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（平成24年度バイオ・ライフサイエンス委員会（第3部会））

Contents

1.
2. 5 i
3. 1 Robert P Lanza 2 2002 p 677 678 1 Saito 3 The Journal of Thoracic and Cardiovascula
4. 2009 1 5 1 29 1 13 3 04 15 2 32 11 3
5. 4 6 ACT 7736896 Methods for producing enriched population of human retinal pigment epithelium cells ACT 11 186720 20 0 5 7 2 0 2004 1 2 3 PCT US2005 00227 3 WO2005 0700 1 1 2006 551392 4 6 1 161 20054077208 HB 2005
6. 1 1 5
7. ft 6 F tt R PE
8. 17 2 4 17 2 4 159 1 53 1 21 9 28 1 0129
9. 1 2 19 4 36 6 2 1 1 1 2 15 16 19 2011 9 30
10. 5 2011 8 1 1 Keller 4 Circulation 2001 Vol 104 pp 2063 2068 1
11. XB 8 10 C 1999 vol 52 No 12 pp 1251 1255 2000 vol 45 No 10 pp 1766 1794 Hirose Biomacromolecules 2000 vol 1 pp 377 381
12. 2 1 2 0 80 1 2
13. 2 9 2 A 1 1 7 R PE 8 1 3 hES RPE 1 4 1 7 RPE R PE 1 8 RPE
14. 1 180
15. close close 3 close 40 Surface Cotact 2
16. 3 3 2012 8 8 3 2 5 01945762 1 1 2003 1 21 13 otz POT 13 1 A myocardium like cultured cell sheet made of myocardial tissue cells that retain contracting and relaxing functions intercellular electrical coupling and orientation 2 A three dimensional structure of myocardium like cultured cells that retain contracting and relaxing functions as well as 34 three dimensional intercellular electrical coupling and orientation said structure forming tu
17. 0010 5 5 3 29 2 1
18. 102 AC T R et inal Pigment E pithel ium RPE A CT 4 2
19. 1 4 P A 6 PAG6 ES RPE B 2 3 RPE R PE FGF FGF RPE B 3 3
20. 3 4 4 RPE BHE 4 1 ES iPS
21. 1 Suzuki 6 2 3 8 8 10 4
22. 5 8 1 24
23. ES RPE RPE 1 2 E DMEM 0070 DMEM
24. 2 4 29 1 1 29 2 44 4 2006 519056 67 15 3 28 79 0114
25. 3 3 2 3 3 1 2003 8 1 2003 285476 2004 2 2 5 1 21 22 23 24 26
26. 4 iPS 8 5 10 1 2 2 1 2 12
27. 5 2 2 3 36 4 1 6 1 1 69 0022
28. 0010 4 4 Suzuki 5 5
29. 0141 1
30. 1 2 3 ii 1 3 1 13 9
31. 1993 144 4 495 500 1998 19 173 1748
32. 1 3 3 3 ES PA6 P A 6 1580 SD I A ES P A 6
33. 25 1 2 1
34. GRUB DUEGTUCU S 2 2006 2 28 2007 9 10 EPC 54 2 56 84 2008 4 6 26 2010 2 25 201049 H7 H 2011 6 28 EPC 54 2 56 83 84 2012 1 9 2013 1 24 EPC 54 56 2010 9 30 2 3 2006 1 31 56 2009 3 30 2009 6 30 2009 7 14 non final OA 35 USC 102 b 2010 1 14 2010 4 15 final OA 35 USC 112 1st 102 b 103 2010 10 13 Notice of Appeal 2010 12 13 RCE 2011 6 16 non final OA 35 USC 112 1st 2nd 102 b 103 201
35. 0022 0026 2 2 1 Nat Med 2006 Apr 12 4 459 65 1 MSC
36. 6 4 2 3 8 10 2012 8 27 1
37. RPE 158 R PE RPE 5
38. WO99 03973 MO00 32749 DD COR 3 2010 016910 2010 7 27 WL
39. IB 4 6 11 13 ii 1 13 J Biomater Sci Polymer Edn 1998 Vol 9 No 12 p 1331 1348 1 3
40. 3 4 29 1 3 29 2 1 SM sheets Ahmad 4 1 1 Suzuki 6 Cir
41. 6 5 7 MN 5 6 8 1 4 1 9 1 4 8 1
42. RPE 2 9 h ES RPE 2 RPE RPE RPE ARPE 19 RPE R PE R PE 3 1 2 4 2 1 3 2
43. 1 2 2 2
44. 1 11 15 112 2 T 2011 12 H 16 2012 7 16 RCE 1 11 15 5
45. 29 2 1 3 MN 1 2 4 1 3 1 Fo 5 0 8 0C 1 IEEE 2 3
46. x m 1 th Geriatric Medicine 2003 Vol 41 No 12 pp 1791 1795 pp 103 115 3 KAWASAKI H et al PNAS 20024 Vol 99 No 3 pp 1580 1585 4 THOMSON J A et al Biology of Reproduction 1996 Vol 55 pp 254 259 5 2003 52376 6 6 KAWAMORITA M etal Human Cell 20024 Vol 15 No 3 pp 178 182 4 5 8 1 h ES RPE a h ES h ES
