine serum albumin Drugs 0.000 description 2
- 210000005013 brain tissue Anatomy 0.000 description 2
- 229940077737 brain-derived neurotrophic factor Drugs 0.000 description 2
- 239000002775 capsule Substances 0.000 description 2
- 239000001768 carboxy methyl cellulose Substances 0.000 description 2
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 2
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 2
- 230000006727 cell loss Effects 0.000 description 2
- 239000003638 chemical reducing agent Substances 0.000 description 2
- 229960005188 collagen Drugs 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 230000007850 degeneration Effects 0.000 description 2
- 210000005258 dental pulp stem cell Anatomy 0.000 description 2
- 210000004207 dermis Anatomy 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000006866 deterioration Effects 0.000 description 2
- 239000007884 disintegrant Substances 0.000 description 2
- 239000002270 dispersing agent Substances 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 239000003797 essential amino acid Substances 0.000 description 2
- 235000020776 essential amino acid Nutrition 0.000 description 2
- 238000012869 ethanol precipitation Methods 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 239000008187 granular material Substances 0.000 description 2
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 2
- 210000004263 induced pluripotent stem cell Anatomy 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 230000003907 kidney function Effects 0.000 description 2
- 101150111214 lin-28 gene Proteins 0.000 description 2
- 230000003908 liver function Effects 0.000 description 2
- 208000002780 macular degeneration Diseases 0.000 description 2
- 239000003550 marker Substances 0.000 description 2
- 108010082117 matrigel Proteins 0.000 description 2
- LXCFILQKKLGQFO-UHFFFAOYSA-N methylparaben Chemical compound COC(=O)C1=CC=C(O)C=C1 LXCFILQKKLGQFO-UHFFFAOYSA-N 0.000 description 2
- 229940053128 nerve growth factor Drugs 0.000 description 2
- 108091027963 non-coding RNA Proteins 0.000 description 2
- 102000042567 non-coding RNA Human genes 0.000 description 2
- 210000001328 optic nerve Anatomy 0.000 description 2
- 239000003002 pH adjusting agent Substances 0.000 description 2
- 230000004203 pancreatic function Effects 0.000 description 2
- 229940049954 penicillin Drugs 0.000 description 2
- 239000000546 pharmaceutical excipient Substances 0.000 description 2
- WVDDGKGOMKODPV-ZQBYOMGUSA-N phenyl(114C)methanol Chemical compound O[14CH2]C1=CC=CC=C1 WVDDGKGOMKODPV-ZQBYOMGUSA-N 0.000 description 2
- 239000002243 precursor Substances 0.000 description 2
- 201000002212 progressive supranuclear palsy Diseases 0.000 description 2
- 229950010131 puromycin Drugs 0.000 description 2
- 230000004264 retinal detachment Effects 0.000 description 2
- 230000002207 retinal effect Effects 0.000 description 2
- 230000003248 secreting effect Effects 0.000 description 2
- 210000000697 sensory organ Anatomy 0.000 description 2
- 239000000600 sorbitol Substances 0.000 description 2
- 235000010356 sorbitol Nutrition 0.000 description 2
- 239000008107 starch Substances 0.000 description 2
- 235000019698 starch Nutrition 0.000 description 2
- 229960005322 streptomycin Drugs 0.000 description 2
- 238000004114 suspension culture Methods 0.000 description 2
- 230000009885 systemic effect Effects 0.000 description 2
- 239000002562 thickening agent Substances 0.000 description 2
- 238000002054 transplantation Methods 0.000 description 2
- 230000000472 traumatic effect Effects 0.000 description 2
- 229960000604 valproic acid Drugs 0.000 description 2
- 230000004304 visual acuity Effects 0.000 description 2
- DGVVWUTYPXICAM-UHFFFAOYSA-N β‐Mercaptoethanol Chemical compound OCCS DGVVWUTYPXICAM-UHFFFAOYSA-N 0.000 description 2
- LNAZSHAWQACDHT-XIYTZBAFSA-N (2r,3r,4s,5r,6s)-4,5-dimethoxy-2-(methoxymethyl)-3-[(2s,3r,4s,5r,6r)-3,4,5-trimethoxy-6-(methoxymethyl)oxan-2-yl]oxy-6-[(2r,3r,4s,5r,6r)-4,5,6-trimethoxy-2-(methoxymethyl)oxan-3-yl]oxyoxane Chemical compound CO[C@@H]1[C@@H](OC)[C@H](OC)[C@@H](COC)O[C@H]1O[C@H]1[C@H](OC)[C@@H](OC)[C@H](O[C@H]2[C@@H]([C@@H](OC)[C@H](OC)O[C@@H]2COC)OC)O[C@@H]1COC LNAZSHAWQACDHT-XIYTZBAFSA-N 0.000 description 1
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 description 1
- 244000215068 Acacia senegal Species 0.000 description 1
- QTBSBXVTEAMEQO-UHFFFAOYSA-M Acetate Chemical compound CC([O-])=O QTBSBXVTEAMEQO-UHFFFAOYSA-M 0.000 description 1
- 108700028369 Alleles Proteins 0.000 description 1
- 241000416162 Astragalus gummifer Species 0.000 description 1
- 108060000903 Beta-catenin Proteins 0.000 description 1
- 102000015735 Beta-catenin Human genes 0.000 description 1
- 101100257372 Caenorhabditis elegans sox-3 gene Proteins 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
- 102000029816 Collagenase Human genes 0.000 description 1
- 108060005980 Collagenase Proteins 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- 208000001840 Dandruff Diseases 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 108010014258 Elastin Proteins 0.000 description 1
- 102000016942 Elastin Human genes 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 101150099612 Esrrb gene Proteins 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 108010080379 Fibrin Tissue Adhesive Proteins 0.000 description 1
- 108010049003 Fibrinogen Proteins 0.000 description 1
- 102000008946 Fibrinogen Human genes 0.000 description 1
- 239000004606 Fillers/Extenders Substances 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 206010019668 Hepatic fibrosis Diseases 0.000 description 1
- 102000011787 Histone Methyltransferases Human genes 0.000 description 1
- 108010036115 Histone Methyltransferases Proteins 0.000 description 1
- 101001139134 Homo sapiens Krueppel-like factor 4 Proteins 0.000 description 1
- 101000984042 Homo sapiens Protein lin-28 homolog A Proteins 0.000 description 1
- 101000713275 Homo sapiens Solute carrier family 22 member 3 Proteins 0.000 description 1
- 101000687905 Homo sapiens Transcription factor SOX-2 Proteins 0.000 description 1
- 206010021143 Hypoxia Diseases 0.000 description 1
- 102000011782 Keratins Human genes 0.000 description 1
- 108010076876 Keratins Proteins 0.000 description 1
- 101150072501 Klf2 gene Proteins 0.000 description 1
- 102100020677 Krueppel-like factor 4 Human genes 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 108010085895 Laminin Proteins 0.000 description 1
- 241000713666 Lentivirus Species 0.000 description 1
- 208000019693 Lung disease Diseases 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 101000804949 Mus musculus Developmental pluripotency-associated protein 2 Proteins 0.000 description 1
- 101100446513 Mus musculus Fgf4 gene Proteins 0.000 description 1
- 101100310657 Mus musculus Sox1 gene Proteins 0.000 description 1
- 101100310648 Mus musculus Sox17 gene Proteins 0.000 description 1
- 101100257376 Mus musculus Sox3 gene Proteins 0.000 description 1
- 108700026495 N-Myc Proto-Oncogene Proteins 0.000 description 1
- 102100030124 N-myc proto-oncogene protein Human genes 0.000 description 1
- 101150072008 NR5A2 gene Proteins 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 101710126211 POU domain, class 5, transcription factor 1 Proteins 0.000 description 1
- 241000590428 Panacea Species 0.000 description 1
- 208000016222 Pancreatic disease Diseases 0.000 description 1
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 1
- 206010036790 Productive cough Diseases 0.000 description 1
- 102100025460 Protein lin-28 homolog A Human genes 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 108091027967 Small hairpin RNA Proteins 0.000 description 1
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 1
- 101150001847 Sox15 gene Proteins 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 241000282898 Sus scrofa Species 0.000 description 1
- 210000001744 T-lymphocyte Anatomy 0.000 description 1
- 101150111019 Tbx3 gene Proteins 0.000 description 1
- 229920001615 Tragacanth Polymers 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
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 206010067584 Type 1 diabetes mellitus Diseases 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 238000004115 adherent culture Methods 0.000 description 1
- 210000000577 adipose tissue Anatomy 0.000 description 1
- 230000001919 adrenal effect Effects 0.000 description 1
- 239000003463 adsorbent Substances 0.000 description 1
- 210000001552 airway epithelial cell Anatomy 0.000 description 1
- 229940035676 analgesics Drugs 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000000730 antalgic agent Substances 0.000 description 1
- 239000000058 anti acne agent Substances 0.000 description 1
- 230000000170 anti-cariogenic effect Effects 0.000 description 1
- 230000002421 anti-septic effect Effects 0.000 description 1
- 229940124340 antiacne agent Drugs 0.000 description 1
- 239000000935 antidepressant agent Substances 0.000 description 1
- 239000002518 antifoaming agent Substances 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000006708 antioxidants Nutrition 0.000 description 1
- 239000002216 antistatic agent Substances 0.000 description 1
- 239000011668 ascorbic acid Substances 0.000 description 1
- 235000010323 ascorbic acid Nutrition 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 210000003403 autonomic nervous system Anatomy 0.000 description 1
- 239000003899 bactericide agent Substances 0.000 description 1
- 239000011230 binding agent Substances 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 239000007844 bleaching agent Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000000601 blood cell Anatomy 0.000 description 1
- 230000003925 brain function Effects 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 235000010216 calcium carbonate Nutrition 0.000 description 1
- 239000004091 cariogenic agent Substances 0.000 description 1
- 239000000969 carrier Substances 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 230000024245 cell differentiation Effects 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 210000003169 central nervous system Anatomy 0.000 description 1
- 239000002738 chelating agent Substances 0.000 description 1
- 210000002932 cholinergic neuron Anatomy 0.000 description 1
- 239000003593 chromogenic compound Substances 0.000 description 1
- 210000004081 cilia Anatomy 0.000 description 1
- 239000012459 cleaning agent Substances 0.000 description 1
- GKTWGGQPFAXNFI-HNNXBMFYSA-N clopidogrel Chemical compound C1([C@H](N2CC=3C=CSC=3CC2)C(=O)OC)=CC=CC=C1Cl GKTWGGQPFAXNFI-HNNXBMFYSA-N 0.000 description 1
- 229960002424 collagenase Drugs 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 210000000555 contractile cell Anatomy 0.000 description 1
- 230000007797 corrosion Effects 0.000 description 1
- 238000005260 corrosion Methods 0.000 description 1
- 210000003792 cranial nerve Anatomy 0.000 description 1
- 239000006071 cream Substances 0.000 description 1
- 101150027734 cript gene Proteins 0.000 description 1
- 238000012136 culture method Methods 0.000 description 1
- 230000001934 delay Effects 0.000 description 1
- 239000003398 denaturant Substances 0.000 description 1
- 239000002781 deodorant agent Substances 0.000 description 1
- 238000009795 derivation Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- UGMCXQCYOVCMTB-UHFFFAOYSA-K dihydroxy(stearato)aluminium Chemical compound CCCCCCCCCCCCCCCCCC(=O)O[Al](O)O UGMCXQCYOVCMTB-UHFFFAOYSA-K 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 229920002549 elastin Polymers 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 210000001671 embryonic stem cell Anatomy 0.000 description 1
- 239000003974 emollient agent Substances 0.000 description 1
- 230000001804 emulsifying effect Effects 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 229940088598 enzyme Drugs 0.000 description 1
- 210000001339 epidermal cell Anatomy 0.000 description 1
- 210000003499 exocrine gland Anatomy 0.000 description 1
- 210000002744 extracellular matrix Anatomy 0.000 description 1
- 210000001813 extracellular matrix secreting cell Anatomy 0.000 description 1
- 238000005562 fading Methods 0.000 description 1
- 239000012894 fetal calf serum Substances 0.000 description 1
- 229940012952 fibrinogen Drugs 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 108091006047 fluorescent proteins Proteins 0.000 description 1
- 239000004088 foaming agent Substances 0.000 description 1
- 235000013355 food flavoring agent Nutrition 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 239000003205 fragrance Substances 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 210000004602 germ cell Anatomy 0.000 description 1
- 210000001654 germ layer Anatomy 0.000 description 1
- 229960002743 glutamine Drugs 0.000 description 1
- 229940075507 glyceryl monostearate Drugs 0.000 description 1
- 239000000118 hair dye Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 210000003494 hepatocyte Anatomy 0.000 description 1
- 229940121372 histone deacetylase inhibitor Drugs 0.000 description 1
- 239000003276 histone deacetylase inhibitor Substances 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 229920003063 hydroxymethyl cellulose Polymers 0.000 description 1
- 229940031574 hydroxymethyl cellulose Drugs 0.000 description 1
- 230000001146 hypoxic effect Effects 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 239000002917 insecticide Substances 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 108010038862 laminin 10 Proteins 0.000 description 1
- 239000010410 layer Substances 0.000 description 1
- 208000019423 liver disease Diseases 0.000 description 1
- 210000005228 liver tissue Anatomy 0.000 description 1
- 239000000314 lubricant Substances 0.000 description 1
- 210000004216 mammary stem cell Anatomy 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- ZFLWDHHVRRZMEI-UHFFFAOYSA-N methyl 2,6-dimethyl-5-nitro-4-[2-(trifluoromethyl)phenyl]-1,4-dihydropyridine-3-carboxylate Chemical compound COC(=O)C1=C(C)NC(C)=C([N+]([O-])=O)C1C1=CC=CC=C1C(F)(F)F ZFLWDHHVRRZMEI-UHFFFAOYSA-N 0.000 description 1
- 229920000609 methyl cellulose Polymers 0.000 description 1
- 235000010270 methyl p-hydroxybenzoate Nutrition 0.000 description 1
- 239000004292 methyl p-hydroxybenzoate Substances 0.000 description 1
- 239000001923 methylcellulose Substances 0.000 description 1
- 235000010981 methylcellulose Nutrition 0.000 description 1
- 229960002216 methylparaben Drugs 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000003607 modifier Substances 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- 239000001788 mono and diglycerides of fatty acids Substances 0.000 description 1
- 238000010172 mouse model Methods 0.000 description 1
- FMURUEPQXKJIPS-UHFFFAOYSA-N n-(1-benzylpiperidin-4-yl)-6,7-dimethoxy-2-(4-methyl-1,4-diazepan-1-yl)quinazolin-4-amine;trihydrochloride Chemical compound Cl.Cl.Cl.C=12C=C(OC)C(OC)=CC2=NC(N2CCN(C)CCC2)=NC=1NC(CC1)CCN1CC1=CC=CC=C1 FMURUEPQXKJIPS-UHFFFAOYSA-N 0.000 description 1
- 239000006199 nebulizer Substances 0.000 description 1
- 210000000944 nerve tissue Anatomy 0.000 description 1
- 210000001178 neural stem cell Anatomy 0.000 description 1
- 210000004498 neuroglial cell Anatomy 0.000 description 1
- 239000002674 ointment Substances 0.000 description 1
- 239000003605 opacifier Substances 0.000 description 1
- 239000008375 oral care agent Substances 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 208000024691 pancreas disease Diseases 0.000 description 1
- 210000004923 pancreatic tissue Anatomy 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 210000002856 peripheral neuron Anatomy 0.000 description 1
- 229960003742 phenol Drugs 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
- 230000000704 physical effect Effects 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 210000004694 pigment cell Anatomy 0.000 description 1
- 210000002826 placenta Anatomy 0.000 description 1
- 239000004014 plasticizer Substances 0.000 description 1
- 238000007747 plating Methods 0.000 description 1
- 238000005498 polishing Methods 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 239000003380 propellant Substances 0.000 description 1
- 230000002685 pulmonary effect Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 239000005871 repellent Substances 0.000 description 1
- 230000002940 repellent Effects 0.000 description 1
- 230000000241 respiratory effect Effects 0.000 description 1
- 230000029058 respiratory gaseous exchange Effects 0.000 description 1
- 230000036387 respiratory rate Effects 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 210000000844 retinal pigment epithelial cell Anatomy 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 239000011435 rock Substances 0.000 description 1
- 208000017520 skin disease Diseases 0.000 description 1
- 230000036560 skin regeneration Effects 0.000 description 1
- 239000004055 small Interfering RNA Substances 0.000 description 1
- 150000003384 small molecules Chemical class 0.000 description 1
- 210000000329 smooth muscle myocyte Anatomy 0.000 description 1
- 235000010413 sodium alginate Nutrition 0.000 description 1
- 239000000661 sodium alginate Substances 0.000 description 1
- 229940005550 sodium alginate Drugs 0.000 description 1
- 235000019333 sodium laurylsulphate Nutrition 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 230000002269 spontaneous effect Effects 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 208000024794 sputum Diseases 0.000 description 1
- 210000003802 sputum Anatomy 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 210000000352 storage cell Anatomy 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000002344 surface layer Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 239000000375 suspending agent Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 210000004243 sweat Anatomy 0.000 description 1
- 239000012749 thinning agent Substances 0.000 description 1
- 235000010487 tragacanth Nutrition 0.000 description 1
- 239000000196 tragacanth Substances 0.000 description 1
- 229940116362 tragacanth Drugs 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 239000006097 ultraviolet radiation absorber Substances 0.000 description 1
- 210000003954 umbilical cord Anatomy 0.000 description 1
- 241000701161 unidentified adenovirus Species 0.000 description 1
- VBEQCZHXXJYVRD-GACYYNSASA-N uroanthelone Chemical compound C([C@@H](C(=O)N[C@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CS)C(=O)N[C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C(C)C)[C@@H](C)O)NC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H](CCSC)NC(=O)[C@H](CS)NC(=O)[C@@H](NC(=O)CNC(=O)CNC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CS)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CS)NC(=O)CNC(=O)[C@H]1N(CCC1)C(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CC(N)=O)C(C)C)[C@@H](C)CC)C1=CC=C(O)C=C1 VBEQCZHXXJYVRD-GACYYNSASA-N 0.000 description 1
- 210000003556 vascular endothelial cell Anatomy 0.000 description 1
- 108700026220 vif Genes Proteins 0.000 description 1
- 230000031836 visual learning Effects 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000000080 wetting agent Substances 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/98—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/48—Reproductive organs
- A61K35/54—Ovaries; Ova; Ovules; Embryos; Foetal cells; Germ cells
- A61K35/545—Embryonic stem cells; Pluripotent stem cells; Induced pluripotent stem cells; Uncharacterised stem cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Cell Biology (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Zoology (AREA)
- Immunology (AREA)
- Epidemiology (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Pharmacology & Pharmacy (AREA)
- Reproductive Health (AREA)
- Dermatology (AREA)
- Hematology (AREA)
- Gynecology & Obstetrics (AREA)
- Birds (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Cosmetics (AREA)
Abstract
Description
本発明は、iPS細胞培養上清を含む医薬組成物、およびその製造方法、化粧品およびその製造方法、抗加齢組成物、疾患発症抑制方法、疾患治療方法、組織異常治療方法ならびに美容方法、に関する。 The present invention relates to a pharmaceutical composition containing an iPS cell culture supernatant, a method for producing the same, a cosmetic and a method for producing the same, an anti-aging composition, a disease onset inhibiting method, a disease treating method, a tissue abnormality treating method and a cosmetic method. .
従来の医療によっては治療困難な疾病に対する代替技術として、幹細胞を利用した再生医療が注目されている。
幹細胞を用いた再生医療は、全ての難病にとっての新しい臨床プラットフォームにおける有望なツールである。胚性幹細胞(ES細胞)および体性幹細胞を初めとする種々の幹細胞が報告されている。体性幹細胞のうち、骨髄、脂肪組織、皮膚、臍帯、胎盤等の種々の組織から単離される間葉系幹細胞(MSC)が皮膚再生における臨床的応用に特に用いられてきた。
Regenerative medicine using stem cells has attracted attention as an alternative technique for diseases that are difficult to treat depending on conventional medicine.
Regenerative medicine using stem cells is a promising tool in a new clinical platform for all intractable diseases. Various stem cells including embryonic stem cells (ES cells) and somatic stem cells have been reported. Among somatic stem cells, mesenchymal stem cells (MSC) isolated from various tissues such as bone marrow, adipose tissue, skin, umbilical cord, and placenta have been particularly used for clinical application in skin regeneration.
また、人工多能性幹細胞(induced pluripotent stem cells;iPS細胞)は、体細胞に数種類の初期化因子を導入することによって作成された細胞であり、三胚葉系列すべてに分化できる「分化多能性」および未分化状態を保持したまま増殖できる「自己複製性」を有している(例えば、特許第5098028号明細書参照)。この分化多能性を利用して、iPS細胞を分化させて移植材料を得ようとする試みも行われている(例えば、国際公開公報WO2011/136378号参照)。 In addition, induced pluripotent stem cells (iPS cells) are cells created by introducing several types of reprogramming factors into somatic cells, and can be differentiated into all three germ layers. And “self-replicating” capable of growing while maintaining an undifferentiated state (see, for example, Japanese Patent No. 5098028). Attempts have also been made to differentiate iPS cells and obtain transplant materials using this differentiation pluripotency (see, for example, International Publication No. WO2011-136378).
一方、幹細胞を培養して得られる幹細胞培養上清を標的組織の損傷部を修復するために用いる試みもなされている(例えば、国際公開公報WO2011/118795号参照)。 On the other hand, an attempt has been made to use a stem cell culture supernatant obtained by culturing stem cells to repair a damaged portion of a target tissue (see, for example, International Publication No. WO2011 / 118795).
iPS細胞を再生医療に用いようとする試みは、iPS細胞自体の分化多能性や自己複製性を利用するものであった。この場合、例えば、iPS細胞を分化させて得られた移植材料の移植が必要であり、患者に対する負担がかかる。さらに、拒絶を避けようとすれば患者自身の体細胞からiPS細胞をいちいち作製することが必要であり、治療の準備に要する時間や労力は比較的大きいものとなる。
一方、歯髄幹細胞などの、体内に存在する体性幹細胞を培養して得られた培養上清は、損傷修復において一定の効果を示すものの、適用対象となる組織の種類によってその有効性は変動し、体内の組織一般に対する普遍的な有効性を有するものとは言い難かった。
本発明は、上記の状況に鑑みてなされたものであり、iPS細胞の培養上清を利用した新たな医薬組成物およびその製造方法、化粧品およびその製造方法、抗加齢組成物、疾患発症抑制方法、疾患治療方法、組織異常治療方法ならびに美容方法を提供することを課題とする。
Attempts to use iPS cells for regenerative medicine have made use of the pluripotency and self-replication of iPS cells themselves. In this case, for example, transplantation of a transplant material obtained by differentiating iPS cells is necessary, which places a burden on the patient. Furthermore, if it is intended to avoid rejection, it is necessary to prepare iPS cells from the patient's own somatic cells one by one, and the time and labor required for preparation for treatment become relatively large.
On the other hand, culture supernatants obtained by culturing somatic stem cells existing in the body, such as dental pulp stem cells, exhibit a certain effect in damage repair, but their effectiveness varies depending on the type of tissue to which they are applied. It was difficult to say that it has universal effectiveness for the body tissues in general.
The present invention has been made in view of the above situation, and a new pharmaceutical composition using the culture supernatant of iPS cells and a method for producing the same, a cosmetic and a method for producing the same, an anti-aging composition, and a disease onset suppression It is an object to provide a method, a disease treatment method, a tissue abnormality treatment method, and a beauty method.
本発明の第1の態様に係る医薬組成物は、iPS細胞を培養することによって得られたiPS細胞培養上清を含む、医薬組成物である。
本発明の第2の態様に係る医薬組成物の製造方法は、
(1)iPS細胞を培養するステップ;および
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、医薬組成物の製造方法である。
本発明の第3の態様に係る使用は、iPS細胞を培養することによって得られたiPS細胞培養上清の、医薬組成物の製造における使用である。
The pharmaceutical composition according to the first aspect of the present invention is a pharmaceutical composition comprising an iPS cell culture supernatant obtained by culturing iPS cells.
The method for producing a pharmaceutical composition according to the second aspect of the present invention comprises:
(1) culturing iPS cells; and (2) collecting the culture supernatant obtained by culturing the iPS cells;
Is a method for producing a pharmaceutical composition.
The use according to the third aspect of the present invention is the use of an iPS cell culture supernatant obtained by culturing iPS cells in the production of a pharmaceutical composition.
本発明の第4の態様に係る方法は、前記第1の態様に係る医薬組成物を、疾患または組織異常の発症前の対象に、前記疾患または組織異常の発症を抑えるために有効な量投与することを含む、対象において疾患または組織異常の発症前に前記疾患または組織異常の発症を抑える方法である。
本発明の第5の態様に係る方法は、前記第1の態様に係る医薬組成物を、疾患を有する対象に、前記疾患を治療するために有効な量で投与することを含む、疾患の治療方法である。
本発明の第6の態様に係る方法は、前記第1の態様に係る医薬組成物を、組織異常を有する対象に、前記組織異常を治療するために有効な量で投与することを含む、組織異常の治療方法である。
本発明の第7の態様に係る化粧品は、iPS細胞を培養することによって得られたiPS細胞培養上清を含む、化粧品である。
In the method according to the fourth aspect of the present invention, the pharmaceutical composition according to the first aspect is administered to a subject before the onset of a disease or tissue abnormality in an amount effective for suppressing the onset of the disease or tissue abnormality. A method of suppressing the onset of the disease or tissue abnormality before the onset of the disease or tissue abnormality in the subject.
The method according to the fifth aspect of the present invention comprises administering the pharmaceutical composition according to the first aspect to a subject having a disease in an amount effective to treat the disease. Is the method.
The method according to the sixth aspect of the present invention comprises administering the pharmaceutical composition according to the first aspect to a subject having a tissue abnormality in an amount effective for treating the tissue abnormality. This is a method for treating abnormalities.
The cosmetic according to the seventh aspect of the present invention is a cosmetic containing an iPS cell culture supernatant obtained by culturing iPS cells.
本発明の第8の態様に係る美容方法は、前記第7の態様に係る化粧品を対象に適用することにより、対象の美容的外観を改善する、美容方法である。
本発明の第9の態様に係る化粧品の製造方法は、
(1)iPS細胞を培養するステップ;および
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、化粧品の製造方法である。
A beauty method according to an eighth aspect of the present invention is a beauty method for improving the cosmetic appearance of a subject by applying the cosmetic according to the seventh aspect to the subject.
The method for producing a cosmetic according to the ninth aspect of the present invention comprises:
(1) culturing iPS cells; and (2) collecting the culture supernatant obtained by culturing the iPS cells;
A method for producing cosmetics.
本発明によれば、iPS細胞の培養上清を利用した新たな医薬組成物およびその製造方法、化粧品およびその製造方法、抗加齢組成物、疾患発症抑制方法、疾患治療方法、組織異常治療方法ならびに美容方法が提供される。 According to the present invention, a new pharmaceutical composition using the culture supernatant of iPS cells and a production method thereof, a cosmetic and a production method thereof, an anti-aging composition, a disease onset suppression method, a disease treatment method, and a tissue abnormality treatment method As well as a cosmetic method.
<医薬組成物>
本開示に係る医薬組成物は、iPS細胞を培養することによって得られたiPS細胞培養上清を含む。
iPS細胞は分化多能性および自己複製性を有する。先に述べたとおり、iPS細胞を分化させ、得られた分化組織を移植片として用いる試みはあった。しかしながら、iPS細胞から分泌される物質に着目して、iPS細胞の培養上清を用いて例えば、医学的あるいは美容上の効果を得ようとする試みは無かった。それどころか、iPS細胞の培養上清がどのような生物学的な効果を有するのかについて、何ら知られてはいなかった。
<Pharmaceutical composition>
The pharmaceutical composition according to the present disclosure includes an iPS cell culture supernatant obtained by culturing iPS cells.
iPS cells have pluripotency and self-renewal. As described above, there has been an attempt to differentiate iPS cells and use the obtained differentiated tissue as a graft. However, focusing on substances secreted from iPS cells, there has been no attempt to obtain, for example, medical or cosmetic effects using the culture supernatant of iPS cells. On the contrary, nothing was known about what biological effect the culture supernatant of iPS cells had.
本発明の発明者は、iPS細胞培養上清を、医学的、美容的等の効果を得るために用いるという、前例の無い試みを初めて行った。その結果、驚くべきことに、iPS細胞培養上清は、組織一般に対して広く修復能力を有し、またその修復能力は間葉系幹細胞や造血幹細胞などの体性幹細胞の培養上清が示す修復能力と比べて傑出したものであることを、本発明者は発見した。これは、体性幹細胞から分泌される分泌物(成長因子、サイトカイン等を含む)と比較した場合、分化多能性を有するiPS細胞から分泌される分泌物はその物質組成が大きく異なっており、その結果として、どの組織であるかを問わずに、広く一般の組織に対して修復能力を示し、またその修復能力の程度も高いものとなっているものと推定される。具体的には、多くの成長因子、サイトカインについて、iPS細胞の培養上清における含有量は、体性幹細胞の培養上清における含有量に比べて高く、あるいは、特定の成長因子、サイトカインは、iPS細胞の培養上清には含まれるが、体性幹細胞の培養上清には実質的に含まれないと考えられる。また、iPS細胞の培養上清は、体性幹細胞の培養上清には含まれないような抗加齢物質(例えば特定の種類のタンパク質)を含んでおり、この結果として、iPS細胞の培養上清は、体性幹細胞の培養上清では見られないような優れた抗加齢効果を有すると考えられる。
ただし、本発明は、前記推定に何ら拘束されるものではない。
なお、本開示において「修復」とは、組織異常あるいは疾患によって失われた組織の機能の一部又は全部が、当該組織異常または疾患の生じる前における当該組織の機能と比較して維持又は回復していることを意味し、組織の機能が回復することのみならず、機能的な組織として再生することも広く包含する。機能が維持又は回復していることの評価については、組織異常あるいは疾患が生じている組織において異なるが、外観、対象となる機能の程度を評価するために通常用いられるアッセイ等に基づいて行えばよい。
The inventor of the present invention has made an unprecedented attempt to use the iPS cell culture supernatant to obtain medical and cosmetic effects. As a result, surprisingly, iPS cell culture supernatants have a wide repair ability for tissues in general, and the repair ability is shown by the somatic stem cell culture supernatants such as mesenchymal stem cells and hematopoietic stem cells. The inventor has discovered that it is outstanding compared to ability. This is because the secretory product secreted from somatic stem cells (including growth factors, cytokines, etc.) secreted from iPS cells with pluripotency has a greatly different composition. As a result, it is presumed that, regardless of the tissue, the repair ability is widely shown to general tissues, and the degree of the repair ability is high. Specifically, for many growth factors and cytokines, the content of iPS cells in the culture supernatant is higher than the content of somatic stem cells in the culture supernatant, or the specific growth factors and cytokines are iPS cells. It is considered to be contained in the cell culture supernatant but not substantially contained in the somatic stem cell culture supernatant. In addition, the culture supernatant of iPS cells contains an anti-aging substance (for example, a specific type of protein) that is not included in the culture supernatant of somatic stem cells. Kiyoshi is considered to have an excellent anti-aging effect not seen in the culture supernatant of somatic stem cells.
However, the present invention is not restricted by the estimation.
In the present disclosure, the term “repair” means that a part or all of the tissue function lost due to the tissue abnormality or disease is maintained or recovered as compared with the function of the tissue before the tissue abnormality or disease occurs. This means not only that the function of the tissue is restored, but also that it is regenerated as a functional tissue. The evaluation of whether the function is maintained or restored differs depending on the tissue in which the tissue abnormality or disease is occurring, but if it is performed based on the appearance, the assay usually used for evaluating the degree of the target function, etc. Good.
本開示に係る医薬組成物においては、iPS細胞を培養して得られたiPS細胞培養上清を医薬組成物の有効成分として用いる。当該iPS細胞培養上清には、サイトカイン混合物等が含まれているため、種々の組織において修復能力を示す。本発明の一形態では、iPS細胞培養上清中のサイトカインの混合物が組織の内在性幹細胞に対する誘導シグナルとして作用することにより、前記内在性幹細胞が、分化し、増殖し得、これが修復能力の発現をもたらし得ると推定できる。ただし、本発明は、前記推定に何ら拘束されるものではない。 In the pharmaceutical composition according to the present disclosure, the iPS cell culture supernatant obtained by culturing iPS cells is used as an active ingredient of the pharmaceutical composition. Since the iPS cell culture supernatant contains a cytokine mixture and the like, it exhibits repair ability in various tissues. In one form of the present invention, the mixture of cytokines in the iPS cell culture supernatant acts as an induction signal for the tissue's endogenous stem cells, so that the endogenous stem cells can differentiate and proliferate, which expresses the repair ability. It can be estimated that However, the present invention is not restricted by the estimation.
本開示におけるiPS細胞培養上清は、iPS細胞を培養して得られるものであるため、以下の説明ではその製法も併せて説明する。 Since the iPS cell culture supernatant in the present disclosure is obtained by culturing iPS cells, the production method thereof will also be described in the following description.
人工多能性幹細胞(iPS細胞)は、初期化因子を、体細胞に導入することによって作製された、ES細胞とほぼ同等の特性、例えば分化多能性と自己複製による増殖能、を有する体細胞由来の人工の幹細胞である(K. Takahashi and S. Yamanaka (2006) Cell, 126:663−676; K. Takahashi et al. (2007), Cell, 131:861−872; J. Yu et al. (2007), Science, 318:1917−1920; Nakagawa, M.ら,Nat. Biotechnol. 26:101−106 (2008);国際公開WO 2007/069666)。初期化因子は、例えば、DNA又はタンパク質の形態で体細胞に導入することが出来る。初期化因子は、ES細胞に特異的に発現している遺伝子、その遺伝子産物もしくはノンコーディングRNAまたはES細胞の未分化維持に重要な役割を果たす遺伝子、その遺伝子産物もしくはノンコーディングRNA、あるいは低分子化合物によって構成されてもよい。初期化因子に含まれる遺伝子としては、例えば、特開2013−247943に挙げられているように、Oct3/4、Sox2、Sox1、Sox3、Sox15、Sox17、Klf4、Klf2、c−Myc、N−Myc、L−Myc、Nanog、Lin28、Fbx15、ERas、ECAT15−2、Tcl1、beta−catenin、Lin28b、Sall1、Sall4、Esrrb、Nr5a2、Tbx3またはGlis1等が挙げられる。
iPS細胞は、上記の方法の他にも、これまでに報告された各種iPS細胞作製法によって作製することができる。また、今後開発されるiPS細胞作製法を適用することも当然に想定される。
Artificial pluripotent stem cells (iPS cells), which are produced by introducing reprogramming factors into somatic cells, have almost the same characteristics as ES cells, such as differentiation pluripotency and proliferation ability by self-replication. Cell-derived artificial stem cells (K. Takahashi and S. Yamanaka (2006) Cell, 126: 663-676; K. Takahashi et al. (2007), Cell, 131: 861-872; J. Yu et al. (2007), Science, 318: 1917-1920; Nakagawa, M. et al., Nat. Biotechnol. 26: 101-106 (2008); International Publication WO 2007/069666). The reprogramming factor can be introduced into somatic cells, for example, in the form of DNA or protein. The reprogramming factor is a gene that is specifically expressed in ES cells, its gene product or noncoding RNA, a gene that plays an important role in maintaining undifferentiation of ES cells, its gene product or noncoding RNA, or a small molecule It may be constituted by a compound. Examples of genes included in the reprogramming factor include Oct3 / 4, Sox2, Sox1, Sox3, Sox15, Sox17, Klf4, Klf2, c-Myc, N-Myc, as described in JP2013-247934A, for example. , L-Myc, Nanog, Lin28, Fbx15, ERas, ECAT15-2, Tcl1, beta-catenin, Lin28b, Sall1, Sall4, Esrrb, Nr5a2, Tbx3 or Glis1.
In addition to the above method, iPS cells can be prepared by various iPS cell preparation methods reported so far. It is naturally assumed that an iPS cell production method developed in the future will be applied.
iPS細胞作製法の最も基本的な手法は、転写因子であるOct3/4、Sox2、Klf4及びc-Mycの4因子を、ウイルスを利用して細胞へ導入する方法である(Takahashi K, Yamanaka S: Cell 126 (4), 663-676, 2006; Takahashi, K, et al: Cell 131 (5), 861-72, 2007)。ヒトiPS細胞についてはOct4、Sox2、Lin28及びNonogの4因子の導入による樹立の報告がある(Yu J, et al: Science 318(5858), 1917-1920, 2007)。c-Mycを除く3因子(Nakagawa M, et al: Nat. Biotechnol. 26 (1), 101-106, 2008)、Oct3/4及びKlf4の2因子(Kim J B, et al: Nature 454 (7204), 646-650, 2008)、或いはOct3/4のみ(Kim J B, et al: Cell 136 (3), 411-419, 2009)の導入によるiPS細胞の樹立も報告されている。また、遺伝子の発現産物であるタンパク質を細胞に導入する手法(Zhou H, Wu S, Joo JY, et al: Cell Stem Cell 4, 381-384, 2009; Kim D, Kim CH, Moon JI, et al: Cell Stem Cell 4, 472-476, 2009)も報告されている。一方、ヒストンメチル基転移酵素G9aに対する阻害剤BIX-01294やヒストン脱アセチル化酵素阻害剤バルプロ酸(VPA)或いはBayK8644等を使用することによって作製効率の向上や導入する因子の低減などが可能であるとの報告もある(Huangfu D, et al: Nat. Biotechnol. 26 (7), 795-797, 2008; Huangfu D, et al: Nat. Biotechnol. 26 (11), 1269-1275, 2008; Silva J, et al: PLoS. Biol. 6 (10), e 253, 2008)。遺伝子導入法についても検討が進められ、レトロウイルスの他、レンチウイルス(Yu J, et al: Science 318(5858), 1917-1920, 2007)、アデノウイルス(Stadtfeld M, et al: Science 322 (5903), 945-949, 2008)、プラスミド(Okita K, et al: Science 322 (5903), 949-953, 2008;Okita K. et al.: Nat Methods 8, 409-412)、トランスポゾンベクター(Woltjen K, Michael IP, Mohseni P, et al: Nature 458, 766-770, 2009; Kaji K, Norrby K, Pac a A, et al: Nature 458, 771-775, 2009; Yusa K, Rad R, Takeda J, et al: Nat Methods 6, 363-369, 2009)、或いはエピソーマルベクター(Yu J, Hu K, Smuga-Otto K, Tian S, et al: Science 324, 797-801, 2009)を遺伝子導入に利用した技術が開発されている。 The most basic method for producing iPS cells is to introduce four factors, transcription factors Oct3 / 4, Sox2, Klf4 and c-Myc, into cells using viruses (Takahashi K, Yamanaka S : Cell 126 (4), 663-676, 2006; Takahashi, K, et al: Cell 131 (5), 861-72, 2007). Human iPS cells have been reported to be established by introducing four factors, Oct4, Sox2, Lin28 and Nonog (Yu J, et al: Science 318 (5858), 1917-1920, 2007). 3 factors except c-Myc (Nakagawa M, et al: Nat. Biotechnol. 26 (1), 101-106, 2008), Oct3 / 4 and Klf4 (Kim JB, et al: Nature 454 (7204) , 646-650, 2008), or the establishment of iPS cells by introducing only Oct3 / 4 (Kim JB, et al: Cell 136 (3), 411-419, 2009) has been reported. In addition, a method for introducing a protein, which is an expression product of a gene, into a cell (Zhou H, Wu S, Joo JY, et al: Cell Stem Cell 4, 381-384, 2009; Kim D, Kim CH, Moon JI, et al : Cell Stem Cell 4, 472-476, 2009). On the other hand, by using the inhibitor BIX-01294 for histone methyltransferase G9a, the histone deacetylase inhibitor valproic acid (VPA) or BayK8644, production efficiency can be improved and factors to be introduced can be reduced. (Huangfu D, et al: Nat. Biotechnol. 26 (7), 795-797, 2008; Huangfu D, et al: Nat. Biotechnol. 26 (11), 1269-1275, 2008; Silva J , et al: PLoS. Biol. 6 (10), e 253, 2008). Studies on gene transfer methods are also underway, and in addition to retroviruses, lentiviruses (Yu J, et al: Science 318 (5858), 1917-1920, 2007), adenoviruses (Stadtfeld M, et al: Science 322 (5903 ), 945-949, 2008), plasmid (Okita K, et al: Science 322 (5903), 949-953, 2008; Okita K. et al .: Nat Methods 8, 409-412), transposon vector (Woltjen K , Michael IP, Mohseni P, et al: Nature 458, 766-770, 2009; Kaji K, Norrby K, Pac a A, et al: Nature 458, 771-775, 2009; Yusa K, Rad R, Takeda J, et al: Nat Methods 6, 363-369, 2009) or episomal vectors (Yu J, Hu K, Smuga-Otto K, Tian S, et al: Science 324, 797-801, 2009) Technology has been developed.
iPS細胞作製に際して、初期化因子を導入する対象となる体細胞は、哺乳動物(例えば、ヒト、マウス、サル、ブタ、ラット等)由来の生殖細胞以外のいかなる細胞であってもよく、例えば、特開2013−247943に挙げられているように、角質化する上皮細胞(例、角質化表皮細胞)、粘膜上皮細胞(例、舌表層の上皮細胞)、外分泌腺上皮細胞(例、乳腺細胞)、ホルモン分泌細胞(例、副腎髄質細胞)、代謝・貯蔵用の細胞(例、肝細胞)、境界面を構成する内腔上皮細胞(例、I型肺胞細胞)、内鎖管の内腔上皮細胞(例、血管内皮細胞)、運搬能をもつ繊毛のある細胞(例、気道上皮細胞)、細胞外マトリックス分泌用細胞(例、線維芽細胞)、収縮性細胞(例、平滑筋細胞)、血液系細胞、免疫系細胞(例、Tリンパ球)、感覚器官の細胞(例、桿細胞)、自律神経系ニューロン(例、コリン作動性ニューロン)、感覚器と末梢ニューロンの支持細胞(例、随伴細胞)、中枢神経系の神経細胞とグリア細胞(例、星状グリア細胞)、色素細胞(例、網膜色素上皮細胞)、およびそれらの前駆細胞(組織前駆細胞)等が挙げられる。細胞の分化の程度や細胞を採取する動物の齢などに特に制限はなく、未分化な前駆細胞(体性幹細胞も含む)であっても、最終分化した成熟細胞であっても、同様に、本開示において体細胞の起源として使用することができる。ここで未分化な前駆細胞としては、たとえば神経幹細胞、造血幹細胞、間葉系幹細胞、歯髄幹細胞等の組織幹細胞(体性幹細胞)が挙げられる。中でも、安定性の点からは骨髄由来間葉系幹細胞が好ましい。また、修復能力の点からは、体細胞はヒト由来のものであることが好ましい。また、前記体細胞として、前記医薬組成物の投与対象となる対象自身から得た体細胞を用いることが、拒絶の回避の上で有利な場合がある。 The somatic cell into which the reprogramming factor is introduced in the preparation of the iPS cell may be any cell other than a germ cell derived from a mammal (eg, human, mouse, monkey, pig, rat, etc.), for example, As described in JP2013-247934A, keratinized epithelial cells (eg, keratinized epidermal cells), mucosal epithelial cells (eg, epithelial cells of the tongue surface layer), exocrine gland epithelial cells (eg, mammary cells) , Hormone-secreting cells (eg, adrenal medullary cells), metabolism / storage cells (eg, hepatocytes), luminal epithelial cells that make up the interface (eg, type I alveolar cells), lumens of inner chain vessels Epithelial cells (eg, vascular endothelial cells), cilia cells with transport ability (eg, airway epithelial cells), extracellular matrix secreting cells (eg, fibroblasts), contractile cells (eg, smooth muscle cells) , Blood cells, immune system cells (eg, T lymphocytes) Sensory organ cells (eg, sputum cells), autonomic nervous system neurons (eg, cholinergic neurons), sensory organs and peripheral neuron support cells (eg, associated cells), central nervous system neurons and glial cells (eg, cells) , Astroglial cells), pigment cells (eg, retinal pigment epithelial cells), and their precursor cells (tissue precursor cells). There is no particular limitation on the degree of cell differentiation and the age of the animal from which the cells are collected, whether it is undifferentiated progenitor cells (including somatic stem cells) or terminally differentiated mature cells, It can be used as a source of somatic cells in the present disclosure. Examples of undifferentiated progenitor cells include tissue stem cells (somatic stem cells) such as neural stem cells, hematopoietic stem cells, mesenchymal stem cells, and dental pulp stem cells. Among these, bone marrow-derived mesenchymal stem cells are preferable from the viewpoint of stability. From the viewpoint of repair ability, the somatic cell is preferably derived from a human. In addition, it may be advantageous for avoiding rejection to use somatic cells obtained from the subject to whom the pharmaceutical composition is administered as the somatic cells.
iPS細胞誘導のための培養液としては、特開2013−247943に挙げられているように、例えば、10〜15% FBSを含有するDMEM、DMEM/F12又はDME培養液(これらの培養液にはさらに、LIF、penicillin/streptomycin、puromycin、L-グルタミン、非必須アミノ酸類、β-メルカプトエタノールなどを適宜含むことができる。)または市販の培養液[例えば、マウスES細胞培養用培養液(TX-WES培養液、トロンボX社)、霊長類ES細胞培養用培養液(霊長類ES/iPS細胞用培養液、リプロセル社)、無血清培地(mTeSR、Stemcell Technology社)]などが含まれる。 As a culture solution for iPS cell induction, as disclosed in JP2013-247934A, for example, DMEM, DMEM / F12 or DME culture solution containing 10 to 15% FBS (in these culture solutions) Further, LIF, penicillin / streptomycin, puromycin, L-glutamine, non-essential amino acids, β-mercaptoethanol, etc. may be included as appropriate.) Or a commercially available culture solution [for example, a culture solution for mouse ES cell culture (TX- WES culture medium, Thrombo X), culture medium for primate ES cell culture (culture medium for primate ES / iPS cells, Reprocell), serum-free medium (mTeSR, Stemcell Technology)] and the like.