47. F A 1 2 12 42 1 2 A 1 1 2 RP E hES RPE A 2 R PE R PE ES RPE A 3 9 hES RPE R PE
48. 3 R PE 3 FGF 20ng mL 158 1 RPE FGF 1 Dunn et al FGF RPE 2 Jean et a
49. 0 80C 0 lt 80 0 C 80C close contact direct contact close close allowing the thus obtained myocardium like sheet to adhere to another cell sheet on a normal cell culture support or a cell culture support coated with a temperature responsive polymer and repeating this process as appropriate
50. 1 3 3 5 04707319 2 1 1 2006 2 28 24 84 252 1 6 10 11 16 17 22 23 Diacrin D1 US6432711 D2 W001 07568 D3 Circulation vol 104 no 17 pp 2063 2068 54 2 18 21 24 26 D1 Shimizu 5M DA Circulation research vol 90 No 3 pp e40 48 Okano D5 http www j circ or jp engl ish sessions reports 66th ss okano htm 56
51. 30 1 61 2 29 1 3 29 46 2 IR i 1 6 9 26 1 WO02 08387 1 ii 1 4 5 9 23 2 2003 4 vol 104 p 422
52. 12 ABCEMPRBCULEWUDERBRA DNS EE WORISAZUBBMEISIE ELI 8 iPS IPS 2 3 EP1970446 Nuclear reprogramming factor No 06834636 0 5 4 1 2 12 2 EP1970446 1 a Oct
53. 2 1 3 1 2 1 4 1 3 1 5 5 0wm 1 4 1 6 Ila llb Ild IIx CD56 MyoD Myf5 myogen in 1 1 5 1
54. 1 2 1 3 1 2 4 5 0 m 1 3 1 5 Ila IIb
55. 101 ER 1 lt 13 112 1 1 3 lk Berg American Journal of pathology 1984 vol 114 p 187 200 102 b Ek UTO 49 DY 1 10 12 13 Kikuchi Hirose 5 FEM Eppenberger Akins 103 25 1 11 Soejima Li SL HM Kikuchi Hirose Eppenberger Akins 103 2008 7 9 2008 7 12 10 12 101 126
56. 16 1 23 US 60 538 964 44 1 IB 2006 551392 4 4 3 ACT ACT US7736896 WO2005 07001 1 2006 551392 ES RPE ES RPE ES R PE 107 ACT ES RPE ES RPE
57. 0022 0160 D 82 0160
58. 1 ELE 1 63 2 29 1 3 3 29 2 1 1
59. 3 3 1 2002 12 11 2007 3 27 gt 2008 11 5 29 1 3 2 32 2009 1 5 TIER 2010 4 27 29 2 IR 19 2010 7 27 2010 9 10 2010 9H 17 2010 12 17 2010 12 21 2010 12 27 2011 1 21 2011 2 H 10
60. 6 6 2 6 4 36 6 2 1 22 24 26 1 24
61. 1 1 139 RCS 5 RCS PBS aFGF bFGF l ONL 5 8 4 intr
62. 3 RCE 1 2 54 2000 6 30 1 3
63. 12 16 13 15 3 1 4
64. 39 2 1 3 1 EPC54 2 DD 1 5 7 8 D5 3 EPC56 PIPAAm D5 3 2010 1 20
65. 1 2 3 3 2 3 1 3 78 RIA SRO RAIL BEIT OC CML SRAI CE DTRABCE TEAL
66. 16 1 23 US 60 538 964 PCT US2005 002273 N02005 07001 1 17 8 4 2012 5 8 US AU CN EP CA 2 2007 55211 2007 7 20 2008 53098 2008 8 1 4 17 1 24 US 11 041 382 PCT US2005 025860 N02006 080952 18 8 3 AU CN BRPI CA EP 106 3 4 5 2010 528899 2008 10 10 2011 500024 2011 1 6
67. 1 5 3 7 0 8 0 C 1 2 3 4 3 8 4 a 3 31 b c b
68. RPE 2 E 3 1 21 34 42 1 3 hES RPE HoT a MEF hES h ES b c R PE LIF bFGF
69. c RPE R PE 2 6 2 2 RPE hES 132 R PE a nhES b c RPE R PE 2 7 2 2 RPE hES R PE
70. 5 EPC84 D5 J Biomater Sci Polymer Edn 1998 Vol 9 No 12 p 1331 1348 PIPAAm 5 D5 EPC52 2 D5 D5 D5 PET EPC52 2 10
71. LAL 1 2 3 1 1
72. 1 2 1 2 W099 03973 MSCs i
73. 2 0 1 1 4 pp62 63 3 Adv anced cell technologies ACT ES RPE iPS ES RPE ACT a
74. 1 Lanza D OM 2I RISER MAA E BR EX ELIO PARES MAILE SHRED 6H 3 Saito 3 Suzuki 4 39D 1 4 1 4
75. 0320 1 4 1 6 pluripotency 71 2 AME Clee MAIS ER ES 0021