培養法の例としては、たとえば、特開2013−247943に挙げられているように、37℃、5% CO2存在下にて、10% FBS含有DMEM又はDMEM/F12培養液上で体細胞と初期化因子とを接触させ約4〜7日間培養し、その後、細胞をフィーダー細胞(たとえば、マイトマイシンC処理STO細胞、SNL細胞等)上に播種し直し、体細胞と初期化因子の接触から約10日後からbFGF含有霊長類ES細胞培養用培養液で培養し、該接触から約30〜約45日又はそれ以上後にiPS様コロニーを生じさせることができる。 As an example of the culture method, for example, as described in JP2013-247934A, in the presence of 5% CO 2 at 37 ° C. in the presence of 10% FBS-containing DMEM or DMEM / F12 culture solution, Contact with reprogramming factor and culture for about 4-7 days, then re-seed cells on feeder cells (e.g., mitomycin C-treated STO cells, SNL cells, etc.) After 10 days, the cells can be cultured in a bFGF-containing primate ES cell culture medium, and iPS-like colonies can be generated about 30 to about 45 days or more after the contact.
あるいは、37℃、5%CO2存在下にて、フィーダー細胞(たとえば、マイトマイシンC処理STO細胞、SNL細胞等)上で10% FBS含有DMEM培養液(これにはさらに、LIF、ペニシリン/ストレプトマイシン、ピューロマイシン、L-グルタミン、非必須アミノ酸類、β-メルカプトエタノールなどを適宜含むことができる。)で培養し、約25〜約30日又はそれ以上の後にES様コロニーを生じさせることができる。望ましくは、フィーダー細胞の代わりに、初期化される体細胞そのものを用いる(Takahashi K, et al. (2009), PLoS One. 4:e8067またはWO2010/137746)、もしくは細胞外基質(例えば、Laminin(WO2009/123349)およびマトリゲル(BD社))を用いる方法が例示される。 Alternatively, 10% FBS-containing DMEM culture medium (including LIF, penicillin / streptomycin, etc.) on feeder cells (eg, mitomycin C-treated STO cells, SNL cells, etc.) in the presence of 5% CO 2 at 37 ° C. And can be appropriately added with puromycin, L-glutamine, non-essential amino acids, β-mercaptoethanol, etc.), and ES-like colonies can be formed after about 25 to about 30 days or more. Desirably, instead of feeder cells, somatic cells to be initialized themselves are used (Takahashi K, et al. (2009), PLoS One. 4: e8067 or WO2010 / 137746), or an extracellular matrix (for example, Laminin ( WO2009 / 123349) and Matrigel (BD)) are exemplified.
この他にも、血清を含有しない培地を用いて培養する方法も例示される(Sun N, et al. (2009), Proc Natl Acad Sci U S A. 106:15720-15725)。さらに、樹立効率を上げるため、低酸素条件(0.1%以上、15%以下の酸素濃度)によりiPS細胞を樹立しても良い(Yoshida Y, et al. (2009), Cell Stem Cell. 5:237-241またはWO2010/013845)。
上記培養の間には、培養開始2日目以降から毎日1回新鮮な培養液と培養液交換を行う。また、核初期化に使用する体細胞の細胞数は、限定されないが、培養ディッシュ100cm2当たり約5×103〜約5×106細胞の範囲である。
In addition, a method of culturing using a medium not containing serum is also exemplified (Sun N, et al. (2009), Proc Natl Acad Sci US A. 106: 15720-15725). Furthermore, in order to increase establishment efficiency, iPS cells may be established under hypoxic conditions (oxygen concentration of 0.1% or more and 15% or less) (Yoshida Y, et al. (2009), Cell Stem Cell. 5: 237 -241 or WO2010 / 013845).
During the culture, the culture medium is exchanged with a fresh culture medium once a day from the second day onward. The number of somatic cells used for nuclear reprogramming is not limited, but ranges from about 5 × 10 3 to about 5 × 10 6 cells per 100 cm 2 of culture dish.
iPS細胞は、特開2013−247943に記載されているように、形成したコロニーの形状により選択することが可能である。また、Fbxo15、Nanog、Oct/4、Fgf-4、Esg-1及びCript等の多能性幹細胞マーカー(未分化マーカー)の発現などを指標として選択することができる。一方、体細胞が初期化された場合に発現する遺伝子(例えば、Oct3/4、Nanog)と連動して発現する薬剤耐性遺伝子をマーカー遺伝子として導入した場合は、対応する薬剤を含む培養液(選択培養液)で培養を行うことにより樹立したiPS細胞を選択することができる。また、マーカー遺伝子が蛍光タンパク質遺伝子の場合は蛍光顕微鏡で観察することによって、発光酵素遺伝子の場合は発光基質を加えることによって、また発色酵素遺伝子の場合は発色基質を加えることによって、iPS細胞を選択することができる。選択された細胞をiPS細胞として回収する。 iPS cells can be selected according to the shape of the formed colonies as described in JP2013-247934A. In addition, expression of pluripotent stem cell markers (undifferentiation markers) such as Fbxo15, Nanog, Oct / 4, Fgf-4, Esg-1 and Cript can be selected as an index. On the other hand, when a drug resistance gene that is expressed in conjunction with a gene that is expressed when somatic cells are initialized (for example, Oct3 / 4, Nanog) is introduced as a marker gene, a culture solution containing the corresponding drug (selection The established iPS cells can be selected by culturing with the culture medium. When the marker gene is a fluorescent protein gene, iPS cells are selected by observing with a fluorescence microscope, in the case of a luminescent enzyme gene, by adding a luminescent substrate, and in the case of a chromogenic enzyme gene, by adding a chromogenic substrate. can do. Selected cells are collected as iPS cells.
得られたiPS細胞から培養上清を得るには、iPS細胞の培養条件として通常用いられる条件をそのまま適用することができる。例えば、iPS細胞を作製する際の培地および培養条件としては、上記説明で記載された培地および培養条件が挙げられる。なお、iPS細胞を維持する上でフィーダー細胞の使用は必須ではなく、例えばラミニン511を用いることでフィーダー細胞を省略することも可能である(A novel efficient feeder-free culture system for the derivation of human induced pluripotent stem cells
Masato Nakagawa, et al. Scientific Reports 4,Article number:3594doi:10.1038/srep03594 Received 09 October 2013 Accepted 06 December 2013 Published 08 January 2014 (Nature))。また、培養の際には細胞死を抑制するためにROCK阻害剤を添加してもよい。
In order to obtain a culture supernatant from the obtained iPS cells, conditions usually used as culture conditions for iPS cells can be applied as they are. For example, the culture medium and culture conditions for producing iPS cells include the culture medium and culture conditions described in the above description. The use of feeder cells is not essential for maintaining iPS cells. For example, feeder cells can be omitted by using laminin 511 (A novel efficient feeder-free culture system for the derivation of human induced). pluripotent stem cells
Masato Nakagawa, et al. Scientific Reports 4, Article number: 3594doi: 10.1038 / srep03594 Received 09 October 2013 Accepted 06 December 2013 Published 08 January 2014 (Nature)). Moreover, you may add a ROCK inhibitor in order to suppress a cell death in the case of culture | cultivation.
本開示に係る医薬組成物は、血清を含まないものであってもよい。血清を含まないことで安全性が高められる場合がある。例えば、血清を含まない培地(無血清培地)でiPS細胞を培養することによって、血清を含まない培養上清を調製することができる。1回又は複数回の継代培養を行うことにし、最後又は最後から数回の継代培養を無血清培地で培養することによっても、血清を含まない培養上清を得ることができる。一方、回収した培養上清から、透析やカラムによる溶媒置換などを利用して血清を除去することによっても、血清を含まない培養上清を得ることができる。
血清を含まない「iPS細胞の培養上清」を調製するためには、全過程を通して或いは最後又は最後から数回の継代培養についは無血清培地を使用するとよい。尚、基本培地としてはDMEMの他、イスコフ改変ダルベッコ培地(IMDM)(GIBCO社等)、ハムF12培地(HamF12)(SIGMA社、GIBCO社等)、RPMI1640培地等を用いることができる。二種以上の基本培地を併用することにしてもよい。混合培地の一例として、IMDMとHamF12を等量混合した培地(例えば商品名:IMDM/HamF12(GIBCO社)として市販される)を挙げることができる。また、培地に添加可能な成分の例として、血清(ウシ胎仔血清、ヒト血清、羊血清等)、血清代替物(Knockout serum replacement(KSR)など)、ウシ血清アルブミン(BSA)、抗生物質、各種ビタミン、各種ミネラルを挙げることができる。
The pharmaceutical composition according to the present disclosure may not contain serum. Safety may be improved by not containing serum. For example, a culture supernatant not containing serum can be prepared by culturing iPS cells in a serum-free medium (serum-free medium). A serum-free culture supernatant can also be obtained by performing subculture once or a plurality of times and culturing the last or last several subcultures in a serum-free medium. On the other hand, serum-free culture supernatant can also be obtained from the collected culture supernatant by removing the serum using dialysis or solvent replacement using a column.
In order to prepare “iPS cell culture supernatant” that does not contain serum, a serum-free medium may be used throughout the entire process or for the last or several subcultures from the end. In addition to DMEM, Iscov modified Dulbecco medium (IMDM) (GIBCO, etc.), Ham F12 medium (HamF12) (SIGMA, GIBCO, etc.), RPMI1640 medium, etc. can be used as the basic medium. Two or more basic media may be used in combination. As an example of the mixed medium, a medium in which IMDM and HamF12 are mixed in equal amounts (for example, commercially available as trade name: IMDM / HamF12 (GIBCO)) can be mentioned. Examples of ingredients that can be added to the medium include serum (fetal calf serum, human serum, sheep serum, etc.), serum substitutes (Knockout serum replacement (KSR), etc.), bovine serum albumin (BSA), antibiotics, various Vitamins and various minerals can be mentioned.
培養上清を得るための培養時間としては、例えば5時間〜7日間であり、また、1日〜6日間であってもよい。培養温度は例えば36℃〜38℃、例えば37℃、であり、CO2濃度は4〜6%、例えば5%、である。また、培養は、例えば非接着性条件下での三次元培養、例えば浮遊培養(例えば、分散培養、凝集浮遊培養など)により行ってもよい。 The culture time for obtaining the culture supernatant is, for example, 5 hours to 7 days, and may be 1 day to 6 days. The culture temperature is, for example, 36 ° C to 38 ° C, for example, 37 ° C, and the CO 2 concentration is 4 to 6%, for example, 5%. The culture may be performed, for example, by three-dimensional culture under non-adhesive conditions, for example, suspension culture (for example, dispersion culture, aggregated suspension culture, etc.).
培養後に、細胞成分を分離除去することによって、iPS細胞の培養上清を得ることができる。本開示において、培養上清とは、培養液から細胞成分を分離除去した上清そのものだけでなく、各種処理(例えば、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩、保存等)を適宜施した培養上清もその範囲に含む。培養上清の処理方法の詳細は後述する。本開示において培養上清とは、細胞成分を含まない。このため、本開示における培養上清は、培養に用いられたiPS細胞は含んでいない。 After culturing, the culture supernatant of iPS cells can be obtained by separating and removing cell components. In the present disclosure, the culture supernatant is not only the supernatant itself from which the cell components are separated and removed from the culture solution, but also various treatments (for example, centrifugation, concentration, solvent replacement, dialysis, freezing, drying, freeze-drying, dilution) , Desalted, preserved, etc.) are also included in the range. Details of the culture supernatant treatment method will be described later. In the present disclosure, the culture supernatant does not contain cell components. For this reason, the culture supernatant in this indication does not contain the iPS cell used for culture.
本開示に係る医薬組成物は、上記により得られたiPS細胞の培養上清を有効成分として含むものであり、ある実施形態においては、前記医薬組成物は組成物全体としても前記iPS細胞を含まない。また、別の実施形態では、前記医薬組成物は組成物全体としても細胞(細胞の種類は問わない)を含まない。つまり、無細胞である。当該実施形態の医薬組成物はこの特徴によって、iPS細胞自体は当然のこと、iPS細胞を含む各種組成物と明確に区別される。この実施形態の典型例は、iPS細胞を含まず、iPS細胞の培養上清のみで構成された医薬組成物である。 The pharmaceutical composition according to the present disclosure includes the culture supernatant of iPS cells obtained as described above as an active ingredient. In one embodiment, the pharmaceutical composition includes the iPS cells as a whole composition. Absent. In another embodiment, the pharmaceutical composition does not contain cells (regardless of cell type) as a whole composition. That is, it is cell-free. Due to this characteristic, the pharmaceutical composition of the embodiment is clearly distinguished from various compositions containing iPS cells as well as iPS cells themselves. A typical example of this embodiment is a pharmaceutical composition that does not contain iPS cells and is composed only of culture supernatants of iPS cells.
適用される被検体の状態に応じて、期待される治療効果が維持されることを条件として、本開示に係る医薬組成物は他の成分を追加的に含んでもよい。追加的に含まれ得る成分の一例は以下の通りである。
(i)生体吸収性材料
有機系生体吸収性材料としてヒアルロン酸、コラーゲン、フィブリノーゲン(例えばボルヒール(登録商標))等を使用することができる。
Depending on the condition of the subject to be applied, the pharmaceutical composition according to the present disclosure may additionally contain other components, provided that the expected therapeutic effect is maintained. An example of components that can be additionally included is as follows.
(i) Bioabsorbable material As the organic bioabsorbable material, hyaluronic acid, collagen, fibrinogen (for example, Bolheel (registered trademark)) and the like can be used.
(ii)ゲル化材料
ゲル化材料は、生体親和性が高いものを用いることが好ましく、ヒアルロン酸、コラーゲン又はフィブリン糊等を用いることができる。ヒアルロン酸、コラーゲンとしては種々のものを選択して用いることができるが、本開示に係る医薬組成物の適用目的(適用組織)に適したものを採用することが好ましい。用いるコラーゲンは可溶性(酸可溶性コラーゲン、アルカリ可溶性コラーゲン、酵素可溶性コラーゲン等)であることが好ましい。
(ii) Gelling material As the gelling material, a material having high biocompatibility is preferably used, and hyaluronic acid, collagen, fibrin glue, or the like can be used. Various types of hyaluronic acid and collagen can be selected and used, but it is preferable to employ one suitable for the application purpose (application tissue) of the pharmaceutical composition according to the present disclosure. The collagen used is preferably soluble (acid-soluble collagen, alkali-soluble collagen, enzyme-soluble collagen, etc.).
(iii)その他
製剤上許容される他の成分(例えば、担体、賦形剤、崩壊剤、緩衝剤、乳化剤、懸濁剤、無痛化剤、安定剤、保存剤、防腐剤、生理食塩水など)を含有させることもできる。賦形剤としては乳糖、デンプン、ソルビトール、D-マンニトール、白糖等を用いることができる。崩壊剤としてはデンプン、カルボキシメチルセルロース、炭酸カルシウム等を用いることができる。緩衝剤としてはリン酸塩、クエン酸塩、酢酸塩等を用いることができる。乳化剤としてはアラビアゴム、アルギン酸ナトリウム、トラガント等を用いることができる。懸濁剤としてはモノステアリン酸グリセリン、モノステアリン酸アルミニウム、メチルセルロース、カルボキシメチルセルロース、ヒドロキシメチルセルロース、ラウリル硫酸ナトリウム等を用いることができる。無痛化剤としてはベンジルアルコール、クロロブタノール、ソルビトール等を用いることができる。安定剤としてはプロピレングリコール、アスコルビン酸等を用いることができる。保存剤としてはフェノール、塩化ベンザルコニウム、ベンジルアルコール、クロロブタノール、メチルパラベン等を用いることができる。防腐剤としては塩化ベンザルコニウム、パラオキシ安息香酸、クロロブタノール等を用いることができる。抗生物質、pH調整剤、成長因子(例えば、上皮細胞成長因子(EGF)、神経成長因子(NGF)、脳由来神経栄養因子(BDNF))等を含有させることにしてもよい。
(iii) Others Pharmaceutically acceptable other components (for example, carriers, excipients, disintegrants, buffers, emulsifiers, suspensions, soothing agents, stabilizers, preservatives, preservatives, physiological saline, etc. ) Can also be contained. As the excipient, lactose, starch, sorbitol, D-mannitol, sucrose and the like can be used. As the disintegrant, starch, carboxymethylcellulose, calcium carbonate and the like can be used. Phosphate, citrate, acetate, etc. can be used as the buffer. As the emulsifier, gum arabic, sodium alginate, tragacanth and the like can be used. As the suspending agent, glyceryl monostearate, aluminum monostearate, methyl cellulose, carboxymethyl cellulose, hydroxymethyl cellulose, sodium lauryl sulfate and the like can be used. As the soothing agent, benzyl alcohol, chlorobutanol, sorbitol and the like can be used. As the stabilizer, propylene glycol, ascorbic acid or the like can be used. As preservatives, phenol, benzalkonium chloride, benzyl alcohol, chlorobutanol, methylparaben, and the like can be used. As preservatives, benzalkonium chloride, paraoxybenzoic acid, chlorobutanol, and the like can be used. Antibiotics, pH adjusting agents, growth factors (for example, epidermal growth factor (EGF), nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF)) and the like may be included.
本開示に係る医薬組成物の最終的な形態は特に限定されない。形態の例は液体状(液状、ゲル状など)及び固体状(粉状、細粒、顆粒状など)である。本開示に係る医薬組成物は、吸入に適した形態を有していてもよく、例えば、ネブライザーやディフューザーによって霧状に散布可能な液体の形態を有していてもよい。iPS細胞の培養上清は、事前の準備や保存の点において、iPS細胞自体を用いる場合よりも有利であり、疾患の急性期や亜急性期の治療に特に適するといえる。また、細胞成分を含まず、免疫拒絶の問題を克服し得るという点においても、iPS細胞の培養上清の有用性は極めて高い。 The final form of the pharmaceutical composition according to the present disclosure is not particularly limited. Examples of forms are liquid (liquid, gel, etc.) and solid (powder, fine granules, granules, etc.). The pharmaceutical composition according to the present disclosure may have a form suitable for inhalation, and for example, may have a liquid form that can be sprayed in a mist form with a nebulizer or a diffuser. The culture supernatant of iPS cells is more advantageous than the case of using iPS cells themselves in terms of prior preparation and storage, and can be said to be particularly suitable for the treatment of the acute phase and subacute phase of diseases. Moreover, the usefulness of the culture supernatant of iPS cells is extremely high in that it does not contain cellular components and can overcome the problem of immune rejection.
近年、細胞を用いた再生医療の実現に向けた研究が数多くの研究グループによって進められている。細胞を利用する場合、生体から採取した細胞を培養、選択、処理などに施し、その後回収して移植物の成分とする。そのような従来の研究においては、通常、一連の操作の過程で培養上清は廃棄或いは生理的緩衝液などに置換される。従って、最終的な移植物は培養上清を積極的に含むものではない。このことに鑑みれば、iPS細胞の培養上清を含む点において、本開示に係る医薬組成物は、iPS細胞自体の有用性に注目してiPS細胞を有効成分として使用した組成物などとは、文言上は勿論のこと実質的にも峻別される。 In recent years, research for the realization of regenerative medicine using cells has been promoted by many research groups. When cells are used, the cells collected from the living body are subjected to culture, selection, treatment, etc., and then collected and used as a component of the transplant. In such conventional research, the culture supernatant is usually discarded or replaced with a physiological buffer in the course of a series of operations. Therefore, the final transplant does not actively contain the culture supernatant. In view of this, in terms of including the culture supernatant of iPS cells, the pharmaceutical composition according to the present disclosure is a composition using iPS cells as an active ingredient, focusing on the usefulness of iPS cells themselves. Of course, the wording is also distinct.
とはいえ、実施形態によっては、本開示に係る医薬組成物はiPS細胞の培養上清に加えてiPS細胞を含むものであってもよい。このような場合には、作製後に分化誘導をしていない(換言すれば未分化状態を維持させている)iPS細胞を用いることが好ましい。iPS細胞を追加的に用いることにより、治療効果が向上する場合がある。 However, in some embodiments, the pharmaceutical composition according to the present disclosure may include iPS cells in addition to the culture supernatant of iPS cells. In such a case, it is preferable to use iPS cells that have not been induced to differentiate after production (in other words, maintained in an undifferentiated state). The therapeutic effect may be improved by additionally using iPS cells.
本開示に係る医薬組成物は、iPS細胞の培養上清を含有することにより、組織異常や疾患に対して組織を修復する効果を奏する。この効果の程度は、従来の体性幹細胞の培養上清の使用により得られていた効果を遙かに凌駕する。また、本開示に係る医薬組成物は、驚くべきことに、体内の組織一般に対し広く一般的な修復効果を奏する。このため、本開示に係る医薬組成物は、特定の疾患や組織異常に限定されないユニバーサルな医薬組成物として用いることが可能である。つまり、一種の万能薬として用いることが出来る。このため、本発明の医薬組成物は、疾患や組織異常の発症前であるために、当該疾患や組織異常への罹患が未知の段階であっても、対象への投与により対象内における疾患や組織異常の進行を有効に抑制できる。このため、本開示に係る医薬組成物を習慣的に用いることにより、多岐に渡る疾患や状態異常を発症前に治療することが可能である。 The pharmaceutical composition according to the present disclosure has an effect of repairing a tissue against tissue abnormality or disease by containing a culture supernatant of iPS cells. The degree of this effect far surpasses the effect obtained by using the culture supernatant of conventional somatic stem cells. In addition, the pharmaceutical composition according to the present disclosure surprisingly exhibits a wide and general repair effect on general tissues in the body. For this reason, the pharmaceutical composition according to the present disclosure can be used as a universal pharmaceutical composition not limited to a specific disease or tissue abnormality. In other words, it can be used as a kind of panacea. For this reason, since the pharmaceutical composition of the present invention is before the onset of a disease or tissue abnormality, even if the disease or tissue abnormality is unknown, the disease or The progression of tissue abnormality can be effectively suppressed. For this reason, by using the pharmaceutical composition according to the present disclosure habitually, it is possible to treat various diseases and abnormal conditions before onset.
本開示においては、
(1)iPS細胞を培養するステップ;
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ。
を含む、医薬組成物の製造方法も提供される。この製造方法は、前記回収された培養上清に遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、および脱塩の中から選択される一つ以上の処理を施すステップをさらに含んでもよい。このようなステップを含むことにより、医薬組成物の取り扱いや保存、運搬がより容易になる。また、前記製造方法は、前記回収された培養上清に、追加の成分を添加するステップをさらに含んでいてもよい。そのような追加の成分の添加により、医薬組成物全体の物性を変化させ、その特性を向上することが可能である。また、前記製造方法は、体細胞に初期化因子を導入して前記iPS細胞を作成するステップをさらに含んでいてもよい。各々のステップならびに追加の成分等については、本開示に係る医薬組成物の説明において記載した事項がそのまま当てはまる。前記回収された培養上清に遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、および脱塩の中から選択される一つ以上の処理を施すステップと、前記回収された培養上清に、追加の成分を添加するステップの両方を含む場合には、両ステップはどちらを先に行ってもよく、また可能な場合には同時並行して行ってもよい。
In this disclosure,
(1) culturing iPS cells;
(2) A step of recovering the culture supernatant obtained by culturing the iPS cells.
A method for producing a pharmaceutical composition is also provided. In this production method, the collected culture supernatant is subjected to one or more treatments selected from centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, and desalting. May further be included. By including such a step, handling, storage and transportation of the pharmaceutical composition become easier. The production method may further include a step of adding an additional component to the collected culture supernatant. By adding such an additional component, it is possible to change the physical properties of the entire pharmaceutical composition and to improve its properties. The production method may further include a step of creating the iPS cell by introducing an reprogramming factor into a somatic cell. For each step and additional components, the matters described in the explanation of the pharmaceutical composition according to the present disclosure are applied as they are. Subjecting the collected culture supernatant to one or more treatments selected from centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, and desalting; When the culture supernatant includes both steps of adding additional components, both steps may be performed first, and if possible, may be performed simultaneously.
上記ステップ(2)においては、iPS細胞の培養上清を回収する。例えば、スポイトやピペットなどで培養液を吸引して回収することができる。回収した培養上清はそのまま或いは一以上の処理を経た後に本開示に係る医薬組成物の有効成分として使用される。ここでの処理として、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩、保存(例えば、4℃、-80℃)を例示することができる。尚、iPS細胞の培養上清は、複雑高度な精製をしなくとも、所期の作用を示す。このため、本開示に係る医薬組成物は簡便な工程で製造できる。複雑な精製工程を要しないことは、精製に伴う活性の低下を回避できる点においても有利である。 In step (2) above, the culture supernatant of iPS cells is collected. For example, the culture solution can be collected by suction with a dropper or pipette. The collected culture supernatant is used as an active ingredient of the pharmaceutical composition according to the present disclosure as it is or after one or more treatments. Examples of the treatment here include centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, and storage (eg, 4 ° C., −80 ° C.). It should be noted that the culture supernatant of iPS cells exhibits the desired action without complicated and sophisticated purification. For this reason, the pharmaceutical composition according to the present disclosure can be produced by a simple process. The fact that a complicated purification step is not required is also advantageous in that a decrease in activity associated with purification can be avoided.
<iPS細胞培養上清の濃縮方法>
本開示に係る医薬組成物は、製剤化されたものであってもよい。製剤化のためのiPS細胞培養上清の濃縮方法としては、培養上清の濃縮に通常用いられている方法を適用することができる。濃縮方法の例としては、例えば、以下の二つの方法を挙げることができる。
1. スピンカラム濃縮法
培養上清をAmicon Ultra Centrifugal Filter Units-10K(ミリポア社製)を用いて濃縮する(最大75倍濃縮)。具体的な操作手順は次の通りである。
(i) 培養上清(最大15ml)をAmicon Ultra Centrifugal Filter Units-10Kへ投入し、×4000gで約60分間遠心し、200μlまで濃縮する。
(ii) 上記チューブへ培養上清と同量の滅菌PBSを投入し、再度×4000gで約60分間遠心し、ベース溶液をPBSへ置換する。
(iii) 得られた溶液200μlをマイクロテストチューブへ回収し、濃縮iPS細胞培養上清とする。
<Concentration method of iPS cell culture supernatant>
The pharmaceutical composition according to the present disclosure may be formulated. As a method for concentrating the iPS cell culture supernatant for formulation, a method usually used for concentrating the culture supernatant can be applied. Examples of the concentration method include the following two methods.
1. Spin column concentration method The culture supernatant is concentrated using Amicon Ultra Centrifugal Filter Units-10K (Millipore) (maximum 75-fold concentration). The specific operation procedure is as follows.
(i) The culture supernatant (up to 15 ml) is put into Amicon Ultra Centrifugal Filter Units-10K, centrifuged at 4000 g for about 60 minutes, and concentrated to 200 μl.
(ii) Put the same amount of sterile PBS as the culture supernatant into the above tube, and centrifuge again at × 4000g for about 60 minutes to replace the base solution with PBS.
(iii) Collect 200 μl of the obtained solution into a micro test tube to obtain a concentrated iPS cell culture supernatant.
2. エタノール沈殿濃縮法
培養上清をエタノール沈殿法を用いて濃縮する(最大10倍濃縮)。具体的な操作手順は次の通りである。
(i) 培養上清5mlに対し100%エタノール45mlを加え、混和し、-20℃で60分間放置する。
(ii) 4℃、×15000gで15分間遠心する。
(iii) 上澄みを除去し、90%エタノール10mlを加え、よく攪拌する。
(iv) 4℃、×15000gで5分間遠心する。
(v) 上澄みを除去し、得られたペレットを滅菌水500μlに溶解し、マイクロテストチューブへ回収し、濃縮iPS細胞培養上清とする。
2. Ethanol precipitation concentration method Concentrate the culture supernatant using ethanol precipitation method (concentration up to 10 times). The specific operation procedure is as follows.
(i) Add 45 ml of 100% ethanol to 5 ml of the culture supernatant, mix and leave at -20 ° C for 60 minutes.
(ii) Centrifuge for 15 minutes at 4 ° C and 15000g.
(iii) Remove the supernatant, add 10 ml of 90% ethanol and stir well.
(iv) Centrifuge at 4 ° C and 15000g for 5 minutes.
(v) The supernatant is removed, and the resulting pellet is dissolved in 500 μl of sterilized water, collected in a micro test tube, and used as a concentrated iPS cell culture supernatant.
<iPS細胞培養上清の凍結乾燥方法>
また本開示に係る医薬組成物におけるiPS細胞培養上清は、凍結乾燥されたものであってもよい。これにより、良好な保存安定性が得られる。iPS細胞培養上清の凍結乾燥方法としては、培養上清の凍結乾燥に通常用いられている方法を適用することができる。凍結乾燥方法の例としては、例えば、以下の方法を挙げることができる。
(i) 上記方法で得られたiPS細胞培養上清又は濃縮iPS細胞培養上清を-80℃で2時間から半日凍結する。
(ii) 凍結後、サンプルチューブの蓋を開放し、凍結乾燥機へセットする。
(iii) 1〜2日間凍結乾燥を行う。
(iv) 得られたサンプルを凍結乾燥iPS細胞培養上清とする(-80℃で保存可能)。
<Method of freeze-drying iPS cell culture supernatant>
The iPS cell culture supernatant in the pharmaceutical composition according to the present disclosure may be lyophilized. Thereby, good storage stability is obtained. As a lyophilization method of the iPS cell culture supernatant, a method generally used for lyophilization of the culture supernatant can be applied. Examples of the lyophilization method include the following methods.
(i) The iPS cell culture supernatant or concentrated iPS cell culture supernatant obtained by the above method is frozen at −80 ° C. for 2 hours to half a day.
(ii) After freezing, open the lid of the sample tube and set it in the freeze dryer.
(iii) Lyophilize for 1-2 days.
(iv) The obtained sample is used as a lyophilized iPS cell culture supernatant (can be stored at -80 ° C).
本開示においては、本開示に係る医薬組成物を、疾患または組織異常の発症前の対象に、前記疾患または組織異常の発症を抑えるために有効な量投与することを含む、前記対象において疾患または組織異常の発症前に前記疾患または組織異常の発症を抑える方法、も提供される。前記対象は、ヒト、またはヒト以外の哺乳動物(ペット動物、家畜、実験動物を含む。具体的には例えばマウス、ラット、モルモット、ハムスター、サル、ウシ、ブタ、ヤギ、ヒツジ、イヌ、ネコ等)であってもよい。前記対象は、疾患または組織異常を発病するリスクを有すると判定された対象であってもよい。そのようなリスクは、遺伝子診断、家系分析等により判定することが出来る。例えば、特定の遺伝子における特定のアレルの存在が、特定の疾患への罹患確率に相関することが統計的に明らかにされている例がある。 In the present disclosure, the pharmaceutical composition according to the present disclosure is administered to a subject prior to the onset of a disease or tissue abnormality in an amount effective for suppressing the onset of the disease or tissue abnormality. A method for suppressing the onset of the disease or tissue abnormality before the onset of the tissue abnormality is also provided. The subjects include humans or non-human mammals (pet animals, domestic animals, laboratory animals. Specifically, for example, mice, rats, guinea pigs, hamsters, monkeys, cows, pigs, goats, sheep, dogs, cats, etc. ). The subject may be a subject determined to have a risk of developing a disease or tissue abnormality. Such a risk can be determined by genetic diagnosis, family analysis or the like. For example, in some cases, it has been statistically revealed that the presence of a specific allele in a specific gene correlates with the probability of suffering a specific disease.
一般に疾患や組織異常は、進行すればするほど治療はより困難になる。このため、そうした疾患や組織異常の早期発見が重要となる訳であるが、毎年の検診によって早期発見が確実にできるわけではない。この点を考慮すると、疾患や組織異常の発症前に予め治療を開始できることが望ましいが、そのような治療に用いられる薬剤は継続的に用いられるものである以上、投与対象者に与える負担が少ないものであることが望ましく、またその段階ではまだ明らかではない疾患あるいは組織異常の進行を抑えなければならないという点からは、疾患あるいは組織異常一般に広く有効性を有するものであることが望ましい。これらの要求を同時に満たすことは、従来の知見では困難であった。しかし、本開示に係る医薬組成物を用いれば、移植や注射といった侵襲的処置を行わなくても投与可能であり、また前記医薬組成物は疾患あるいは組織異常一般に広く有効性を有する。このことから、前記医薬組成物は、上記のような、発症前における疾患や組織異常の早期処置(進行抑制)に高い有効性を有する。このような発症前における早期処置のことを、本開示においては先制医療と称する。 In general, as disease and tissue abnormality progress, the treatment becomes more difficult. For this reason, early detection of such diseases and tissue abnormalities is important, but early detection is not always reliable. In consideration of this point, it is desirable that treatment can be started in advance before the onset of a disease or tissue abnormality, but since the drug used for such treatment is continuously used, the burden on the administration subject is small. In view of the necessity of suppressing the progression of a disease or tissue abnormality that is not yet clear at that stage, it is desirable that the disease or tissue abnormality generally has wide efficacy. Satisfying these requirements simultaneously has been difficult with conventional knowledge. However, if the pharmaceutical composition according to the present disclosure is used, it can be administered without invasive treatment such as transplantation or injection, and the pharmaceutical composition has wide efficacy in general for diseases or tissue abnormalities. From this, the said pharmaceutical composition has high effectiveness in the early treatment (progression suppression) of the disease and tissue abnormality before onset as mentioned above. Such early treatment before onset is referred to as preemptive medicine in the present disclosure.
前記医薬組成物の投与量は、未処理の培養上清の量に換算して、例えば0.1mg/kg/日〜1000mg/kg/日であり、また、1mg/kg/日〜100mg/kg/日であってもよい。また、投与の方法は特に制限されない。例えば、前記医薬組成物の投与は、非経口投与であることが好ましく、非経口投与としては、全身性投与であっても局所投与であってもよい。局所投与の例としては、目的組織への注入、塗布又は噴霧などを挙げることができる。前記医薬組成物の投与方法の例としては、静脈内投与、動脈内投与、門脈内投与、皮内投与、皮下投与、筋肉内投与、腹腔内投与、経肺投与(経肺吸収)及び鼻腔内投与等を挙げることができる。中でも、鼻腔内投与、経肺投与等は、低侵襲性であり、好ましい。投与スケジュールとしては例えば一日一回〜数回、二日に一回、或いは三日に一回などを採用できる。投与スケジュールの作成においては、対象(レシピエント)の性別、年齢、体重、病態などを考慮することができる。 The dosage of the pharmaceutical composition is, for example, 0.1 mg / kg / day to 1000 mg / kg / day, and 1 mg / kg / day to 100 mg / kg in terms of the amount of untreated culture supernatant. / Day may be sufficient. Moreover, the administration method is not particularly limited. For example, the pharmaceutical composition is preferably administered parenterally, and the parenteral administration may be systemic administration or local administration. Examples of topical administration include injection into the target tissue, application or spraying. Examples of methods for administering the pharmaceutical composition include intravenous administration, intraarterial administration, intraportal administration, intradermal administration, subcutaneous administration, intramuscular administration, intraperitoneal administration, transpulmonary administration (transpulmonary absorption), and nasal cavity. An internal administration etc. can be mentioned. Of these, intranasal administration, transpulmonary administration and the like are preferable because they are minimally invasive. As the administration schedule, for example, once to several times a day, once every two days, or once every three days can be adopted. In preparing the administration schedule, the gender, age, weight, disease state, etc. of the subject (recipient) can be considered.
投与方法の選択は、発症抑制の目標となる組織の種類、疾患の種類等に基づいて当業者により行うことができる。例えば、当該組織が頭部にある場合には、血液脳関門の通過を考慮する必要がなく、低侵襲性であることから、鼻腔内投与等を適用することが特に好ましい。例えば、前記組織が脳である場合には、鼻腔内投与が好ましく適用され得る。また、脳梗塞の先制医療において、鼻腔内投与が好ましく適用され得る。経肺投与は、生活環境の中で長時間投与を行うことが出来る点で投与対象者の負担が少なく、ディフューザー等を使用して前記医薬組成物を霧状に噴霧することにより実施できる。 Selection of the administration method can be performed by those skilled in the art based on the type of tissue, the type of disease, and the like that are the targets of onset suppression. For example, when the tissue is in the head, it is not necessary to consider the passage through the blood brain barrier, and it is particularly preferable to apply intranasal administration or the like because it is minimally invasive. For example, when the tissue is the brain, intranasal administration can be preferably applied. In addition, intranasal administration can be preferably applied in preemptive medical treatment for cerebral infarction. Transpulmonary administration is less burdensome on the subject of administration because it can be administered for a long time in the living environment, and can be carried out by spraying the pharmaceutical composition in a mist form using a diffuser or the like.
なお、実施形態によっては、本開示に係る医薬組成物に加えてiPS細胞も投与することが出来る。例えば、前記医薬組成物とiPS細胞とを同時に、または別々のタイミングで、対象に投与することができる。このような場合には、採取後に分化誘導をしていない(換言すれば未分化状態を維持させている)iPS細胞を用いることが好ましい。前記医薬組成物とiPSを併用する場合には、前記医薬組成物を含有する第1構成要素と、iPS細胞を含有する第2構成要素とからなるキットを用いて投与を行ってもよい。各構成要素は、例えば各々別々のカプセルであったり、あるいは別々のアンプルあるいはバイアルであってもよい。この場合、第1構成要素を投与した治療対象に対して、第1構成要素の投与と同時又は投与後に第2構成要素を投与してもよい。尚、ここでの「同時」は厳密な同時性を要求するものではない。従って、両要素を混合した後に対象へ投与する等、両要素の投与が時間差のない条件下で実施される場合は勿論のこと、片方の投与後、速やかに他方を投与する等、両要素の投与が実質的な時間差のない条件下で実施される場合もここでの「同時」の概念に含まれる。 In some embodiments, iPS cells can also be administered in addition to the pharmaceutical composition according to the present disclosure. For example, the pharmaceutical composition and iPS cells can be administered to a subject simultaneously or at different times. In such a case, it is preferable to use iPS cells that have not been differentiated after collection (in other words, maintained in an undifferentiated state). When using the said pharmaceutical composition and iPS together, you may administer using the kit which consists of the 1st component containing the said pharmaceutical composition, and the 2nd component containing an iPS cell. Each component may be a separate capsule, for example, or a separate ampoule or vial. In this case, you may administer a 2nd component with respect to the treatment target which administered the 1st component simultaneously with the administration of a 1st component, or after administration. Here, “simultaneous” does not require strict simultaneity. Therefore, when both elements are administered under the condition that there is no time difference, for example, both elements are mixed and then administered to the subject, both elements are immediately administered after one is administered. The case where the administration is performed under conditions without a substantial time difference is also included in the concept of “simultaneous” herein.
前記疾患は、特に限定はされない。例えば、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、(I型II型糖尿病に見られるような)ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患(筋萎縮性側索硬化症、アルツハイマー病、パーキンソン病、進行性核上性麻痺、ハンチントン病、多系統萎縮症、脊髄小脳変性症等)、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患(外傷性網膜剥離、網膜裂孔、網膜振盪症、視神経管骨折、糖尿病網膜症、加齢黄斑変性、網膜色素変性症、緑内障、コロイデレミア、レーベル先天盲、錐体ジストロフィ、家族性ドルーゼン、中心性輪紋状脈絡膜ジストロフィ、常染色体優性視神経萎縮など)、神経難病、歯周病、皮膚潰瘍、骨粗鬆症等が挙げられる。前記疾患は、加齢に伴って発生するあるいは進行する疾患であってもよい。また、前記医薬組成物の投与は、加齢に伴う前記疾患の発生あるいは進行を抑えるものであってもよい。 The disease is not particularly limited. For example, gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related changes in heart, hypertension, ischemic heart disease, valvular heart disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, pulmonary fibrosis, (I Langerhans island atrophy (as seen in type II diabetes), liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases (amyotrophic lateral sclerosis, Alzheimer's disease) Parkinson's disease, progressive supranuclear palsy, Huntington's disease, multiple system atrophy, spinocerebellar degeneration etc.), neuronal degeneration / dropout due to cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. , Retinal disease with neuronal damage (traumatic retinal detachment, retinal tear, retinal shaking, optic nerve tube fracture, diabetic retinopathy, age-related macular degeneration, retinitis pigmentosa, glaucoma, colloidemia, label congenital blindness, cone Body Rofi, familial drusen, central areolar choroidal dystrophy, autosomal dominant optic atrophy etc.), intractable neurological diseases, periodontal disease, skin ulcers, osteoporosis, and the like. The disease may be a disease that develops or progresses with aging. Moreover, administration of the pharmaceutical composition may suppress the occurrence or progression of the disease associated with aging.