76. 1 9 1 9 1 9 ii 36 6 2 10 1 9 2 2 5 2012 5 11 i 1 1 2 3 4 1 0ct3 4 2 KIf 2 KIf 4 3 c Myc N Myc L Myc T 5 8 A
77. 1 0128 Lanza 2 1 70 17 2 4 1 16 1
78. 1 RCS 1 3 2 A 12 4 2 1 2 1
79. 1 2006 1 30 2006 2 20 55 2007 1 31 2008 12 22 A 29 1 2 36 6 2 2009 2 23 2009 6 26 29 2 2009 9 28 2009 11 9 2010 1 15 2011 8 1 XE 2011 9 30 2012 1 12 36 6 2 2012 1 24 2012 2 15 2012 3 9 2012 2 7
80. D5 D5 Jh dx SE RE D5 1 3 D5 4 3 2010 2 9 obtainable from the li
81. 6 myogenin MyoD Myf5 Carnac 5 98 CD56 13 12 14 15 50 100 5 US6207451 US6207451 US6207451 27 28 112 1 Diacrin W001 07568 Jin Jour Pharm and Exp Therapeutics 02 01 2003 304 654 660
82. 124 2 9 2 3 3 6 6 1 4 3 6 6 2 A 1 2 12 42 1 2 1 R PE 1793 BARS 377 RCS 2
83. 2008 6 26 2008 8 8 1 24 Diacrin D1 2 Kellar D3 Akram D6 D6 D6 D6 87 Diac
84. RPE ES R PE 1 ES RPE 1 41 34 4 2 2 2 3 3 1 21 34 42 2 2 3 3
85. 1 1 1 R PE 1 5 1 1 1 RPE 1 6 Stargard t 1 4 1 7 1 4 1 8 r d RPE 6 5 tu bby RCS Abyssinian c a t 1 c d Tp r cd e r d 1
86. 2 1 3 2001 7 2 PCT JP2001 05722 2002 513872 2010 168577 01945762 1 10181984 5 10 333473 4679795 2012 2 H 10 5086398 2012 9 14 1302535 2011 4g 1 H 19 A 2275532 2012 6 6 2000 221385 2000 7 21 2 18 HS
87. 21 2 23 4 67 CHICA L HAIL RIK 2011 9 28
88. 6 RPE R PE a c 1 3 F 4 128 1 21 34 42 1 3
89. 2011 12 27 1 3 5 2 3 1 1 28 1 2 1 2
90. 2 3 ii 5 2002 Vol 31 No 1 pp 25 26 81 8 10 5 MAKI 4
91. Ooto et al RPE 1 3 1 3 1 3 137
92. 66 5 5 5 6 6 5 6 5 6 Problems and future perspectives
93. 3 17 3 1 20 N PIPAAm 1 IPDL OR AND DAH OR AND DAR OR AND
94. 1 8 2 4 1 8 9 Hirose 10 HAEC
95. 1 3 3 6 4 RPE 3 ES PA6 R PE 4 ES PA6 6 R PE 157 D 2 C 1 3 ES PA 6 RPE 4 5 PAG6
96. 1 5 3 2002 513872 0004 0004 3T3 6 2002 513872 2010 7 27
97. 2 RPE ES R PE ES R PE R PE DNA N N A2E R PE
98. 1 3 1 4 Ila IIb IId IIx CD56 MyoD Myf5 myog enin 1 1 3 1 5 509 4 6 Ila 73 IIb IId IIx CD56 MyoD Myf5 myog en i n 1 5
99. 1 1 6 1 6 1 1 1 6 1 6 10 1 3 6 8 10 1 1
100. lt 2 E 3 OR PE 3 F 4 4 foe LTWSRBERLE 3 1 3 22 25 27 RIB 1 3 5 12 15 22 27RU29 35Q 0 RPE MMI RPE R PE RPE
101. another cell sheet a normal cell culture support Ik normal to adhere to another cell sheet on a normal cell culture support or a cell culture support coated with a temperature repponsive polymer AV on to adhere to another cell sheet which is present on a normal cell or a cell culture support or thus obtained myocaridium like sheet
102. 20 C CD56 50 CD56 CD56 CD56
103. 1 4 10 1 3 12 3 1 24 3 1 4 i 12 8 10 13 2
104. D13 CD56 2012 1 9 CD56 4 2010 9 30 2 2011 8 11 2012 3 14 04707319 2 1 A method for producing a three dimensional structure for implanting into a heart of a human to treat dilated pha
105. c R PE 2 h ES RPE a nES b c R PE 3 hES RPE a hES c R PE
106. 0010 3 64 3 Diacrin 4 1 2 3 4 5 0007
107. 1 7 2 4 1 3 c EB 1 7 2 4 4 1 7 2 4 1 3 2 4 2 2 7 1 7 2 4 5 3 1
108. c RPE 4 1 3 5 RPE P a x 6 CRALBPG PEDF RPEG65 1 4 6 cobblestone 1 5 7 b h ES FGF L IF Plasmanate 1 6 8 b
109. photobleaching Pa x 6 ik permeabilization Pa x 6 1 P a x 6
110. RR OF 16 7 8 10 Sox2 iPS Sox2 Sox2 iPS isolated 5
111. iPS EP 2 iPS 2 1 1 0 lt iPS 200 7 5 US8048999 Nuclear reprogramming factor EP1970446 Nuclear reprogramming factor 3 2 3 2
112. p 145 WO98 30679 27 29 RPE
113. 2008 2 7 2009 12 2 2011 3 30 2011 4 5 2012 3 13 2012 5 9 2012 5 11 2012 5 14 2012 6 26 2012 7 30 2012 8 28 2009 12 2 4 2 5 2 2 1 1 3 Oc t wet KI f M y c 2 3 0Ct3 4 KIf4 c Myc 1 3 Sox
114. 3 22 10043 24 1 27
115. E 3 1 21 34 42 RPE R PE 1 4 PA 6 PA6 ES RPE
116. Tzs 3 5 48 3 Soejima Li Kikuchi Hirose Eppenberger Akins 3 2006 11 15 1 10 12 13 Shimizu
117. 6 2 5 2 6 4 36 6 2 3 1 Suzuki 5 1
118. 3 d b c e 3 f Ce 3 g e h Cg 3 i g RU h Cj 3 3 1 2 5 7 1 3 9
119. 3 ES PA6 3 ES 6 8 30 6 8 9 3 1 3 gt 5 1 0 32 ES RPE 8 10 30 lt 32 5 D 1 4 2 1 6 1 3
120. 4 P A 6 ES 1 4 1 3 PA 6 PA 6 ES RPE 1 3 RPE 1 3 1 3 R PE
121. 1 7 2 4 1 3 4 d 4 4 b 6 RPE 1
122. T5 8 A c M y c T58A 36 6 2 2 2 7 c M y c T 5 8 T58A c My c Tzs 2 2 8 2012 8 28 2012 8 22 2 2 9
123. 1 3 ES R PE b 3 Fig 5D P a x 6 RP EE 3 Pax6 RPE RPE Pa x 6 R PE P a x 6 RPE P a x 6 126 C 2
124. 2005 12 13 PCT 2006 12 6 c Myc c Myc Article 123 2 D1 Cell 126 663 676 Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors 2006 8 25 2006 8 A 10 2006 12 6 1 4 9 1 0 1 4 D1 D1 2005 359537 00 14
125. 1 9 10 D1 D3 D6 EPC54 2 24 26 Shimizu D7 J Artif Organs 2002 5 pp 216 222 D7 PIPAAm PIPAAm D7 PIPAAm 24 26 D7 EPC56
126. PA 6 R PE MEF h ES EB R PE 0070 070 6 R PE R PE a c
127. RPE 1 D aw GCL 9 IPL PORT AE S20 WM HIE oos L ONL REAM HRAL Mende migm RPE l e ES RPE RPE RPE i
128. 1 11 1 2 37 C 32 37 C 32 C 1 2
129. 1 2 3 8 9 Hirose 10 ii Ahmad 4 N 4
130. 1 2 4 5 4679795 2012 1 19 12 1 0 8 0
131. 13 6 8 10 1 1 3 4 6 8 10 1 6 8 10 1 2 4 6 8 10 1 84 4 2012 8 27 13 1
132. Ald 2003 144139 2 192138 2009 2 20 1 29 1 1 3 29 2 67 15 3 28
133. 3 ES 3 PA6 1580 3 ES PA6 6 144 5 6 RPE
134. 2 RPE R PE RPE RPE tt 2 RPE 2 gt RPE ES
135. RPE 1 3 RPE
136. 0055 R PE RPE gt 1 2 MEF hES R PE MEF ME F RPE ME F MEF RPE h ES 1 3
137. 1 65 1 5 6 reference 39 42 6 6
138. gt 3 Sox family gene or gene product gene product of Sox2 5 bFGF and or SCF 6 bFGF 0Oct3 4 KIf4 Sox2 and bFGF 7 gene or gene product TERT 8 1 1 13 KIf12 Sox1 Sox3 Sox15 Sox17 L Myc or N Myc 2 1 NRF 3 2 3 1 D1 Cell 2006 15 3 8 25
139. 3 1 1 1 4 10 1 3 3 1 2 1 4 10 1 3 1 1 1 3 1 4 10 1 3 1 1
140. 4 6 1 3 4 ES ME F 4 R PE MEF PA6 2 3 4 3 4
141. ES ES 127 0004 0008 3 6 ES RPE 4 5 3 6 R PE ES
142. Oyamada SIL Etz Shimizu 103 1 1 3 2 3 2 2 3
143. 4 8 D CM 3 3 C 2 5 4 83
144. 1 19 1 4 1 29 2 1 Circulation Research 2002 vol 90 p e40 e48 68
145. 1 2 1 3 1 4 1 59 5 4 6 1 7 1 8 1 9 1 1 0
146. 2010 12 21 1 1 1 2 4 Ete BRI I 11 27
147. 3 1580 RARE 2 1584 2 3 PA6 ES Abstract 1581 5 3 E S ES PAG6 RPE B 2
148. iii 1 2 8 23 Diacrin 3 US6432711 iv 1 4 6 9 23 Diacrin 4 WO01 07568 v 1 6 9 23 Suzuki 5 Circulation 2001 Vol 104 pp 2063 2068
149. 4 1 3 5 RPE P a x 6 CRALBPG PE DF RPE65 1 4 6 cobblestone 1 5 7 b h ES FGF L IF Plasmanate 116 1 6 8 b 6 gt 1 7 9 b 6 8
150. 1584 BARS 2 R PE ES ES 3 O 3 29 2 1 6 4 ES 256 3 5 6 4 257 Fig 5 151
151. 3 3 32 3 1 4 1 4 RU 1 10 13 1 3 1 5 1 4 3 1 6 1 4 1 1 1 5 1 3 2 2012 5 24 1 6