前記組織異常は、若年層における正常な組織と比較した場合の、組織の変性を意味する。いわゆる老化も組織異常の中に含まれる。老化の代表的な症状としては、例えば、視力低下、聴力低下、筋力低下、骨密度減少、腎機能低下、肝機能低下、膵臓機能低下、神経細胞数減少、記憶力低下、などが挙げられる。また、老化の症状は、組織の萎縮、線維化、細胞数の減少、色素沈着、寿命を短縮させるような組織のあるいは全身の変化などであってもよい。また、組織異常には、組織の物理的又は生理的欠陥に起因した傷害部あるいは障害部も包含される
前記疾患あるいは組織異常の発生する部位としては、食道、胃、膵臓、肝臓、脾臓、腎臓、小腸、十二指腸、大腸、皮膚、眼、耳、鼻腔、気管、肺、血管、脳、骨、筋肉、咽頭、神経(脊髄、末梢神経等)が挙げられる。
前記組織異常は、加齢に伴って発生するあるいは進行する組織異常であってもよい。また、前記医薬組成物の投与は、加齢に伴う前記組織異常の発生あるいは進行を抑えるものであってもよい。
Said tissue abnormality refers to tissue degeneration as compared to normal tissue in younger age groups. So-called aging is also included in tissue abnormalities. Representative symptoms of aging include, for example, decreased visual acuity, decreased hearing, decreased muscle strength, decreased bone density, decreased renal function, decreased liver function, decreased pancreatic function, decreased number of neurons, decreased memory ability, and the like. The aging symptoms may be tissue atrophy, fibrosis, a decrease in the number of cells, pigmentation, a change in the tissue or the whole body that shortens the life span, and the like. In addition, tissue abnormalities include injured or damaged parts caused by physical or physiological defects in the tissue. The sites where the diseases or tissue abnormalities occur include the esophagus, stomach, pancreas, liver, spleen, kidney. , Small intestine, duodenum, large intestine, skin, eye, ear, nasal cavity, trachea, lung, blood vessel, brain, bone, muscle, pharynx, nerve (spinal cord, peripheral nerve, etc.).
The tissue abnormality may be a tissue abnormality that occurs or progresses with aging. Moreover, administration of the pharmaceutical composition may suppress the occurrence or progression of the tissue abnormality associated with aging.
上記の方法において、疾患または組織異常の発症の抑制は、内在性の幹細胞の能力に基づいて達成されるものであってもよい。本開示に係る医薬組成物は、サイトカインを始めとする種々の成分を含んでおり、これらの成分は内在性の幹細胞の能力を刺激して、疾患または組織異常の発症を抑制することが可能だからである。 In the above method, suppression of the onset of a disease or tissue abnormality may be achieved based on the ability of endogenous stem cells. The pharmaceutical composition according to the present disclosure includes various components including cytokines, and these components can stimulate the ability of endogenous stem cells to suppress the onset of diseases or tissue abnormalities. It is.
もちろん、本開示に係る医薬組成物の用途は先制医療に制限されるものではない。このため、本開示によれば、本開示に係る医薬組成物を、疾患を有する対象に、前記疾患を治療するために有効な量で投与することを含む、疾患の治療方法も提供される。
前記対象は、ヒト、またはヒト以外の哺乳動物(ペット動物、家畜、実験動物を含む。具体的には例えばマウス、ラット、モルモット、ハムスター、サル、ウシ、ブタ、ヤギ、ヒツジ、イヌ、ネコ等)であってもよい。
Of course, the use of the pharmaceutical composition according to the present disclosure is not limited to preemptive medicine. For this reason, according to the present disclosure, there is also provided a method for treating a disease, comprising administering the pharmaceutical composition according to the present disclosure to a subject having the disease in an amount effective for treating the disease.
The subjects include humans or non-human mammals (pet animals, domestic animals, laboratory animals. Specifically, for example, mice, rats, guinea pigs, hamsters, monkeys, cows, pigs, goats, sheep, dogs, cats, etc. ).
前記疾患の治療方法における、前記医薬組成物の投与量は、未処理の培養上清の量に換算して、例えば0.1mg/kg/日〜1000mg/kg/日であり、また、1mg/kg/日〜100mg/kg/日であってもよい。また、投与の方法は特に制限されない。例えば、前記医薬組成物の投与は、非経口投与であることが好ましく、非経口投与としては、全身性投与であっても局所投与であってもよい。局所投与の例としては、目的組織への注入、塗布又は噴霧などを挙げることができる。前記医薬組成物の投与方法の例としては、静脈内投与、動脈内投与、門脈内投与、皮内投与、皮下投与、筋肉内投与、腹腔内投与、経肺投与(経肺吸収)及び鼻腔内投与等を挙げることができる。中でも、鼻腔内投与、経肺投与等は、低侵襲性であり、好ましい。投与スケジュールとしては例えば一日一回〜数回、二日に一回、或いは三日に一回などを採用できる。投与スケジュールの作成においては、対象(レシピエント)の性別、年齢、体重、病態などを考慮することができる。 In the method for treating diseases, the dosage of the pharmaceutical composition is, for example, 0.1 mg / kg / day to 1000 mg / kg / day in terms of the amount of untreated culture supernatant, and 1 mg / kg It may be kg / day to 100 mg / kg / day. Moreover, the administration method is not particularly limited. For example, the pharmaceutical composition is preferably administered parenterally, and the parenteral administration may be systemic administration or local administration. Examples of topical administration include injection into the target tissue, application or spraying. Examples of methods for administering the pharmaceutical composition include intravenous administration, intraarterial administration, intraportal administration, intradermal administration, subcutaneous administration, intramuscular administration, intraperitoneal administration, transpulmonary administration (transpulmonary absorption), and nasal cavity. An internal administration etc. can be mentioned. Of these, intranasal administration, transpulmonary administration and the like are preferable because they are minimally invasive. As the administration schedule, for example, once to several times a day, once every two days, or once every three days can be adopted. In preparing the administration schedule, the gender, age, weight, disease state, etc. of the subject (recipient) can be considered.
なお、本開示において「治療」の範囲には、疾患や組織異常を根治する処置だけでなく、根治に至らないまでも疾患や組織異常の進行を停止させる、あるいは処置をしない場合に比べて遅らせる、処置も含まれる。例えば、加齢による疾患や組織異常の進行を停止させるあるいは遅らせることも、治療の範囲に含まれる。また、本開示において治療の開始時期は、発症が見られてからでもよく、また、前述のとおり発症前に未然に行うものであってもよい。 In the present disclosure, the range of “treatment” includes not only a treatment to cure a disease or tissue abnormality, but also stops the progression of the disease or tissue abnormality until it is not completely cured, or delays compared to a case in which no treatment is performed. Treatment is also included. For example, it is also within the scope of treatment to stop or delay the progression of diseases and tissue abnormalities due to aging. Further, in the present disclosure, the start time of treatment may be after onset is observed, or may be performed before onset as described above.
前記疾患の治療方法における、投与方法の選択は、疾患組織の種類、疾患の種類等に基づいて当業者により行うことができる。例えば、当該組織が頭部にある場合には、血液脳関門の通過を考慮する必要がなく、低侵襲性であることから、鼻腔内投与等を適用することが特に好ましい。例えば、前記組織が脳である場合には、鼻腔内投与が好ましく適用され得る。また、脳梗塞の治療に対して、鼻腔内投与が好ましく適用され得る。経肺投与は、生活環境の中で長時間投与を行うことが出来る点で投与対象者の負担が少なく、ディフューザー等を使用して前記医薬組成物を霧状に噴霧することにより実施できる。 Selection of the administration method in the disease treatment method can be performed by those skilled in the art based on the type of disease tissue, the type of disease, and the like. For example, when the tissue is in the head, it is not necessary to consider the passage through the blood brain barrier, and it is particularly preferable to apply intranasal administration or the like because it is minimally invasive. For example, when the tissue is the brain, intranasal administration can be preferably applied. In addition, intranasal administration can be preferably applied to the treatment of cerebral infarction. Transpulmonary administration is less burdensome on the subject of administration because it can be administered for a long time in the living environment, and can be carried out by spraying the pharmaceutical composition in a mist form using a diffuser or the like.
なお、実施形態によっては、本開示に係る医薬組成物に加えてiPS細胞も投与することが出来る。例えば、前記医薬組成物とiPS細胞とを同時に、または別々のタイミングで、対象に投与することができる。このような場合には、採取後に分化誘導をしていない(換言すれば未分化状態を維持させている)iPS細胞を用いることが好ましい。前記医薬組成物とiPSを併用する場合には、前記医薬組成物を含有する第1構成要素と、iPS細胞を含有する第2構成要素とからなるキットを用いて投与を行ってもよい。各構成要素は、例えば各々別々のカプセルであったり、あるいは別々のアンプルあるいはバイアルであってもよい。 In some embodiments, iPS cells can also be administered in addition to the pharmaceutical composition according to the present disclosure. For example, the pharmaceutical composition and iPS cells can be administered to a subject simultaneously or at different times. In such a case, it is preferable to use iPS cells that have not been differentiated after collection (in other words, maintained in an undifferentiated state). When using the said pharmaceutical composition and iPS together, you may administer using the kit which consists of the 1st component containing the said pharmaceutical composition, and the 2nd component containing an iPS cell. Each component may be a separate capsule, for example, or a separate ampoule or vial.
この場合、第1構成要素を投与した治療対象に対して、第1構成要素の投与と同時又は投与後に第2構成要素を投与してもよい。第1構成要素と第2構成要素を同時に投与するという使用法は急性期や亜急性期の疾患への適用に特に適する。尚、ここでの「同時」は厳密な同時性を要求するものではない。従って、両要素を混合した後に対象へ投与する等、両要素の投与が時間差のない条件下で実施される場合は勿論のこと、片方の投与後、速やかに他方を投与する等、両要素の投与が実質的な時間差のない条件下で実施される場合もここでの「同時」の概念に含まれる。 In this case, you may administer a 2nd component with respect to the treatment target which administered the 1st component simultaneously with the administration of a 1st component, or after administration. The use of administering the first component and the second component simultaneously is particularly suitable for application to acute and subacute disease. Here, “simultaneous” does not require strict simultaneity. Therefore, when both elements are administered under the condition that there is no time difference, for example, both elements are mixed and then administered to the subject, both elements are immediately administered after one is administered. The case where the administration is performed under conditions without a substantial time difference is also included in the concept of “simultaneous” herein.
前記疾患の治療方法によって治療される疾患は、特に限定はされない。例えば、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、(I型II型糖尿病に見られるような)ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患(筋萎縮性側索硬化症、アルツハイマー病、パーキンソン病、進行性核上性麻痺、ハンチントン病、多系統萎縮症、脊髄小脳変性症等)、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患(外傷性網膜剥離、網膜裂孔、網膜振盪症、視神経管骨折、糖尿病網膜症、加齢黄斑変性、網膜色素変性症、緑内障、コロイデレミア、レーベル先天盲、錐体ジストロフィ、家族性ドルーゼン、中心性輪紋状脈絡膜ジストロフィ、常染色体優性視神経萎縮など)、神経難病、歯周病、皮膚潰瘍、骨粗鬆症等が挙げられる。 The disease to be treated by the disease treatment method is not particularly limited. For example, gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related changes in heart, hypertension, ischemic heart disease, valvular heart disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, pulmonary fibrosis, (I Langerhans island atrophy (as seen in type II diabetes), liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases (amyotrophic lateral sclerosis, Alzheimer's disease) Parkinson's disease, progressive supranuclear palsy, Huntington's disease, multiple system atrophy, spinocerebellar degeneration etc.), neuronal degeneration / dropout due to cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. , Retinal disease with neuronal damage (traumatic retinal detachment, retinal tear, retinal shaking, optic nerve tube fracture, diabetic retinopathy, age-related macular degeneration, retinitis pigmentosa, glaucoma, colloidemia, label congenital blindness, cone Body Rofi, familial drusen, central areolar choroidal dystrophy, autosomal dominant optic atrophy etc.), intractable neurological diseases, periodontal disease, skin ulcers, osteoporosis, and the like.
前記疾患の部位としては、食道、胃、膵臓、肝臓、脾臓、腎臓、小腸、十二指腸、大腸、皮膚、眼、耳、鼻腔、気管、肺、血管、脳、骨、筋肉、咽頭、神経(脊髄、末梢神経等)が挙げられる。前記疾患は、加齢に伴って発生するあるいは進行する疾患であってもよい。また、前記医薬組成物の投与は、加齢に伴う前記疾患の発生あるいは進行を抑えるものであってもよい。 The site of the disease includes the esophagus, stomach, pancreas, liver, spleen, kidney, small intestine, duodenum, large intestine, skin, eye, ear, nasal cavity, trachea, lung, blood vessel, brain, bone, muscle, pharynx, nerve (spinal cord) Peripheral nerve, etc.). The disease may be a disease that develops or progresses with aging. Moreover, administration of the pharmaceutical composition may suppress the occurrence or progression of the disease associated with aging.
上記の方法において、疾患の治療は、内在性の幹細胞の能力に基づいて達成されるものであってもよい。本開示に係る医薬組成物は、サイトカインを始めとする種々の成分を含んでおり、これらの成分は内在性の幹細胞の能力を刺激して、疾患の治療を達成することが可能だからである。 In the above method, treatment of the disease may be achieved based on the ability of endogenous stem cells. This is because the pharmaceutical composition according to the present disclosure includes various components including cytokines, and these components can stimulate the ability of endogenous stem cells to achieve treatment of a disease.
また、本開示によれば、本開示に係る医薬組成物を、組織異常を有する対象に、前記組織異常を治療するために有効な量で投与することを含む、組織異常の治療方法、も提供される。
前記対象は、ヒト、またはヒト以外の哺乳動物(ペット動物、家畜、実験動物を含む。具体的には例えばマウス、ラット、モルモット、ハムスター、サル、ウシ、ブタ、ヤギ、ヒツジ、イヌ、ネコ等)であってもよい。
前記組織異常の治療方法における、前記医薬組成物の投与量は、未処理の培養上清の量に換算して、例えば0.1mg/kg/日〜1000mg/kg/日であり、また、1mg/kg/日〜100mg/kg/日であってもよい。また、投与の方法は特に制限されない。例えば、前記医薬組成物の投与は、非経口投与であることが好ましく、非経口投与としては、全身性投与であっても局所投与であってもよい。局所投与の例としては、目的組織への注入、塗布又は噴霧などを挙げることができる。前記医薬組成物の投与方法の例としては、静脈内投与、動脈内投与、門脈内投与、皮内投与、皮下投与、筋肉内投与、腹腔内投与、経肺投与(経肺吸収)及び鼻腔内投与等を挙げることができる。中でも、鼻腔内投与、経肺投与等は、低侵襲性であり、好ましい。投与スケジュールとしては例えば一日一回〜数回、二日に一回、或いは三日に一回などを採用できる。投与スケジュールの作成においては、対象の性別、年齢、体重、病態などを考慮することができる。
In addition, according to the present disclosure, there is also provided a method for treating a tissue abnormality, comprising administering the pharmaceutical composition according to the present disclosure to a subject having a tissue abnormality in an amount effective for treating the tissue abnormality. Is done.
The subjects include humans or non-human mammals (pet animals, domestic animals, laboratory animals. Specifically, for example, mice, rats, guinea pigs, hamsters, monkeys, cows, pigs, goats, sheep, dogs, cats, etc. ).
In the method for treating tissue abnormalities, the dosage of the pharmaceutical composition is, for example, 0.1 mg / kg / day to 1000 mg / kg / day in terms of the amount of untreated culture supernatant, and 1 mg / Kg / day to 100 mg / kg / day. Moreover, the administration method is not particularly limited. For example, the pharmaceutical composition is preferably administered parenterally, and the parenteral administration may be systemic administration or local administration. Examples of topical administration include injection into the target tissue, application or spraying. Examples of methods for administering the pharmaceutical composition include intravenous administration, intraarterial administration, intraportal administration, intradermal administration, subcutaneous administration, intramuscular administration, intraperitoneal administration, transpulmonary administration (transpulmonary absorption), and nasal cavity. An internal administration etc. can be mentioned. Of these, intranasal administration, transpulmonary administration and the like are preferable because they are minimally invasive. As the administration schedule, for example, once to several times a day, once every two days, or once every three days can be adopted. In preparing the administration schedule, the sex, age, weight, disease state, etc. of the subject can be considered.
前記組織異常の治療方法における、投与方法の選択は、組織異常を生じている組織の種類、疾患の種類等に基づいて当業者により行うことができる。例えば、当該組織が頭部にある場合には、血液脳関門の通過を考慮する必要がなく、低侵襲性であることから、鼻腔内投与等を適用することが特に好ましい。例えば、前記組織が脳である場合には、鼻腔内投与が好ましく適用され得る。経肺投与は、生活環境の中で長時間投与を行うことが出来る点で投与対象者の負担が少なく、ディフューザー等を使用して前記医薬組成物を霧状に噴霧することにより実施できる。 Selection of the administration method in the tissue abnormality treatment method can be performed by those skilled in the art based on the type of tissue in which the tissue abnormality has occurred, the type of disease, and the like. For example, when the tissue is in the head, it is not necessary to consider the passage through the blood brain barrier, and it is particularly preferable to apply intranasal administration or the like because it is minimally invasive. For example, when the tissue is the brain, intranasal administration can be preferably applied. Transpulmonary administration is less burdensome on the subject of administration because it can be administered for a long time in the living environment, and can be carried out by spraying the pharmaceutical composition in a mist form using a diffuser or the like.
なお、実施形態によっては、本開示に係る医薬組成物に加えてiPS細胞も投与することが出来る。例えば、前記医薬組成物とiPS細胞とを同時に、または別々のタイミングで、対象に投与することができる。このような場合には、採取後に分化誘導をしていない(換言すれば未分化状態を維持させている)iPS細胞を用いることが好ましい。 In some embodiments, iPS cells can also be administered in addition to the pharmaceutical composition according to the present disclosure. For example, the pharmaceutical composition and iPS cells can be administered to a subject simultaneously or at different times. In such a case, it is preferable to use iPS cells that have not been differentiated after collection (in other words, maintained in an undifferentiated state).
この場合、第1構成要素を投与した治療対象に対して、第1構成要素の投与と同時又は投与後に第2構成要素を投与してもよい。第1構成要素と第2構成要素を同時に投与するという使用法は急性期や亜急性期の組織異常への適用に特に適する。尚、ここでの「同時」は厳密な同時性を要求するものではない。従って、両要素を混合した後に対象へ投与する等、両要素の投与が時間差のない条件下で実施される場合は勿論のこと、片方の投与後、速やかに他方を投与する等、両要素の投与が実質的な時間差のない条件下で実施される場合もここでの「同時」の概念に含まれる。 In this case, you may administer a 2nd component with respect to the treatment target which administered the 1st component simultaneously with the administration of a 1st component, or after administration. The use of administering the first component and the second component simultaneously is particularly suitable for application to acute and subacute tissue abnormalities. Here, “simultaneous” does not require strict simultaneity. Therefore, when both elements are administered under the condition that there is no time difference, for example, both elements are mixed and then administered to the subject, both elements are immediately administered after one is administered. The case where the administration is performed under conditions without a substantial time difference is also included in the concept of “simultaneous” herein.
前記組織異常としては、視力低下、聴力低下、筋力低下、骨密度減少、腎機能低下、肝機能低下、膵臓機能低下、神経細胞数減少、記憶力低下、などに代表される老化の他、物理的外傷や、組織が化学物質にさらされることにより生じる化学的外傷、などが挙げられる。また、組織の物理的又は生理的欠陥に起因した傷害部あるいは障害部も含まれる。また、前記老化の症状は、組織の萎縮、線維化、細胞数の減少、色素沈着、寿命を短縮させるような組織のあるいは全身の変化などであってもよい。
前記組織異常の部位としては、食道、胃、膵臓、肝臓、脾臓、腎臓、小腸、十二指腸、大腸、皮膚、眼、耳、鼻腔、気管、肺、血管、脳、骨、筋肉、咽頭、神経(脊髄、末梢神経等)が挙げられる。
前記組織異常は、加齢に伴って発生するあるいは進行する組織異常であってもよい。また、前記医薬組成物の投与は、加齢に伴う前記組織異常の発生あるいは進行を抑えるものであってもよい。
Examples of the tissue abnormalities include aging represented by decreased visual acuity, decreased hearing, decreased muscle strength, decreased bone density, decreased renal function, decreased liver function, decreased pancreatic function, decreased number of neurons, decreased memory ability, etc. Examples include trauma and chemical trauma caused by exposure of tissues to chemical substances. Further, an injured part or a damaged part due to a physical or physiological defect of the tissue is also included. The aging symptom may be tissue atrophy, fibrosis, a decrease in the number of cells, pigmentation, a tissue change or systemic change that shortens the life span, and the like.
Examples of the tissue abnormalities include esophagus, stomach, pancreas, liver, spleen, kidney, small intestine, duodenum, large intestine, skin, eye, ear, nasal cavity, trachea, lung, blood vessel, brain, bone, muscle, pharynx, nerve ( Spinal cord, peripheral nerve, etc.).
The tissue abnormality may be a tissue abnormality that occurs or progresses with aging. Moreover, administration of the pharmaceutical composition may suppress the occurrence or progression of the tissue abnormality associated with aging.
上記の方法において、組織異常の治療は、内在性の幹細胞の能力に基づいて達成されるものであってもよい。本開示に係る医薬組成物は、サイトカインを始めとする種々の成分を含んでおり、これらの成分は内在性の幹細胞の能力を刺激して、組織異常の治療を達成することが可能だからである。 In the above method, treatment of tissue abnormality may be achieved based on the ability of endogenous stem cells. This is because the pharmaceutical composition according to the present disclosure includes various components including cytokines, and these components can stimulate the ability of endogenous stem cells to achieve treatment of tissue abnormalities. .
本開示によれば、また、iPS細胞を培養することによって得られたiPS細胞培養上清の、医薬組成物の製造における使用も提供される。前記iPS細胞培養上清、前記医薬組成物および使用方法の詳細については、前述の説明(本開示に係る医薬組成物およびその製造方法、ならびに本開示に係る各種治療方法の説明等を参照)のとおりである。 According to the present disclosure, use of an iPS cell culture supernatant obtained by culturing iPS cells in the production of a pharmaceutical composition is also provided. For details of the iPS cell culture supernatant, the pharmaceutical composition and the method of use, refer to the above description (see the description of the pharmaceutical composition according to the present disclosure and the production method thereof, and various treatment methods according to the present disclosure). It is as follows.