152. 3 P a x 6 3 P a x 6 RPE 3 3 1 R PE IPE ZO 1 1 RCS
153. R PE 1580 BAHAR 3 1581 1 P A 6 3 3 h ES ES 3 1580 2
154. 1 2 A 1 M1 2 1 1793 3 7 2 Abstract 108 109 1 RPE h ES RPE 1 2 A 2 RPE
155. 6 NS I PD TE PEACOAT ER 6 2 2 5098028 7 2 2 1 7 2 2 2 2011 3 30 lk 9 2 2 3 2011 4 5 kk 10 2 2 4 10 2 2 5 2012 5 11 essnee 11 2 2 6 ki 12 2 2 7 iki 12 2 2 8 2012 8 28 kk 12 VER ut NEED NE 12 2 3 kk nnne nnne 13 2 4 iPS kk kk 17 3 22 222222 17 2 18 3 2 kk 19 3 2 1 kk 19 3 2 2 PCT JP2001 005722 21 3 2 3 2002 513872 ik 23 3 2 4 HRB 2010 168577 Lin 31 3 2 5 01945762 ee 34 3 2 6
156. D8 W099 66036 Smith D9 US5543318 37 1 2 12 EPC54 2 1 12 D5 EPC56 D5 PET D5 D5 PET
157. 1 1 6 2012 1 12 2012 1 24 7 4943844 2012 2 3 gt 19 1 2
158. Manasche D13 Lancet 2001 col 357 no 9252 pp 279 280 89 CD56 CD56 1 CD56 50 8 10 509
159. iPS iPS 6 0 0 1 1 HE BR UE oem 6 2 iPS
160. c 1 1 1 0 1 2 1 0 1 1 1 1 3 NMN 1 0 1 2 1 2 2012 2 7 1 3 29 1 1 29 2 44 4 2006 519056
161. 1 5 1 1 1 R PE 1 6 Stargard t 1 4 1 7 1 4 1 8 117 r d RPE 65 tu bby RCS Abyssinian c a t 1 c d 4 Tp r cd e r d 1 2 3 rcd1 rcd 2 r cd3 p d B riard RPE 65 1 7
162. 4 i 1 8 10 13 5 7 8 10 1 2 1998 19 25 173 174 37 C 32 37C 32C 1 2 23
163. 4 1 6 NRF 0oct3 4 KIf4 cl c Myc 0ct3 4 KIf4 Sox2 bFGF gt 1 4 2 7 8 1 0 Sox2 iPS 1 Sox2 iPS In vivo Isolated 1
164. 1 0 80 C 2 EPC84 0 80C D6 0 lt 80C
165. 2 3 6 1 005 1 0052 EB L IF FGF Plas 160 manate MEF hES 6 1 0051 nhES EB 6 8 EB 1 2 1B 0052 R 6
166. 3 29 2 i 24 26 2 5 1 2 5 62 1 ii 1 lt 6 8 26 6 Journal of Artificial Organs 2002 vol 5 pp 216 222
167. iPS iPSTrend Trend iPS 2011 2 18 4 5 ACT 2 0 0 6 551392 PCTUS2005 002273 WO200 5 07 0011 ACT ES RPE 4 5 1 1 2008 O1 30 2 2010 12 01 3 2011 06 01 4 2011 06 01 5 2011 07 23 6 2011 08 26 7 2012 02 27 8 2012 02 27 108 9 2012 03 08 10 2012 04 27 11 2012 05 29 12 2012 05 29 4 5 2
168. 2010 12 13 RCE 24 a o 27 1 2 8 17 22 4 2 2011 6 H 16 non final 27 28 1 9 16 18 21 23 112 1 98
169. 4 6 9 11 13 HAO 2008 9 18 1 36 D1 D2 D4 Cardiac tissue engineering 2000 10 21 195 35 p 203 204 D6 5 192138 2 D3 D4 12
170. ES ES 3 ES E S 3 PA6 158 ORAM 3 143 PA 6
171. 101 2 WO001 07568 US66207451
172. W000 32749 26 100 200 1
173. R PE R PE 1 1793 BARS 3 1 RCS RPE C 2 1 21 34 42 3 2 C 1 ES 3 ES 3 P A 6
174. 1 R PE 1 1 1 4 R PE R PE a 10 b PEDF 110 EPC 1 1 5 1 4 1 6 RPE HLA ES ES HL A 8 17 ES 8
175. 67 15 3 28 30 1 30 1 2 3 29 1 3 29 2 1 2003 4 vol 104 p 422
176. 3 4 4 2 4 1 4 3 MEF 1 134 4 4 1 S ARDS J FU PRES AA CHO RKA 1 4 5 9 BRS 1 2 3 8 2 3 A F 2 A 1 2 1 2 RPE R PE R PE
177. Carnac 5 Mol Biol Cell 9 1891 1902 CD56 13 12 14 15 50 1006 W001 07568 0007 97 W001 07568 US6207451 6 8
178. RPE FGF 0070 R P FGF 3 3 P ax 6 Pa x 6 RPE progenito r R PE 3 138 Fig 5D Pa x 6 P a x 6 Pa x 6 F i g 5D P a x 6
179. 2 2012 5 31 1 3 1 1 2006 519056 67 77 SM
180. 24 2 2 7 2 4 2 2 7 a 1 TMEF b c 22 4ng ml _ bFGF 1ng ml LIF 8 4 8 496PLASMANATE d 4 4 a 2 4 3 8 MEF 1 7 149 2 4 2
181. RPE R PE 19 10 12 US 60 998 668 RA F 19 10 12 US 60 998 766 20 1 2 US 61 009 911 20 1 2 US 61 009 908 PCT US2008 011669 WO02009 051671 21 4 23 US AU CA EP KR WO 2011 208002 2011 9 22 2012 0241100 2012 2 9 17 1 24 US 11 041 382 44 1 2007 552111 2 0 12 4 0 4 9 1 20 1 2 2 27 2012 148085 2012 8 9