本開示によれば、iPS細胞を培養することによって得られたiPS細胞培養上清を含む化粧品も提供できる。iPS細胞培養上清およびその製法の詳細については、前記の医薬組成物およびその製法の詳細と同様である。 According to the present disclosure, a cosmetic containing an iPS cell culture supernatant obtained by culturing iPS cells can also be provided. The details of the iPS cell culture supernatant and the production method thereof are the same as the details of the pharmaceutical composition and the production method.
前記化粧品の製造においても、上述のとおり、iPS細胞の培養後に、細胞成分を分離除去することによって、iPS細胞の培養上清を得ることができるが、培養上清は、培養液から細胞成分を分離除去した上清そのもののみを指すわけではなく、各種処理(例えば、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩、保存等)を適宜施した培養上清であってもよい。培養上清の処理方法の詳細は、前記医薬組成物における培養上清の処理方法として前述したとおりである。 Also in the production of the cosmetic product, as described above, the culture supernatant of iPS cells can be obtained by separating and removing the cell components after culturing iPS cells. It is not limited to the separated supernatant itself, but on cultures that have been appropriately subjected to various treatments (eg, centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, storage, etc.) It may be Qing. The details of the culture supernatant treatment method are as described above as the culture supernatant treatment method in the pharmaceutical composition.
本開示に係る化粧品は、上記により得られたiPS細胞の培養上清を有効成分として含むものであり、ある実施形態においては、前記化粧品は組成物全体としても前記iPS細胞を含まない。また、別の実施形態では、前記化粧品は組成物全体としても細胞(細胞の種類は問わない)を含まない。つまり、無細胞である。 The cosmetic according to the present disclosure contains the culture supernatant of iPS cells obtained as described above as an active ingredient, and in one embodiment, the cosmetic does not contain the iPS cells as a whole composition. Moreover, in another embodiment, the said cosmetics do not contain a cell (the kind of cell is not ask | required) as the whole composition. That is, it is cell-free.
適用される被検体の状態に応じて、期待される治療効果が維持されることを条件として、本開示に係る医薬組成物は他の成分を追加的に含んでもよい。追加的に含まれ得る成分の例としては、エモリエント剤、界面活性助剤、外用鎮痛剤、角質柔軟剤、可塑剤、滑沢剤、可溶化剤、還元剤、緩衝剤、忌避剤、殺虫剤、起泡剤、吸着剤、キレート剤、結合剤、減粘剤研磨・スクラブ剤、抗アクネ剤、抗う蝕剤、抗黴剤、抗菌剤、口腔衛生剤、口腔ケア剤、抗ケーキング剤、酵素剤、抗フケ剤、香味剤、香料、殺菌剤、酸化剤、酸化防止剤、紫外線吸収剤・散乱剤、収れん剤、消臭剤、消泡剤、人工爪剤、親水性増粘剤、制汗剤、洗浄剤、増量剤、褪色防止剤、帯電防止剤、脱毛剤、着色剤、爪コンディショニング剤、乳化安定剤、乳化剤、粘着剤、パーマネント・ウェーブ用還元剤、剥離剤、非活性剤系分散剤、非水系増粘剤、皮膚コンディショニング剤、皮膚ブリーチ剤、皮膚保護剤、皮膜形成剤、表面改質剤、腐蝕防止剤、物理的脱毛剤、不透明化剤、分散剤、噴射剤、ヘアコンディショニング剤、ヘアスタイリング剤、閉塞剤、pH調整剤、変性剤、防腐剤、保湿・湿潤剤、保水剤、毛髪着色剤、口腔ヘルスケア用医薬品、および皮膚防御剤溶剤などが挙げられる。 Depending on the condition of the subject to be applied, the pharmaceutical composition according to the present disclosure may additionally contain other components, provided that the expected therapeutic effect is maintained. Examples of ingredients that can be additionally included include emollients, surfactants, external analgesics, keratin softeners, plasticizers, lubricants, solubilizers, reducing agents, buffers, repellents, insecticides , Foaming agent, adsorbent, chelating agent, binder, thinning agent polishing / scrub agent, anti-acne agent, anti-cariogenic agent, antidepressant agent, antibacterial agent, oral hygiene agent, oral care agent, anti-caking agent, enzyme Agent, anti-dandruff agent, flavoring agent, fragrance, bactericidal agent, oxidizing agent, antioxidant, ultraviolet absorber / scattering agent, astringent, deodorant, antifoaming agent, artificial nail agent, hydrophilic thickener, antibacterial agent Sweat agent, cleaning agent, extender, anti-fading agent, antistatic agent, hair remover, colorant, nail conditioning agent, emulsifying stabilizer, emulsifier, adhesive, permanent wave reducing agent, release agent, inactive agent system Dispersant, non-aqueous thickener, skin conditioning agent, skin bleach, skin protectant, film form Agent, surface modifier, corrosion inhibitor, physical hair remover, opacifier, dispersant, propellant, hair conditioning agent, hair styling agent, occlusive agent, pH adjuster, denaturant, antiseptic, moisturizing / wetting Agents, water retention agents, hair coloring agents, oral health care drugs, and skin protection solvents.
本開示に係る化粧品の最終的な形態は特に限定されない。形態の例は、液状、乳液状、クリーム状、軟膏状、スプレー状、ゲル状などである。 The final form of the cosmetic according to the present disclosure is not particularly limited. Examples of forms are liquid, emulsion, cream, ointment, spray, gel and the like.
本開示においては、
(1)iPS細胞を培養するステップ;
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ。
を含む、化粧品の製造方法も提供される。この製造方法は、前記回収された培養上清に遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、および脱塩の中から選択される一つ以上の処理を施すステップをさらに含んでもよい。また、前記製造方法は、前記回収された培養上清に、追加の成分を添加するステップをさらに含んでいてもよい。また、前記製造方法は、体細胞に初期化因子を導入して前記iPS細胞を作成するステップをさらに含んでいてもよい。各々のステップについては、本開示に係る医薬組成物の説明において記載した事項がそのまま当てはまる。追加的成分については、本開示に係る化粧品の説明において記載した事項がそのまま当てはまる。前記回収された培養上清に遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、および脱塩の中から選択される一つ以上の処理を施すステップと、前記回収された培養上清に、追加の成分を添加するステップの両方を含む場合には、両ステップはどちらを先に行ってもよく、また可能な場合には同時並行して行ってもよい。
In this disclosure,
(1) culturing iPS cells;
(2) A step of recovering the culture supernatant obtained by culturing the iPS cells.
A method for producing a cosmetic product is also provided. In this production method, the collected culture supernatant is subjected to one or more treatments selected from centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, and desalting. May further be included. The production method may further include a step of adding an additional component to the collected culture supernatant. The production method may further include a step of creating the iPS cell by introducing an reprogramming factor into a somatic cell. About each step, the matter described in description of the pharmaceutical composition based on this indication is applied as it is. For the additional components, the matters described in the description of the cosmetic product according to the present disclosure are applied as they are. Subjecting the collected culture supernatant to one or more treatments selected from centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, and desalting; When the culture supernatant includes both steps of adding additional components, both steps may be performed first, and if possible, may be performed simultaneously.
本開示によれば、本開示に係る化粧品を対象に適用することにより対象の美容的外観を改善する美容方法、も提供される。
対象は、例えばヒトであり、対象組織は例えば、皮膚や毛髪である。前記化粧品は、例えば、顔や首、腕、足といった部位に適用できる。また、火傷や外傷の痕を消すために、こうした部位に適用することも可能である。
適用は、一日一回〜三回程度行ってもよく、あるいは、ディフューザーなどを使用して霧状に噴霧することで、連続して適用することも可能である。
According to the present disclosure, a cosmetic method for improving the cosmetic appearance of a subject by applying the cosmetic according to the present disclosure to the subject is also provided.
The target is, for example, a human, and the target tissue is, for example, skin or hair. The cosmetic can be applied to parts such as the face, neck, arms, and feet, for example. It can also be applied to these areas to eliminate burns and trauma marks.
The application may be performed once to three times a day, or may be continuously applied by spraying in a mist using a diffuser or the like.
本開示に係る化粧品は、iPS細胞培養上清を含むため、例えば皮膚のしわの原因となるコラーゲンやエラスチン組織への傷害などの、美容的外観を損なう変化を修復する作用を示す。これによって、本開示に係る化粧品は、対象における美容的外観に関係する部位に適用されることで、対象の美容的外観を改善する効果を示すものと考えられる。
本開示に係る化粧品は、加齢に伴って発生するあるいは進行する美容上の欠陥、例えばしわ、たるみ、くすみ、シミ、などの発生あるいは悪化を抑えるものであってもよい。
Since the cosmetic product according to the present disclosure includes the iPS cell culture supernatant, the cosmetic product exhibits an effect of repairing changes that impair the cosmetic appearance, such as injury to collagen and elastin tissue that cause skin wrinkles. Thus, the cosmetic product according to the present disclosure is considered to exhibit an effect of improving the cosmetic appearance of the target by being applied to a part related to the cosmetic appearance of the target.
The cosmetic product according to the present disclosure may be one that suppresses the occurrence or deterioration of cosmetic defects, such as wrinkles, sagging, dullness, and spots, that occur or progress with aging.
上記のとおり、iPS細胞を培養することによって得られたiPS細胞培養上清は、加齢に伴う疾患、組織異常、美容上の欠陥などの発生または進行を幅広く抑えることが出来る。このため、本開示によれば、iPS細胞を培養することによって得られたiPS培養上清を含む、抗加齢組成物も提供できる。抗加齢組成物に含まれるiPS培養上清について、その製法、由来、予備的処理、形態、含有量などの詳細については、上記の医薬組成物についての説明において説明された内容と同様である。加齢の発現は、組織の萎縮、線維化、細胞数の減少、色素沈着、寿命を短縮させるような組織のあるいは全身の変化などであってもよい。また、対象に前記抗加齢組成物を、抗加齢作用を発揮するのに有効な量投与することを含む、対象における加齢を抑制する方法も提供される。対象への前記抗加齢組成物の投与は、対象の寿命を延長するものであってもよい。 As described above, the iPS cell culture supernatant obtained by culturing iPS cells can widely suppress the occurrence or progression of aging-related diseases, tissue abnormalities, cosmetic defects, and the like. Therefore, according to the present disclosure, an anti-aging composition containing an iPS culture supernatant obtained by culturing iPS cells can also be provided. About the iPS culture supernatant contained in the anti-aging composition, the details of the production method, origin, preliminary treatment, form, content and the like are the same as the contents explained in the explanation of the pharmaceutical composition above. . The expression of aging may be tissue atrophy, fibrosis, cell number decrease, pigmentation, tissue or systemic changes that shorten life span, and the like. Also provided is a method for inhibiting aging in a subject, comprising administering to the subject an amount of the anti-aging composition effective to exert an anti-aging effect. Administration of the anti-aging composition to a subject may extend the life of the subject.
本発明の実施形態には以下のものが含まれる。
<1> iPS細胞を培養することによって得られたiPS細胞培養上清を含む、医薬組成物。
<2> 前記iPS細胞を含まない、<1>に記載の医薬組成物。
<3> 無細胞である、<1>または<2>に記載の医薬組成物。
<4> 前記iPS細胞培養上清が、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群から選択される1以上によって処理されたものである、<1>〜<3>のいずれか一項に記載の医薬組成物。
<5> 前記iPS細胞が、骨髄由来間葉系幹細胞に由来するものである、<1>〜<4>のいずれか一項に記載の医薬組成物。
<6> 血清を含まない、<1>〜<5>のいずれか一項に記載の医薬組成物。
Embodiments of the present invention include the following.
<1> A pharmaceutical composition comprising an iPS cell culture supernatant obtained by culturing iPS cells.
<2> The pharmaceutical composition according to <1>, which does not include the iPS cell.
<3> The pharmaceutical composition according to <1> or <2>, which is cell-free.
<4> The iPS cell culture supernatant is treated with one or more selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting and storage. The pharmaceutical composition according to any one of <1> to <3>.
<5> The pharmaceutical composition according to any one of <1> to <4>, wherein the iPS cells are derived from bone marrow-derived mesenchymal stem cells.
<6> The pharmaceutical composition according to any one of <1> to <5>, which does not contain serum.
<7> 対象において疾患または組織異常の発症前に前記疾患または組織異常の発症を抑えるために用いられる、<1>〜<6>のいずれか一項に記載の医薬組成物。
<8> 前記対象が、前記疾患または組織異常を発病するリスクを有すると判定された対象である、<7>に記載の医薬組成物。
<9> 前記疾患が、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患、神経難病、歯周病、皮膚潰瘍、および骨粗鬆症からなる群から選択される、<7>または<8>に記載の医薬組成物。
<10> 前記組織異常が老化である、<7>〜<9>のいずれか一項に記載の医薬組成物。
<11> 前記疾患や組織異常が特定の疾患や組織異常に限定されない、ユニバーサルな医薬組成物である、<7>に記載の医薬組成物。
<12> 疾患を治療するために用いられる、<1>〜<6>のいずれか一項に記載の医薬組成物。
<13> 前記疾患が、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患、神経難病、歯周病、皮膚潰瘍、および骨粗鬆症からなる群から選択される、<12>に記載の医薬組成物。
<7> The pharmaceutical composition according to any one of <1> to <6>, which is used to suppress the onset of the disease or tissue abnormality before the onset of the disease or tissue abnormality in the subject.
<8> The pharmaceutical composition according to <7>, wherein the subject is a subject determined to have a risk of developing the disease or tissue abnormality.
<9> The above diseases are gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related changes in heart, hypertension, ischemic heart disease, valvular disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, lung Fibrosis, Langerhans Island atrophy, liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases, cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. <7> or <8>, wherein the pharmaceutical composition is selected from the group consisting of dropout, periventricular leukomalacia, retinal disease with neuronal damage, intractable neuropathy, periodontal disease, skin ulcer, and osteoporosis object.
<10> The pharmaceutical composition according to any one of <7> to <9>, wherein the tissue abnormality is aging.
<11> The pharmaceutical composition according to <7>, which is a universal pharmaceutical composition in which the disease or tissue abnormality is not limited to a specific disease or tissue abnormality.
<12> The pharmaceutical composition according to any one of <1> to <6>, which is used for treating a disease.
<13> The disease is gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related changes in heart, hypertension, ischemic heart disease, valvular disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, lung Fibrosis, Langerhans Island atrophy, liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases, cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. <12> The pharmaceutical composition according to <12>, selected from the group consisting of dropout, periventricular leukomalacia, retinal disease associated with nerve cell damage, intractable nerve disease, periodontal disease, skin ulcer, and osteoporosis.
<14> 組織異常を治療するために用いられる、<1>〜<6>のいずれか一項に記載の医薬組成物。
<15> 前記組織異常が老化である、<14>に記載の医薬組成物。
<16> (1)iPS細胞を培養するステップ;および
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、医薬組成物の製造方法。
<17> 体細胞に初期化因子を導入して前記iPS細胞を作成するステップをさらに含む、<16>に記載の方法。
<18> 前記回収した培養上清に対して、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群より選択される少なくとも1の処理を行うステップをさらに含む、<16>または<17>に記載の方法。
<19> 前記回収した培養上清に追加的成分を添加するステップをさらに含む、<16>〜<18>のいずれか一項に記載の方法。
<20> iPS細胞を培養することによって得られたiPS細胞培養上清の、医薬組成物の製造における使用。
<14> The pharmaceutical composition according to any one of <1> to <6>, which is used for treating a tissue abnormality.
<15> The pharmaceutical composition according to <14>, wherein the tissue abnormality is aging.
<16> (1) a step of culturing iPS cells; and (2) a step of collecting a culture supernatant obtained by culturing the iPS cells,
A method for producing a pharmaceutical composition, comprising:
<17> The method according to <16>, further comprising a step of introducing the reprogramming factor into the somatic cell to produce the iPS cell.
<18> The collected culture supernatant is subjected to at least one treatment selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, and storage. The method according to <16> or <17>, further comprising a step.
<19> The method according to any one of <16> to <18>, further comprising a step of adding an additional component to the collected culture supernatant.
<20> Use of an iPS cell culture supernatant obtained by culturing iPS cells in the production of a pharmaceutical composition.
<21> 前記医薬組成物が前記iPS細胞を含まない、<20>に記載の使用。
<22> 前記医薬組成物が無細胞である、<20>または<21>に記載の使用。
<23> 前記iPS細胞培養上清が、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群から選択される1以上によって処理されたものである、<20>〜<22>のいずれか一項に記載の使用。
<24> 前記iPS細胞が、骨髄由来間葉系幹細胞に由来するものである、<20>〜<23>のいずれか一項に記載の使用。
<25> 前記医薬組成物が血清を含まない、<20>〜<24>のいずれか一項に記載の使用。
<26> 前記医薬組成物が、対象において疾患または組織異常の発症前に前記疾患または組織異常の発症を抑えるために用いられる、<20>〜<25>のいずれか一項に記載の使用。
<27> 前記対象が、前記疾患または組織異常を発病するリスクを有すると判定された対象である、<26>に記載の使用。
<21> The use according to <20>, wherein the pharmaceutical composition does not contain the iPS cell.
<22> The use according to <20> or <21>, wherein the pharmaceutical composition is cell-free.
<23> The iPS cell culture supernatant is treated with one or more selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, and storage. The use according to any one of <20> to <22>.
<24> The use according to any one of <20> to <23>, wherein the iPS cells are derived from bone marrow-derived mesenchymal stem cells.
<25> The use according to any one of <20> to <24>, wherein the pharmaceutical composition does not contain serum.
<26> The use according to any one of <20> to <25>, wherein the pharmaceutical composition is used for suppressing the onset of the disease or tissue abnormality before the onset of the disease or tissue abnormality in the subject.
<27> The use according to <26>, wherein the subject is a subject determined to have a risk of developing the disease or tissue abnormality.
<28> 前記疾患が、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患、神経難病、歯周病、皮膚潰瘍、および骨粗鬆症からなる群から選択される、<26>または<27>に記載の使用。
<29> 前記組織異常が老化である、<26>〜<28>のいずれか一項に記載の使用。
<30> 前記疾患や組織異常が特定の疾患や組織異常に限定されず、前記医薬組成物がユニバーサルな医薬組成物である、<26>に記載の使用。
<31> 前記医薬組成物が、疾患を治療するために用いられる、<20>〜<25>のいずれか一項に記載の使用。
<32> 前記疾患が、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患、神経難病、歯周病、皮膚潰瘍、および骨粗鬆症からなる群から選択される、<31>に記載の使用。
<33> 前記医薬組成物が、組織異常を治療するために用いられる、<20>〜<25>のいずれか一項に記載の使用。
<34> 前記組織異常が老化である、<33>に記載の使用。
<28> The above diseases are gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, aging change of heart, hypertension, ischemic heart disease, valvular heart disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, lung Fibrosis, Langerhans Island atrophy, liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases, cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. The use according to <26> or <27>, which is selected from the group consisting of dropout, periventricular leukomalacia, retinal disease with neuronal damage, intractable neuropathy, periodontal disease, skin ulcer, and osteoporosis.
<29> The use according to any one of <26> to <28>, wherein the tissue abnormality is aging.
<30> The use according to <26>, wherein the disease or tissue abnormality is not limited to a specific disease or tissue abnormality, and the pharmaceutical composition is a universal pharmaceutical composition.
<31> The use according to any one of <20> to <25>, wherein the pharmaceutical composition is used for treating a disease.
<32> The disease is gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related changes in heart, hypertension, ischemic heart disease, valvular disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, lung Fibrosis, Langerhans Island atrophy, liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases, cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. The use according to <31>, which is selected from the group consisting of dropout, periventricular leukomalacia, retinal disease with neuronal damage, intractable neuropathy, periodontal disease, skin ulcer, and osteoporosis.
<33> The use according to any one of <20> to <25>, wherein the pharmaceutical composition is used for treating a tissue abnormality.
<34> The use according to <33>, wherein the tissue abnormality is aging.
<35> <1>〜<6>のいずれか一項に記載の医薬組成物を、疾患または組織異常の発症前の対象に、前記疾患または組織異常の発症を抑えるために有効な量投与することを含む、対象において疾患または組織異常の発症前に前記疾患または組織異常の発症を抑える方法。
<36> 前記対象が、前記疾患または組織異常を発病するリスクを有すると判定された対象である、<35>に記載の方法。
<37> 前記疾患が、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患、神経難病、歯周病、皮膚潰瘍、および骨粗鬆症からなる群から選択される、<35>または<36>に記載の方法。
<38> 前記組織異常が老化である、<35>〜<37>のいずれか一項に記載の方法。
<39> 前記疾患や組織異常が特定の疾患や組織異常に限定されず、前記医薬組成物がユニバーサルな医薬組成物である、<35>に記載の方法。
<40> 前記疾患または組織異常の発症抑制が、内在性の幹細胞の能力に基づいて達成される、<35>〜<39>のいずれか一項に記載の方法。
<35> The pharmaceutical composition according to any one of <1> to <6> is administered to a subject before the onset of the disease or tissue abnormality in an amount effective to suppress the onset of the disease or tissue abnormality. A method of suppressing the onset of the disease or tissue abnormality before the onset of the disease or tissue abnormality in the subject.
<36> The method according to <35>, wherein the subject is a subject determined to have a risk of developing the disease or tissue abnormality.
<37> The disease is gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related changes in heart, hypertension, ischemic heart disease, valvular disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, lung Fibrosis, Langerhans Island atrophy, liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases, cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. <35> or <36>, wherein the method is selected from the group consisting of dropout, periventricular leukomalacia, retinal disease with neuronal damage, intractable nerve disease, periodontal disease, skin ulcer, and osteoporosis.
<38> The method according to any one of <35> to <37>, wherein the tissue abnormality is aging.
<39> The method according to <35>, wherein the disease or tissue abnormality is not limited to a specific disease or tissue abnormality, and the pharmaceutical composition is a universal pharmaceutical composition.
<40> The method according to any one of <35> to <39>, wherein the onset suppression of the disease or tissue abnormality is achieved based on the ability of endogenous stem cells.