182. ik 1 amp Oct KIf Sox 2 3 4 2 2 3 2011 4 5 2 2 4 i 36 4 1 36 6 1 2 0 509
183. 141 2 58 2E 2F 1 RPE 1 7 9 4 RPE RPE ES RPE 2 7
184. 4 9 10 14 D2 D3 ES isolated
185. 5 EPC84 D5 2 D5 3 3 D5 3 2 38 2009 9 A 10 EPC84 0 80
186. HOT 1 4 1 1 0016 5
187. 258 10 9 5G ES 0004 0008 3 6 ES RPE RPE 1 3 4 5 3 6 R PE
188. 7 1 6 1 8 7 9 7 8 1 75 1 0 a s 0 8 0C b
189. R PE 3 PA6 EE S PA6 4 6 s 1 gt 3 3 s 4 6 ES P A 6 5 E 1 2 1 34 4 2
190. 3 4 3 4 D 2 3 PA 6 3 ES PA6 D 3 6 6 R PE 2 6 D 1 D 3
191. 1 2 B Preparation and transplantation of monolayered MSCs 1 MSC 1 Abstract FAS 3 1 0101 MSC MSC 1 LVEDP dP dt RRS MSC LVEDP Fig 5a dP dt MSC Fig 5b c DFB MSC 4 DFB
192. 4 2 2 Irina Klimanskaya Robert Lanza ACT ES RPE 2 6 ACT ES R PE Cell Cure Neuroscience 103 4 3 10 RPE
193. N 5 8 1 3 1 0 5 9 1 3 1 1 5 1 0 1 3 1 2 3 1 4 1 1 1 3 1 3 1 4 1 1 1 3 3 3
194. 0061 7 b h ES FGF L IF P 1 a s m anate 7 1 6 FGF L IF Plasman ate 1 FGF FGF RPE 159
195. 1 9 2 1 R PE RPE RPE 1 2 BB 114 RP E 1 7 19 21 3 7 1 2 4 B 2 3 1
196. 136 2 4 RPE 29 h ES 4 3 4 4 4 3 4 4 gt 1 ME F 4 AD 1 1 3 1
197. 5 6 3 3 3 3 2 2010 4 27 1 p
198. 6 b c RPE 2 h ES RPE a nES b 129 c RPE 3 h ES RPE a hES b
199. 76 1 9 1 1 1 3 29 2 i
200. 2 1 2 4 2 2 h ES RPE R PE E S RPE RPE RPE 2 2 1 3 RPE 142 1 3 R PE
201. 3 8 r d RPE 65 tu bby RCS Abyssinian c at c d Tp rc d er d 1 2 3 rcd1 rcd 2 r c d3 p d B riard RPE 6 5 3 7 3 9 1 3 8 120 4 0 3 4 4 1 3 4 4 2
202. R PE bFGF TGF IBMX bmp 2 bmp 4 EB ES R PE ECM I 1 V Matrigel ES RP E Pax6 Pax2 mitf RPE CRALBP PEDF REPG65 RT PCR R PE RPE
203. 1 4 1 1 1 1 1 1 3 1 1
204. 2 9 2 2 1 4 2 3 6 6 1 2 iii RPE 23 6 1 hE 123 S R PE iv h ES RPE h ES R PE
205. 4 3 4183742 4411362 4411363 25 5 4 iPS 5098028 2 2 5098028 5098028 PCT JP2006 324881 2007 550210 4
206. this process 2 4 2 3 3 this process another cell sheet normal on 2 EPC54 2 EPC56 1
207. 1 27 28 23 17 22 3 2010 4 15 Diacrin W001 07568 L 2 18 24 8 10 9 myogenin Myo Myf5
208. 5 2004 131 42 2008 4 5 10 3 ES R PE Irin a KIimanskaya 9 Irina Klimanskaya et al Nature Reviews Drug Discovery 7 131 42 2008 3 E S PA amp RPE 1 3 9 3 i 1 Zhao S et al Brain RIs 1995A 148 pr 24 677 2 300 810 11 Zhao S et al Int Rev Cytol 1 997 171 225 266 1 Haruta M et al Invest Ophthal mol Vis Sci 2004 Mar 45 3 1020 1025
209. 6 2 1 5 7 16 1 W02002 008387 1 lt 5 7 16 3 i 1 5 0 8 0 33 1 5
210. b 85 c d 9 8 1 0 8 9 1 1 N SF 8 1 0 1 1 2 8 1 1 1 1 3 8 1 2 1
211. T homson 1998 ES RPE E 5 6 1 Non Final Rejection Kawasaki ES RPE Thomson Kawasaki E S ES 10 3 177 FER Kawasaki 2 0 0 3 2 9 1 3
212. 3 ES PA 6 RPE 156 ES ES PA6 C 2 D 2 1 42 2 6 D 1 4 R PE MEF PA6 2
213. EE 8 10 1 lt 2 EB h ES Costar EB 4 5 5 1 8 13 2 ES ES RPE
214. RT P CR RPE RPE 0069 hEDC EB 6 8 EB 1 29 EB 121 3 E B EB 0072 nh ES EB hES hES MEF
215. 1 0 1 1 13 1 2 1 4 1 13 1 1 5 1 4 1 6 a 0 8 0
216. 102 a 2007 7 15 Shimizu new issue 2007 4E 10 3 2007 10 15 2007 7 15 RCE 4 2 2008 1 H9 lt 11
217. R PE 147 hES 0 0 4 8 7 5 1276 2 Wichterle et al Cell 2002 110 385 397 3 Zeng et al Stem Cells 2004 22 925 940 4 48 3 86 6 Lawrence et al IOVS 2000 41 518 528 0140 6736 12 60028 2 2012 1 2 4
218. a b 1 13 Berg 5 2008 10 15 1 lt 9 13 112 1 5 9 2009 4 15 16 1
219. d RPE RPE R PE 9 R PE a hES EB b c 12 PO d 3 e R PE 8 1 0 1 9 1 1 RPE RPE 8 1 2 RPE R PE
220. ES R PE ES R PE RCS 1793 1794 BAS 1 125 3 E S R PE Abstract 1583 2 1584 RAS 2 1 3 ES RPE RPE 2 23 6 1
221. 6 5 6 3 6
222. 2 2 2010 2 25 2003 3 28 67 88 EPO EPO 2010 9 7 1 21 22 23 24 1 24
223. 3 YES 3 RPE R PE ES 3 ES 10 50 3 ES ES P A 6
224. EPC53 c 4 1 2 10 11 4 EPC84 EPC 29 1 10 EPC 84 1 2 11 d
225. ES gt 1795 1 pp 103 115 3 KAWASAKI H et al PNAS 20024 Vol 99 No 3 pp 1580 1585 4 THOMSON J A et al Biology of Reproduction 1996 Vol 55 pp 254 259 5 2003 523766 82 4 5 12 23 8 2 3 1 4 24 2 2 7 152 2 4 2 2 7
226. RPE Pax 6 ES Pax R PE Kawasaki Thomson Final Rejection 103 1 1 2 ii ATC i Kawasaki RPE Pax RP E Pax R PE R PE
227. 1 1 9 1 1 1 1 1 0 1 1 2 1 1 1 3 1 1 1 2 1 4 1 3 1 5 1 1 0 1 1 6 1 5 1 7 1 5 1 6 1 8 1 7 2 2 2
228. 2 Jean et al RPE F 4 1 21 34 42 RPE RPE
229. 1 3 1 3 J a 3 MER PE RPE b P a x 6 R PE R PE a 2 2 FGF RPE 1 1793 1794 RAS 1 3 R PE 1 4 3G Fig 5 6 1 1794 BA 2 RPE
230. R PE ES ES RPE XE 2 1 21 34 4 2 D 2 1 4 2 2 6 4 ES 256 3 5 6 4 257 Fig 5 258 10 9
231. b c 74 1 7 1 6 1 8 1 6 1 7 1 9 N 1 6 1 7 3 3 4 2009 223536 1 2011 10 28 13 1
232. 1 8 RPE 4 5 3 1 BELEE RRAPI ESBgU C oc NS Si OR TESP BRSIT Hd 5RPE HRE RP ERME RPEXIXRP ERMIR 2 5 7 lt 1 2 1 1 3 1 4 3 5 2 6 r d
233. RPE R PE 1 2 RPE 153 RPE RPE RPE RPE 1 2 RP E B 2 12 42 1 3 2 B 1
234. 3 P A 6 C 2 3 ES PA6 3 3 2 ES ES PAG ES 10 50 ES ES ES
235. Oct3 4 KIf 4 Sox 2 o 3 1 2 1 1 2 1 4 11 1 2 1 2 2 1 5 1 4 1 iPS 2 2 6 1 3 T58A
236. 1 2mM GlutaM AX I 0 1mM 4ng ml bFGF 1ng ml LIF 8 4 8 496PL ASMANATE R PE 2 3 RPE P ax 6 CRALBPG PE DF RPE65 2 2 R PE 2 4 cobblestone 118 2 2 RPE 2 5 2 2 RPE hES RP E a MEF hES h ES b
237. 2 1 5 Oct Oct 3 4 Oct 1 A Oct6 1 4 1 6 KIf KIf 1 KIf 2 KIf 4 KIf5 1 4 1 7 Myc c Myc N Myc Myc 1 4 1 8 Sox Sox 1 Sox 2 Sox 3 Sox 7 Sox1 5 Sox17 83 Sox 1 8 1 4 1 9 1 8 1 0ct KIf M y c Sox
238. 2 4 ee 2008 12 12 29 1 1 29 2 1 4 5 9 26 18 2 20 67 15 3 28 184 14 30 1
239. 4 Sox 1 Sox 2 Sox 3 Sox 1 5 Sox 1 7 0ct3 4 KIf 4 c Myc Sox 2 2 1 2 3 1 Oct3 4 2 KIf2 KIf 4 3 Sox 1 Sox 2 Sox 3 Sox 1 5 Sox 1 7
240. a hES c RPE R PE 2 8 2 5 2 7 R PE 2 9 b FGF L IF PLASMAN ATE 2 5 27 RPE 3 0 b 6 gt 25 2 7 RPE 3 1 b
241. b KIf c Myc 2 3 0ct3 4 KIf4 c Myc 1 3 Sox 1 2 4 Sox2 3 5 bFGF SCF 1 4 6 3 0ct3 4 KIf4 c Myc bFGF 1 5 7 TERT 1 6 8 SV40 Large T antigen HPV16 E6 HPV16 E7 Bmil 1 gt 1 7 9 Fbx15 Nanog
242. 1 3 12 1993 14 4 495 500 E gt 5 8 10 12 1998 19 173 174 Tissue Engineering 2001 vol 7 no 2 pp 141 51 Cardiac tissue engineering 2000 22 10 21 H 19525 35 p 203 204 3 2 3 2002 513872 1 2002 12 A 11 2000 6 30 3 2000 5 10 42
243. I PE 1 2 4 2 1 3 4 C 1 2 1 34 4 2 3 2 2 1 2 2 1 34 4 2 2 1 1 1 E S RPE 2
244. CD56 50 CD56 9 14 4 19 Lc 67 3 3 2011 6 28 DHEER CD56 2003 3 28 67 Memon N
245. G6 8 b 6 8 1 7 6 6 6