<41> <1>〜<6>のいずれか一項に記載の医薬組成物を、疾患を有する対象に、前記疾患を治療するために有効な量で投与することを含む、疾患の治療方法。
<42> 前記疾患が、胃潰瘍、肝硬変、肝不全、胆石、胆道炎、黄疸、心臓の加齢変化、高血圧、虚血性心疾患、心臓弁膜疾患、不整脈、心不全、動脈硬化、肺炎、肺気腫、肺線維症、ランゲルハンス島萎縮、肝線維症、慢性気管支炎、脳卒中、脳虚血、脳梗塞、認知症、てんかん、神経変性疾患、脳虚血や脳内出血等に伴う脳梗塞による神経細胞の変性・脱落、脳室周囲白質軟化症、神経細胞の障害を伴う網膜疾患、神経難病、歯周病、皮膚潰瘍、および骨粗鬆症からなる群から選択される、<41>に記載の方法。
<43> 前記疾患の治療が、内在性の幹細胞の能力に基づいて達成される、<41>または<42>に記載の方法。
<44> <1>〜<6>のいずれか一項に記載の医薬組成物を、組織異常を有する対象に、前記組織異常を治療するために有効な量で投与することを含む、組織異常の治療方法。
<45> 前記組織異常が老化である、<44>に記載の方法。
<46> 前記組織異常の治療が、内在性の幹細胞の能力に基づいて達成される、<45>に記載の方法。
<47>
前記医薬組成物の投与が、静脈内投与、動脈内投与、門脈内投与、皮内投与、皮下投与、筋肉内投与、腹腔内投与、経肺投与及び鼻腔内投与からなる群より選択された方法により行われる、<41>〜<46>のいずれか一項に記載の方法。
<41> A method for treating a disease, comprising administering the pharmaceutical composition according to any one of <1> to <6> to a subject having a disease in an amount effective for treating the disease. .
<42> The disease is gastric ulcer, cirrhosis, liver failure, gallstone, cholangitis, jaundice, age-related change of heart, hypertension, ischemic heart disease, valvular disease, arrhythmia, heart failure, arteriosclerosis, pneumonia, emphysema, lung Fibrosis, Langerhans Island atrophy, liver fibrosis, chronic bronchitis, stroke, cerebral ischemia, cerebral infarction, dementia, epilepsy, neurodegenerative diseases, cerebral infarction associated with cerebral ischemia or intracerebral hemorrhage, etc. <41> The method according to <41>, which is selected from the group consisting of dropout, periventricular leukomalacia, retinal disease associated with nerve cell damage, intractable nerve disease, periodontal disease, skin ulcer, and osteoporosis.
<43> The method according to <41> or <42>, wherein the treatment of the disease is achieved based on the ability of endogenous stem cells.
<44> A tissue abnormality comprising administering the pharmaceutical composition according to any one of <1> to <6> to a subject having a tissue abnormality in an amount effective for treating the tissue abnormality. Treatment methods.
<45> The method according to <44>, wherein the tissue abnormality is aging.
<46> The method according to <45>, wherein the treatment of the tissue abnormality is achieved based on the ability of endogenous stem cells.
<47>
The administration of the pharmaceutical composition was selected from the group consisting of intravenous administration, intraarterial administration, intraportal administration, intradermal administration, subcutaneous administration, intramuscular administration, intraperitoneal administration, transpulmonary administration, and intranasal administration. The method according to any one of <41> to <46>, which is performed by a method.
<48> 前記医薬組成物の投与が、経肺投与または鼻腔内投与にてより行われる<41>〜<47>のいずれか一項に記載の方法。
<49> iPS細胞を培養することによって得られたiPS細胞培養上清を含む、化粧品。
<50> 前記iPS細胞を含まない、<49>に記載の化粧品。
<51>
無細胞である、<49>または<50>に記載の化粧品。
<52>
前記iPS細胞培養上清が、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群から選択される1以上によって処理されたものである、<49>〜<51>のいずれか一項に記載の化粧品。
<53>
前記iPS細胞が、骨髄由来間葉系幹細胞に由来するものである、<49>〜<52>のいずれか一項に記載の化粧品。
<54>
血清を含まない、<49>〜<53>のいずれか一項に記載の化粧品。
<55>
シミ、しわ、たるみおよびくすみからなる群から選択される皮膚の状態を改善するために用いられる、<49>〜<54>のいずれか一項に記載の化粧品。
<48> The method according to any one of <41> to <47>, wherein the pharmaceutical composition is administered by pulmonary administration or intranasal administration.
<49> A cosmetic comprising an iPS cell culture supernatant obtained by culturing iPS cells.
<50> The cosmetic according to <49>, which does not include the iPS cell.
<51>
The cosmetic according to <49> or <50>, which is cell-free.
<52>
The iPS cell culture supernatant is treated with one or more selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, and storage. Cosmetics as described in any one of 49>-<51>.
<53>
The cosmetic according to any one of <49> to <52>, wherein the iPS cells are derived from bone marrow-derived mesenchymal stem cells.
<54>
The cosmetic according to any one of <49> to <53>, which does not contain serum.
<55>
The cosmetic according to any one of <49> to <54>, which is used for improving a skin condition selected from the group consisting of spots, wrinkles, sagging and dullness.
<56>
<49>〜<54>のいずれか一項に記載の化粧品を対象に適用することにより、対象の美容的外観を改善する、美容方法。
<57>
前記美容的外観の改善が、内在性の幹細胞の能力に基づいて達成される、<56>に記載の美容方法。
<58>
前記化粧品が対象の皮膚に塗布され、シミ、しわ、たるみおよびくすみからなる群から選択される皮膚の状態を改善する、<56>または<57>に記載の美容方法。
<59>
(1)iPS細胞を培養するステップ;および
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、化粧品の製造方法。
<60>
体細胞に初期化因子を導入して前記iPS細胞を作成するステップをさらに含む、<59>に記載の方法。
<56>
<49>-<54> The cosmetic method which improves the cosmetic appearance of object by applying the cosmetics as described in any one to object.
<57>
The cosmetic method according to <56>, wherein the improvement of the cosmetic appearance is achieved based on the ability of endogenous stem cells.
<58>
The cosmetic method according to <56> or <57>, wherein the cosmetic is applied to the subject's skin to improve the skin condition selected from the group consisting of spots, wrinkles, sagging and dullness.
<59>
(1) culturing iPS cells; and (2) collecting the culture supernatant obtained by culturing the iPS cells;
A method for producing cosmetics, comprising:
<60>
The method according to <59>, further comprising the step of introducing the reprogramming factor into the somatic cell to produce the iPS cell.
<61>
前記回収した培養上清に対して、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群より選択される少なくとも1の処理を行うステップをさらに含む、<59>または<60>に記載の方法。
<62>
前記回収した培養上清に追加的成分を添加するステップをさらに含む、<59>〜<61>のいずれか一項に記載の方法。
<63> iPS細胞を培養することによって得られたiPS細胞培養上清を含む、抗加齢組成物。
<64> 前記iPS細胞を含まない、<63>に記載の抗加齢組成物。
<65> 無細胞である、<63>または<64>に記載の抗加齢組成物。
<66> 前記iPS細胞培養上清が、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群から選択される1以上によって処理されたものである、<63>〜<65>のいずれか一項に記載の抗加齢組成物。
<67> 前記iPS細胞が、骨髄由来間葉系幹細胞に由来するものである、<63>〜<66>のいずれか一項に記載の抗加齢組成物。
<68> 血清を含まない、<63>〜<67>のいずれか一項に記載の抗加齢組成物。
<69> (1)iPS細胞を培養するステップ;および
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、抗加齢組成物の製造方法。
<70> 体細胞に初期化因子を導入して前記iPS細胞を作成するステップをさらに含む、<69>に記載の方法。
<71> 前記回収した培養上清に対して、遠心処理、濃縮、溶媒の置換、透析、凍結、乾燥、凍結乾燥、希釈、脱塩及び保存からなる群より選択される少なくとも1の処理を行うステップをさらに含む、<69>または<70>に記載の方法。
<72> 前記回収した培養上清に追加的成分を添加するステップをさらに含む、<69>〜<71>のいずれか一項に記載の方法。
<73> 対象に<63>〜<68>のいずれか一項に記載の抗加齢組成物を、抗加齢作用を発揮するのに有効な量投与することを含む、対象における加齢を抑制する方法。
<61>
The recovered culture supernatant is further subjected to at least one treatment selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting and storage. The method according to <59> or <60>.
<62>
The method according to any one of <59> to <61>, further comprising a step of adding an additional component to the collected culture supernatant.
<63> An anti-aging composition comprising an iPS cell culture supernatant obtained by culturing iPS cells.
<64> The anti-aging composition according to <63>, which does not include the iPS cell.
<65> The anti-aging composition according to <63> or <64>, which is cell-free.
<66> The iPS cell culture supernatant is treated with one or more selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, and storage. The anti-aging composition according to any one of <63> to <65>.
<67> The anti-aging composition according to any one of <63> to <66>, wherein the iPS cells are derived from bone marrow-derived mesenchymal stem cells.
<68> The anti-aging composition according to any one of <63> to <67>, which does not contain serum.
<69> (1) a step of culturing iPS cells; and (2) a step of collecting a culture supernatant obtained by culturing the iPS cells.
The manufacturing method of an anti-aging composition containing this.
<70> The method according to <69>, further comprising a step of introducing the reprogramming factor into the somatic cell to produce the iPS cell.
<71> The collected culture supernatant is subjected to at least one treatment selected from the group consisting of centrifugation, concentration, solvent replacement, dialysis, freezing, drying, lyophilization, dilution, desalting, and storage. The method according to <69> or <70>, further comprising a step.
<72> The method according to any one of <69> to <71>, further comprising a step of adding an additional component to the collected culture supernatant.
<73> Aging in a subject comprising administering to the subject an anti-aging composition according to any one of <63> to <68> in an amount effective to exert an anti-aging effect. How to suppress.
以下に本発明の実施例について説明するが、これに限定されるものではない。また実施例中の%は、特に断らない限り、重量(質量)基準である。 Examples of the present invention will be described below, but the present invention is not limited thereto. Further,% in the examples is based on weight (mass) unless otherwise specified.
実施例1:老化促進モデルマウスにおける種々の組織における先制医療
老化促進モデルマウス(Senescence Accelerated Mouse:SAM)を用いて、本開示に係るiPS細胞培養上清を投与した場合と投与しない場合における疾患および組織変性における差異を調べた。
具体的には、下記のようにして、ヒトiPS細胞の培養上清を得た。なお、下記において培養はいずれも37℃で行った。
(iPS細胞)
iPS細胞の作製元となる細胞としてヒト骨髄由来間葉系細胞を用いた以外は、Takahashi K, et al. "Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors" Cell. 131(5): 861-72 (2007)に記載された方法に従い作成されたiPS細胞を、国立大学法人京都大学から入手した。具体的には、ヒトOct3/4、Sox2、Klf4およびc−Mycを含む、レトロウィルス(pMXベクター)をヒト骨髄由来間葉系細胞に導入し、導入6日後に細胞をトリプシン処理し、マイトマイシンCで処理されたSNLフィーダー細胞上にプレーティングした(McMahon and Bradley, 1990)。翌日、培地を10%FBS含有DMEMから、4ng/mlのbFGFを添加した霊長類ES細胞用培地(primate ES cell culture、株式会社リプロセル製)に変えた。
約2週間後に、ヒトES細胞様の形態を有する球状コロニーが現れた。約25日目に平坦で、ヒトES細胞に類似したコロニーが観察された。各細胞の形態はヒトES細胞のものに類似し、コロニーの中央においては時折自発的な分化も見られた。また、フィーダー細胞依存性を有していたり、マトリゲル被覆プレート上マウス胚性線維芽細胞(MEF)馴化霊長類ES細胞用培地では未分化な状態を保つものの、無調整培地においては未分化状態を保たないといった点においても、得られた細胞はヒトES細胞に類似した性質を示した。
こうして得られたヒトiPS細胞を用いて、iPS細胞培養上清の作製を行った。
Example 1: Preemptive medicine in various tissues in senescence-accelerated model mice Diseases with and without administration of the iPS cell culture supernatant according to the present disclosure using senescence-accelerated mouse (SAM) Differences in tissue degeneration were examined.
Specifically, a culture supernatant of human iPS cells was obtained as follows. In the following, culture was performed at 37 ° C.
(IPS cells)
Except for using human bone marrow-derived mesenchymal cells as a source of iPS cells, Takahashi K, et al. "Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors" Cell. 131 (5): 861 -PS (2007), iPS cells prepared according to the method described in the above were obtained from Kyoto University. Specifically, a retrovirus (pMX vector) containing human Oct3 / 4, Sox2, Klf4 and c-Myc was introduced into human bone marrow-derived mesenchymal cells, 6 days after the introduction, the cells were trypsinized, and mitomycin C Were plated on SNL feeder cells treated with (McMahon and Bradley, 1990). On the next day, the medium was changed from 10% FBS-containing DMEM to a primate ES cell culture medium (primate ES cell culture, manufactured by Reprocell) supplemented with 4 ng / ml bFGF.
After about 2 weeks, spherical colonies having human ES cell-like morphology appeared. On day 25, colonies that were flat and similar to human ES cells were observed. The morphology of each cell was similar to that of human ES cells, and spontaneous differentiation was sometimes seen in the center of the colony. In addition, although it has feeder cell dependency or remains undifferentiated in the mouse embryonic fibroblast (MEF) primate ES cell culture medium on the matrigel-coated plate, it remains undifferentiated in the unconditioned medium. The obtained cells showed similar properties to human ES cells in that they were not retained.
The iPS cell culture supernatant was prepared using the human iPS cells thus obtained.
(培養上清の調製)
前記ヒトiPS細胞は接着培養用シャーレで接着培養することにより維持した。培養に際しては、4ng/mlのヒト塩基性FGF(bFGF)を添加した霊長類ES細胞用培地(無血清;株式会社リプロセル製;以後このbFGF添加霊長類ES細胞用培地を、iPS細胞用培地と称する)を用いた。培養中は1週間毎に継代が必要であるが、これはヒトiPS細胞を0.25%トリプシン、0.1mg/mLのコラゲナーゼIV、1mMのCaCl2および20%のKSRを含む溶液で処理することにより行った。
上記の通り維持されたヒトiPS細胞を、ES細胞解離液(株式会社リプロセル製)を用いて接着培養用シャーレから剥がし、非接着培養用シャーレに入れたiPS細胞用培地中で1週間浮遊培養した。この結果、胚様体(Embryo Body:EB)が形成された。
形成された胚様体(EB)を接着培養用シャーレ上に播種し、10%FBSおよび1%アンチアンチ(登録商標、抗真菌剤)を含有するDMEM中で1週間成長(outgrowth)させた(細胞のoutgrowthについては、Stem Cells Dev. 22, 102-113, 2013を参照)。
次に、上記ヒトiPS細胞を0.05%トリプシン−EDTA溶液を用いて接着培養用シャーレから剥がし、新たな接着培養用シャーレに播種した。これにより、ヒトiPS細胞はsingle cell化した。培地として10%FBSおよび1%アンチアンチ(登録商標、抗真菌剤)を含有するDMEMを用い、一週間培養した。
ヒトiPS細胞が70〜80%以上コンフルエントになったことを確認した後に、培地を無血清培地(FBSを含まない、1%アンチアンチ(登録商標、抗真菌剤)含有DMEM)に置換し、2日間(48時間)培養後、上清を回収した。回収した上清を1500回転で5分遠心し、上清を再度回収した後に3000回転で3分遠心し、上清を再度回収したものを以後の実験においてヒトiPS細胞培養上清として用いた。前記遠心はトミー精工製卓上多本架遠心機LC-120を用いて行った。
(Preparation of culture supernatant)
The human iPS cells were maintained by adhesion culture in a petri dish for adhesion culture. In culture, primate ES cell medium supplemented with 4 ng / ml human basic FGF (bFGF) (serum-free; manufactured by Reprocell Corporation; hereinafter, this bFGF-added primate ES cell medium is referred to as iPS cell medium. Used). During culture, passage is required every week, which involves treating human iPS cells with a solution containing 0.25% trypsin, 0.1 mg / mL collagenase IV, 1 mM CaCl 2 and 20% KSR. It was done by doing.
The human iPS cells maintained as described above were detached from the adhesion culture petri dish using an ES cell dissociation solution (manufactured by Reprocell Co., Ltd.), and suspended in the iPS cell culture medium placed in the non-adhesion culture petri dish for 1 week. . As a result, an embryoid body (EB) was formed.
The formed embryoid bodies (EBs) were seeded on adherent culture dishes and grown for 1 week in DMEM containing 10% FBS and 1% anti-anti (registered trademark, antifungal agent) ( For cell outgrowth, see Stem Cells Dev. 22, 102-113, 2013).
Next, the human iPS cells were detached from the adhesion culture petri dish using a 0.05% trypsin-EDTA solution and seeded in a new adhesion culture petri dish. As a result, human iPS cells became single cells. Using DMEM containing 10% FBS and 1% anti-anti (registered trademark, antifungal agent) as a medium, the cells were cultured for one week.
After confirming that human iPS cells were confluent at 70 to 80% or more, the medium was replaced with serum-free medium (DMEM containing 1% anti-anti (registered trademark, antifungal agent) without FBS). After culturing for one day (48 hours), the supernatant was collected. The collected supernatant was centrifuged at 1500 rpm for 5 minutes, and the supernatant was collected again and then centrifuged at 3000 rpm for 3 minutes. The supernatant was collected again and used as a human iPS cell culture supernatant in subsequent experiments. The centrifugation was performed using a tabletop multi-centrifuge LC-120 manufactured by Tommy Seiko.
SAMP8(6週齢、日本エスエルシー株式会社)を準備した。また、上記で得られたiPS細胞培養上清を精製水にて1%濃度に希釈したものを超音波にて霧状にして、動物ケージ内に充満させた。また、iPS細胞培養上清を用いずに、精製水のみを霧状にして動物ケージ内に充満させた。実験群(雄雌それぞれn=5)に対しては、上記のiPS細胞培養上清含有ミストで充満した動物ケージ内に12時間、次に上記ミストを含まない動物ケージ内に12時間、のサイクルを行った。対照群(雄雌それぞれn=5)に対しては、上記の精製水ミストで充満した動物ケージ内に12時間、次に上記ミストを含まない動物ケージ内に12時間、のサイクルを行った。 SAMP8 (6 weeks old, Nippon SLC Co., Ltd.) was prepared. Further, the iPS cell culture supernatant obtained above was diluted with purified water to a concentration of 1% and atomized with ultrasound to fill the animal cage. Further, without using the iPS cell culture supernatant, only purified water was atomized to fill the animal cage. For the experimental group (n = 5 for each male and female), a cycle of 12 hours in an animal cage filled with the mist containing the above iPS cell culture supernatant and then 12 hours in an animal cage not containing the mist. Went. For the control group (n = 5 for each male and female), a cycle of 12 hours in an animal cage filled with the purified water mist was followed by 12 hours in an animal cage not containing the mist.
実験開始後7週目、40週目および60週目に、竹田俊男によって提唱された老化度判定基準(grading score system;"Grading score system: a method for evaluation of the degree of senescence in senescence accelerated mouse (SAM)" authored by Hosokawa M, Kasai R, Higuchi K, Takeshita S, Shimizu K, Hamamoto H, Honma A, Irino M, Toda K, Matsumura A, et al., Mech Ageing Dev. 1984 Jul;26(1):91-102)に基づいて評価を行い、観察期間中の生存期間を計測した。 The grading score system: a method for evaluation of the degree of senescence in senescence accelerated mouse (the grading score system proposed by Toshio Takeda; SAM) '' authored by Hosokawa M, Kasai R, Higuchi K, Takeshita S, Shimizu K, Hamamoto H, Honma A, Irino M, Toda K, Matsumura A, et al., Mech Ageing Dev. 1984 Jul; 26 (1) : 91-102) and the survival time during the observation period was measured.
100%の湿度の環境下では、上記マウスは、肺胞を介して1呼吸サイクル毎に約2%の水分を吸収する。上記のiPS細胞培養上清を含むミストの吸収により、1呼吸サイクル毎に約0.01%の培養上清が吸収されることになる。マウスの呼吸数は1分間に約50回であるため、1分間に約0.5%のiPS細胞培養上清が血中に吸収されたと考えられる。 In a 100% humidity environment, the mouse absorbs approximately 2% of water every breathing cycle through the alveoli. By absorption of the mist containing the above iPS cell culture supernatant, about 0.01% of the culture supernatant is absorbed for each respiratory cycle. Since the respiratory rate of the mouse is about 50 times per minute, it is considered that about 0.5% of the iPS cell culture supernatant was absorbed into the blood per minute.
図1には、6週齢(実験開始時)における肺の状態、および30週齢における実験群(iPS−CM)と対照群それぞれの肺の状態を示す。図1から分かるように、対照群においては肺線維症が進行しているのに対して、実験群では肺線維症の進行は顕著に抑えられている。このことから、本開示に係る医薬組成物には、肺線維症を始めとする肺疾患や肺の組織異常の発症を抑制する効果があることが分かる。 FIG. 1 shows the state of the lungs at 6 weeks of age (at the start of the experiment) and the state of the lungs of the experimental group (iPS-CM) and the control group at 30 weeks of age. As can be seen from FIG. 1, pulmonary fibrosis is progressing in the control group, whereas the progression of pulmonary fibrosis is significantly suppressed in the experimental group. From this, it can be seen that the pharmaceutical composition according to the present disclosure has an effect of suppressing the onset of pulmonary diseases such as pulmonary fibrosis and abnormal tissue of the lung.
図2には、6週齢(実験開始時)における膵臓ランゲルハンス島の状態、および30週齢における実験群(iPS−CM)と対照群それぞれの膵臓ランゲルハンス島の状態を示す。図2から分かるように、対照群においてはランゲルハンス島の萎縮が進行しているのに対して、実験群ではランゲルハンス島萎縮の進行は顕著に抑えられている。このことから、本開示に係る医薬組成物には、(I型II型糖尿病に見られるような)ランゲルハンス島萎縮を始めとする膵臓疾患や膵臓の組織異常の発症を抑制する効果があることが分かる。 FIG. 2 shows the state of pancreatic islets at 6 weeks of age (at the start of the experiment), and the state of pancreatic islets in the experimental group (iPS-CM) and control group at 30 weeks of age. As can be seen from FIG. 2, the Langerhans island atrophy is progressing in the control group, while the Langerhans island atrophy is remarkably suppressed in the experimental group. Therefore, the pharmaceutical composition according to the present disclosure has an effect of suppressing the onset of pancreatic diseases and pancreatic tissue abnormalities such as Langerhans island atrophy (as seen in type I and type II diabetes). I understand.
図3には、6週齢(実験開始時)における肝臓の状態、および30週齢における実験群(iPS−CM)と対照群それぞれの肝臓の状態を示す。図3から分かるように、対照群においては肝線維症が進行しているのに対して、実験群では肝線維症の進行は顕著に抑えられている。このことから、本開示に係る医薬組成物には、肝線維症を始めとする肝臓疾患や肝臓の組織異常の発症を抑制する効果があることが分かる。 FIG. 3 shows the state of the liver at 6 weeks of age (at the start of the experiment), and the state of the livers of the experimental group (iPS-CM) and the control group at the age of 30 weeks. As can be seen from FIG. 3, liver fibrosis is progressing in the control group, whereas the progression of liver fibrosis is significantly suppressed in the experimental group. From this, it can be seen that the pharmaceutical composition according to the present disclosure has an effect of suppressing the onset of liver diseases such as liver fibrosis and liver tissue abnormalities.
図4には、7週齢、30週齢および70週齢におけるラット真皮内におけるコラーゲン蓄積の状態を示す。図4から分かるように、対照群においてはすでに7週齢でコラーゲン含有量の減少が起こっているのに対し、実験群ではコラーゲン含有量の減少は40週齢や70週齢でも顕著に抑えられている。このことから、本開示に係る医薬組成物には、コラーゲン含有量の減少を始めとする皮膚疾患や皮膚の組織異常の発症を抑制する効果があることが分かる。 FIG. 4 shows the state of collagen accumulation in the rat dermis at 7 weeks, 30 weeks and 70 weeks of age. As can be seen from FIG. 4, in the control group, the decrease in collagen content has already occurred at 7 weeks of age, whereas in the experimental group, the decrease in collagen content is significantly suppressed even at 40 and 70 weeks of age. ing. From this, it can be seen that the pharmaceutical composition according to the present disclosure has an effect of suppressing the onset of skin diseases and skin tissue abnormalities including a decrease in collagen content.
図5には、32週齢において、実験群および対照群のそれぞれに対してモリスの水迷路学習課題(Morris, R (May 1984). "Developments of a water-maze procedure for studying spatial learning in the rat", Journal of neuroscience methods 11 (1): 47−60)を実施した結果を示す。具体的には、直径150cmの円筒状容器内に30cmの深さまで水を張り、その中に水面より1cm〜2cmまで立ち上がっている視認できないプラットフォームを設けた。このプラットフォームを利用して水迷路を脱出するまでの時間を測定したところ、対照群と比較して、実験群においては実験回数の増加と共に所要時間が顕著に減少しており、高い学習記憶能力が維持されていることが分かる。
この結果から、本開示に係る医薬組成物には、学習記憶能力低下を始めとする脳機能低下や脳疾患の発症を抑制する効果があることが分かる。
Fig. 5 shows Morris, R (May 1984). "Developments of a water-maze procedure for studying spatial learning in the rat."", Journal of neuroscience methods 11 (1): 47-60). Specifically, water was stretched to a depth of 30 cm in a cylindrical container having a diameter of 150 cm, and an invisible platform rising from 1 cm to 2 cm from the water surface was provided therein. Using this platform, we measured the time to escape from the water maze. Compared to the control group, the experimental group showed a significant decrease in the required time with an increase in the number of experiments, and high learning memory ability. It can be seen that it is maintained.
From this result, it can be seen that the pharmaceutical composition according to the present disclosure has an effect of suppressing the onset of brain function deterioration and brain diseases including learning memory ability decline.
図6には、30週齢における脳内の状態を示す。図6から分かるように、対照群においては神経の変性、神経細胞の減少が起こっているのに対し、実験群では神経組織は正常に保たれ、神経細胞の減少も顕著に抑えられている。このことから、本開示に係る医薬組成物には、神経変性の防止、神経細胞減少の防止を始めとする脳疾患や脳の組織異常の発症を抑制する効果があることが分かる。 FIG. 6 shows the state in the brain at 30 weeks of age. As can be seen from FIG. 6, in the control group, nerve degeneration and nerve cell decrease occurred, whereas in the experimental group, nerve tissue was kept normal and the decrease in nerve cells was remarkably suppressed. From this, it can be seen that the pharmaceutical composition according to the present disclosure has an effect of suppressing the onset of brain diseases and brain tissue abnormalities including prevention of neurodegeneration and prevention of neuronal cell loss.
図7には、30週齢における脳内の神経細胞数測定の対象部位を示す。神経細胞数測定の結果を図8に示す。図8中で「Pre」は対照群、「Post」は実験群を表し、縦軸は細胞数の減少を表している。図8から分かるように、対照群においては神経細胞の減少が起こっているのに対し、実験群では神経細胞数の減少は顕著に抑えられている。このことから、本開示に係る医薬組成物には、神経細胞減少の防止を始めとする脳疾患や脳の組織異常の発症を抑制する効果があることが分かる。 FIG. 7 shows a target site for measuring the number of nerve cells in the brain at 30 weeks of age. The results of measuring the number of nerve cells are shown in FIG. In FIG. 8, “Pre” represents the control group, “Post” represents the experimental group, and the vertical axis represents the decrease in the number of cells. As can be seen from FIG. 8, in the control group, a decrease in the number of neurons occurs, whereas in the experimental group, the decrease in the number of neurons is remarkably suppressed. From this, it can be seen that the pharmaceutical composition according to the present disclosure has the effect of suppressing the onset of brain diseases such as prevention of neuronal cell loss and brain tissue abnormalities.
図9には、実験群(iPS−CM群)および対照群それぞれについての、生存曲線を示した。図9から分かるように、対照群においては、雄の場合も雌の場合も、実験開始後早い時期から生存率が低下し始め、63週の平均寿命であった。一方、実験群においては雄の場合も雌の場合も大幅な寿命の延長が見られ、120週の平均寿命であった。このことから、本開示に係る医薬組成物には、老化の進行を抑制する効果があることが分かる。 FIG. 9 shows survival curves for the experimental group (iPS-CM group) and the control group. As can be seen from FIG. 9, in the control group, the survival rate began to decrease at an early stage after the start of the experiment in both the male and female cases, and the life expectancy was 63 weeks. On the other hand, in the experimental group, the lifespan was significantly extended in both the male and female cases, with an average life of 120 weeks. From this, it can be seen that the pharmaceutical composition according to the present disclosure has an effect of suppressing the progress of aging.
(プラスミドを用いて作製したヒトiPS細胞)
また、iPS細胞の作製元となる細胞としてヒト骨髄由来間葉系細胞を用いた以外は、Okita K. et al.: Nat Methods 8, 409-412に記載された方法に従って作製されたヒトiPS細胞でも実験を行った。具体的には、プラスミドpCXLE-hOCT3/4-shp53(OCT3/4を有するpCXLE-hOCT3/4 (Addgene accession code 27076)のBamHIサイトにマウスU6プロモータに駆動された、p53用shRNA発現カセットを挿入したもの)、pCXLE-hSK(Addgene accession code 27078、SOX2およびKLF4を有する)およびpCXLE-hUL(Addgene accession code 27080、L-MYCおよびLIN28を有する)を用意し、100μlキットと共にマイクロポレーター(インビトロゲン社)を用いて6×105個のヒト骨髄由来間葉系細胞にエレクトロポレーションで導入した。導入条件は、1800V、20ms、一回のパルスであった。導入の7日後に細胞をトリプシン処理し、1×105細胞をSNLまたはMEFフィーダー層で覆われた100mmディッシュに再プレーティングした。次の日に培地を上記iPS細胞用培地に交換した。プレーティングの26〜32日後にコロニーをカウントした。ヒトES細胞に類似したコロニーを選抜した。こうして得られたヒトiPS細胞を用いて、培養上清の作製を、上記と同様にして行った。そして、この培養上清を用いて上記と同様にSAMP8マウスに吸入させた場合の効果を調べた。
その結果、レトロウィルスを用いて作製されたヒトiPS細胞の場合と同様の結果が得られた。このように、iPS細胞の作製方法によらず、iPS細胞培養上清は先制医療や抗加齢などの点において優れた性質を有していた。
(Human iPS cells prepared using plasmid)
In addition, human iPS cells prepared according to the method described in Okita K. et al .: Nat Methods 8, 409-412, except that human bone marrow-derived mesenchymal cells were used as the source cells of iPS cells. But I did an experiment. Specifically, the p53 shRNA expression cassette driven by the mouse U6 promoter was inserted into the BamHI site of plasmid pCXLE-hOCT3 / 4-shp53 (pCXLE-hOCT3 / 4 with OCT3 / 4 (Addgene accession code 27076)) ), PCXLE-hSK (with Addgene accession code 27078, SOX2 and KLF4) and pCXLE-hUL (with Addgene accession code 27080, L-MYC and LIN28) and a microporator (Invitrogen) ) Was introduced into 6 × 10 5 human bone marrow-derived mesenchymal cells by electroporation. The introduction conditions were 1800 V, 20 ms, and one pulse. Cells were trypsinized 7 days after transfection and 1 × 10 5 cells were re-plated into 100 mm dishes covered with SNL or MEF feeder layers. On the next day, the medium was replaced with the medium for iPS cells. Colonies were counted 26-32 days after plating. Colonies similar to human ES cells were selected. Using the human iPS cells thus obtained, the culture supernatant was prepared in the same manner as described above. And the effect at the time of making it inhale to a SAMP8 mouse | mouth similarly to the above using this culture supernatant was investigated.
As a result, the same results as in the case of human iPS cells prepared using a retrovirus were obtained. Thus, regardless of the iPS cell production method, the iPS cell culture supernatant had excellent properties in terms of preemptive medicine and anti-aging.
以上説明したとおり、本開示に係るiPS細胞培養上清が有するユニバーサルかつ強力な作用により、本開示に係る医薬組成物や化粧品を用いれば、疾患や組織異常の先制医療や治療(進行抑制も含む)あるいは美容的外観の維持を達成することが可能となる。 As described above, due to the universal and powerful action of the iPS cell culture supernatant according to the present disclosure, if the pharmaceutical composition or cosmetic according to the present disclosure is used, preemptive medical treatment or treatment of disease or tissue abnormality (including progression inhibition) ) Or maintaining a cosmetic appearance.
この発明は、上記発明の実施の形態及び実施例の説明に何ら限定されるものではない。特許請求の範囲の記載を逸脱せず、当業者が容易に想到できる範囲で種々の変形態様もこの発明に含まれる。
本明細書に記載された全ての文献、特許出願、および技術規格は、個々の文献、特許出願、および技術規格が参照により取り込まれることが具体的かつ個々に記された場合と同程度に、本明細書中に援用されて取り込まれる。
The present invention is not limited to the description of the embodiments and examples of the invention described above. Various modifications may be included in the present invention as long as those skilled in the art can easily conceive without departing from the description of the scope of claims.
All documents, patent applications, and technical standards mentioned in this specification are to the same extent as if each individual document, patent application, and technical standard were specifically and individually described to be incorporated by reference, Incorporated herein by reference.
Claims (34)
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、医薬組成物の製造方法。 (1) culturing iPS cells; and (2) collecting the culture supernatant obtained by culturing the iPS cells;
A method for producing a pharmaceutical composition, comprising:
(2)前記iPS細胞の培養により得られた培養上清を回収するステップ、
を含む、化粧品の製造方法。 (1) culturing iPS cells; and (2) collecting the culture supernatant obtained by culturing the iPS cells;
A method for producing cosmetics, comprising:
Priority Applications (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP2015003493A JP2016128396A (en) | 2015-01-09 | 2015-01-09 | Pharmaceutical compositions containing ips cell culture supernatant and production methods thereof, cosmetics and production methods thereof, antiaging compositions, methods for inhibiting disease onset, methods for treating diseases, methods for treating tissue disorders, and cosmetic methods |
PCT/JP2016/050728 WO2016111377A1 (en) | 2015-01-09 | 2016-01-12 | MEDICINAL COMPOSITION COMPRISING iPS CELL CULTURE SUPERNATANT AND METHOD FOR PRODUCING SAME, COSMETIC AND METHOD FOR PRODUCING SAME, ANTIAGING COMPOSITION, METHOD FOR INHIBITING DISEASE ONSET, METHOD FOR TREATING DISEASE, METHOD FOR TREATING TISSUE ABNORMALITY AND BEAUTIFICATION METHOD |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP2015003493A JP2016128396A (en) | 2015-01-09 | 2015-01-09 | Pharmaceutical compositions containing ips cell culture supernatant and production methods thereof, cosmetics and production methods thereof, antiaging compositions, methods for inhibiting disease onset, methods for treating diseases, methods for treating tissue disorders, and cosmetic methods |
Publications (1)
Publication Number | Publication Date |
---|---|
JP2016128396A true JP2016128396A (en) | 2016-07-14 |
Family
ID=56356060
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP2015003493A Pending JP2016128396A (en) | 2015-01-09 | 2015-01-09 | Pharmaceutical compositions containing ips cell culture supernatant and production methods thereof, cosmetics and production methods thereof, antiaging compositions, methods for inhibiting disease onset, methods for treating diseases, methods for treating tissue disorders, and cosmetic methods |
Country Status (2)
Country | Link |
---|---|
JP (1) | JP2016128396A (en) |
WO (1) | WO2016111377A1 (en) |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN107158165A (en) * | 2017-05-23 | 2017-09-15 | 广州润虹医药科技有限公司 | Include the anti-apolexis composition of IPS cell extracts |
WO2018203499A1 (en) | 2017-05-02 | 2018-11-08 | 剛士 田邊 | Pharmaceutical composition and cosmetic composition |
US12036245B2 (en) | 2018-11-07 | 2024-07-16 | I Peace, Inc. | Pharmaceutical composition and cosmetic composition |
Families Citing this family (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN110755361A (en) * | 2019-09-26 | 2020-02-07 | 盛世荣恩生物科技有限公司 | Skin care product with iPS cell culture solution as main component and preparation method thereof |
JP7007770B1 (en) * | 2021-06-21 | 2022-02-10 | パナジー株式会社 | Eye symptom improver and eye symptom improvement method |
WO2023176187A1 (en) * | 2022-03-15 | 2023-09-21 | アイ ピース, インコーポレイテッド | Drug, cosmetic, and method for providing same |
Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2010093848A1 (en) * | 2009-02-13 | 2010-08-19 | Invitrx, Inc. | Skin cream |
WO2011118795A1 (en) * | 2010-03-26 | 2011-09-29 | 国立大学法人名古屋大学 | Composition for treatment of damaged part |
WO2013118877A1 (en) * | 2012-02-10 | 2013-08-15 | 株式会社ジャパニック | Cosmetic product or skin regeneration promoter comprising nonhuman stem cell culture supernatant as starting material, and method for ion introduction for protein |
US20140093486A1 (en) * | 2012-10-01 | 2014-04-03 | Taipei Veterans General Hospital | Method for preparing induced pluripotent stem cells and its applications |
-
2015
- 2015-01-09 JP JP2015003493A patent/JP2016128396A/en active Pending
-
2016
- 2016-01-12 WO PCT/JP2016/050728 patent/WO2016111377A1/en active Application Filing
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2010093848A1 (en) * | 2009-02-13 | 2010-08-19 | Invitrx, Inc. | Skin cream |
WO2011118795A1 (en) * | 2010-03-26 | 2011-09-29 | 国立大学法人名古屋大学 | Composition for treatment of damaged part |
WO2013118877A1 (en) * | 2012-02-10 | 2013-08-15 | 株式会社ジャパニック | Cosmetic product or skin regeneration promoter comprising nonhuman stem cell culture supernatant as starting material, and method for ion introduction for protein |
US20140093486A1 (en) * | 2012-10-01 | 2014-04-03 | Taipei Veterans General Hospital | Method for preparing induced pluripotent stem cells and its applications |
Non-Patent Citations (6)
Title |
---|
BUCCINI, S. ET AL.: "Cardiac progenitors derived from reprogrammed mesenchymal stem cells contribute to angiomyogenic rep", BASIC RES. CARDIOL., vol. 107, no. 6, JPN6016011122, 2012, pages 301, ISSN: 0003893079 * |
DENG, C.: "INDUCED PLURIPOTENT STEM CELL-DERIVED CONDITIONED MEDIUM ALLEVIATE HYPERTROPHIC SCARS VIA INHIBITING", WOUND REPAIR AND REGENERATION, vol. 22, no. 2, JPN6016011112, 2014, pages 37, ISSN: 0003893075 * |
GAZDHAR, A. ET AL.: "The secretome of induced pluripotent stem cells reduces lung fibrosis in part by hepatocyte growth f", STEM CELL RESEARCH & THERAPY, vol. 5, no. 6, JPN6016011111, 2014, pages 123, ISSN: 0003893074 * |
MORA, A. L. ET AL.: "Aging and lung injury repair: a role for bone marrow derived mesenchymal stem cells", J. CELL BIOCHEM., vol. 105, no. 3, JPN6016011115, 2008, pages 641 - 647, ISSN: 0003893077 * |
NEEL, S. ET AL.: "Induced pluripotent stem (iPS) cells inhibit apoptosis and fibrosis in streptozotocin-induced diabet", MOL. PHARM., vol. 8, no. 5, JPN6016011114, 2011, pages 2350 - 2357, ISSN: 0003893076 * |
ZHANG, Y. ET AL.: "Rat induced pluripotent stem cells protect H9C2 cells from cellular senescence via a paracrine mecha", CARDIOLOGY, vol. 128, no. 1, JPN6016011119, 2014, pages 43 - 50, XP093046844, ISSN: 0003893078, DOI: 10.1159/000357423 * |
Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2018203499A1 (en) | 2017-05-02 | 2018-11-08 | 剛士 田邊 | Pharmaceutical composition and cosmetic composition |
JPWO2018203499A1 (en) * | 2017-05-02 | 2020-01-23 | 剛士 田邊 | Pharmaceutical and cosmetic compositions |
CN107158165A (en) * | 2017-05-23 | 2017-09-15 | 广州润虹医药科技有限公司 | Include the anti-apolexis composition of IPS cell extracts |
US12036245B2 (en) | 2018-11-07 | 2024-07-16 | I Peace, Inc. | Pharmaceutical composition and cosmetic composition |
Also Published As
Publication number | Publication date |
---|---|
WO2016111377A1 (en) | 2016-07-14 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
WO2016111377A1 (en) | MEDICINAL COMPOSITION COMPRISING iPS CELL CULTURE SUPERNATANT AND METHOD FOR PRODUCING SAME, COSMETIC AND METHOD FOR PRODUCING SAME, ANTIAGING COMPOSITION, METHOD FOR INHIBITING DISEASE ONSET, METHOD FOR TREATING DISEASE, METHOD FOR TREATING TISSUE ABNORMALITY AND BEAUTIFICATION METHOD | |
JP7551975B2 (en) | Neuronal extracellular vesicles | |
JP6296622B2 (en) | Method for producing composition for treatment of damaged area | |
AU2014250190B2 (en) | Method for producing reprogrammed derivative neuronal stem cell from non-neuronal cell by using HMGA2 | |
CA3113484A1 (en) | Methods and compositions for the clinical derivation of an allogenic cell and therapeutic uses | |
Kim et al. | Extensive characterization of feline intra-abdominal adipose-derived mesenchymal stem cells | |
JP7094567B2 (en) | Method for producing neural crest cells and sympathetic nerve cells | |
WO2014057997A1 (en) | Reprogramming peptide and use thereof | |
US20210230551A1 (en) | Enhancement of fibroblast plasticity for treatment of disc degeneration | |
JP6856240B2 (en) | Method for preparing vascular endothelial progenitor cells | |
Murphy et al. | Cell therapy for cystic fibrosis | |
Panda et al. | Non-viral reprogramming and induced pluripotent stem cells for cardiovascular therapy | |
JP7058431B2 (en) | Freeze-dried preparation containing megakaryocytes and platelets | |
WO2016111378A1 (en) | COMPOSITION FOR TREATING NEURODEGENERATIVE DISEASE COMPRISING iPS CELL CULTURE SUPERNATANT AND METHOD FOR PRODUCING SAME, METHOD FOR INHIBITING NEURODEGENERATIVE DISEASE ONSET AND METHOD FOR TREATING NEURODEGENERATIVE DISEASE | |
Gardlik | Inducing pluripotency using in vivo gene therapy | |
TWI852685B (en) | Human induced pluripotent stem cells-differentiated cardiomyocytes, methods of producing the same, and uses thereof in treating cardiac diseases | |
WO2021045238A1 (en) | Method for preparing skin-derived skeletal muscle cells | |
Xiao-cheng et al. | Prospective use of skin-derived precursors in neural regeneration | |
TWI769410B (en) | New induced pluripotent stem cells (ipscs) and applications thereof | |
EP4310176A1 (en) | Pericyte having basic fibroblast growth factor (bfgf) gene introduced therein | |
JP6654323B2 (en) | Cells capable of forming stratified epithelial tissue and method for producing the same | |
KR20200047096A (en) | Composition for inducing dedifferentiation for induced pluripotent stem cells | |
Mohanty et al. | Biological Basis and Molecular Mechanism of Regeneration | |
Nematollahi-Mahani et al. | Biological and biochemical characteristics of human umbilical cord mesenchymal cells | |
Gaskell et al. | Development of robust, scalable and synthetic systems for the maintenance of pluripotency and subsequent differentiation |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
A621 | Written request for application examination |
Free format text: JAPANESE INTERMEDIATE CODE: A621 Effective date: 20170721 |
|
A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20180403 |
|
A02 | Decision of refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A02 Effective date: 20181009 |