246. OE 2004 2 2 PCT JP2004 001024 2006 519056 2009 223536 2012 0087825 04707319 2 10183361 4 10183373 9 10 567728 4943844 2012 3 9 2003 285476 2003 8 1 3 2 1 3 2 1 2000 7 21 2001 7 2 ae a e 1 3 4 10 12 2 4 10 12 3 5 7 9 6 9 3 11 12 13
247. RPE 4 4 1 ACT Klimanskaya PCT US2005 002273 WO2005 070011 2005 1 2 4 US 11 041 382 E S RPE 2001 133721 200 1 4 2 7 20 0 2 325571 200211 12 ES RP E 2 1
248. 4 2 1 iPS ES RPE ES R PE 2 0 0 4 ES RPE iPS RPE 2013 iPS RPE iPS IPS
249. Abstract 108 109 1 2 2 12 15 34 1 1 1 RPE R PE R PE 1 2 2 2 1 5 2 3 1 2 42 1 2 B 2 12 42 1 3 2 1
250. 1 RPE 140 ES 1 1 2 4 2 RPE 1 2 1 2 4 2 ES RPE 2 h ES RPE R PE 2 ES R PE 2 ES RPE
251. 3 3 3 3 2 1 3 3 1 2 3 4 1 3 1 3 5 1 4 1 3 6
252. Lees 162 4 6 3 Non Final Rejection 165 4 6 4 ra NAA obses dodo yer voveo ant as 166 4 BD WIE cuida meta cum aT ncm an eT C DA CE 167 4 6 6 Final Rejection 168 dO SE tn de en f Cn cat hi 168 46 8 eis eerie wr n NIFI ER CREE 169 4 6 9 Non Final Rejection 169 4 6 10 kb 169 4 6 11 i 169 4 6 12 i 174 NG 177 ce 0 A EE E E AAE AA TET 102 1 iPS 4 iPS
253. Ild II x CD56 MyoD Myf 5 myogenin 1009 1 3 1 6 1 1 7 1 1 8 1 1 9 1 2 0 1 9 2 1 60 2 0 2 2 1 2 1 1
254. P a x 6 4 5 6 2 9 1 3 2 9 2 E S RP E Pa x 6 i 122 1 Geriatric Medicine 20084 Vol 41 No 12 pp
255. lt gt 29 2 36 6 1 2010 7 27 2 2003 1 21 2007 7 31 2008 9 18 2009 5 27 2009 9 10 2010 1 H 20H 2010 2 9 2010 7 27 2010 12 23 20114 1 H 19 EPC 53 c 3 54 2 84 EPC 54 2 56 84 2010 9 29 3 2003 1 H 21
256. IIa IIx CD56 MyoD Myf5 myogeni n 1 1 4 1 6 1 5 1 1 7 6 8 1 a 0 8 0C
257. 1 1 5 8 23 96 HI 1 2 6 10 16 26 I 2 2009 7 14 1 5 8 23 Diacrin W001 07568 US6207451 102 b 3 2010 1 14 23
258. 2 9 2 TLO Wisconsin Alumni Reseach Foundation WARF ES US7029913 ES ES 2010 4 2 8 pp151 2 0 1 1 ES 2 Ci ATC Kawasaki ES
259. RP E R PE 1 ES B 1795 2 KAWASAKI H et al PNAS 20024 Vol 99 No 3 pp 1580 1585 4 5 4 1 8 lt 13 115 1 hES RPE a MEF hES h ES b
260. 2 hd if eITAULcdsU Uhh RES AL 3 O 3 3 1 2 3 4 3 1 2 3 3 5 0 8 0 1 2 21 3 6
261. 3 14 0ct3 4 KIf4 c Myc 3 c Myc 1 Myc family c Myc 1 A nuclear reprogramming factor for a somatic cell which comprises a an Oct family gene or gene product b a KIf family gene or gene product and c a Myc family gene or gene product and or a cytokine iPS iPS
262. 1 5 3 1 2 5 2 1 0 80C
263. 1 9 ES WoT Kawasaki 1 9 Kawasaki R PE Kawasaki R PE 1 9 RPE65 P a x 166 Kawasaki ES Pax 6 T Thomson Kawasaki 1 9 RP E 1 9 1 0 3 We first discuss the differences between the claimed methods and the methods performed by Kawasaki et al Kawasaki et al teach a method of directed differentiation They co culture ES cells with PA6 cells which express
264. 7 7 84 86 8 12 15 13 15 Akram D6 US6407451 D6 1 5 11 21 D6 EPC54 2 9 10
265. P a x 6 Pa x 6 R PE RPE B 4 1 RCS 1 1 3 2 PA 6 ES RPE 154
266. 1 7 IIa IIb IId Il x CD56 MyoD Myf 5 myogenin 5 0 6 8 1Ia IIb IId Il x CD56 MyoD Myf 5 myogenin 1 0 09 6 7 9 1 8 1 1 0 1 8 1 1 1 1 0 1 2
267. 1 6 RPE HLA ES ES HL A 8 1 7 E S 8 1 8 R PE 1 9 R PE RPE RPE RP E 1 2 0 r d RPE 6 5 tubby RCS Abyssini an cat SABE cd 4 x THAA RHABE T preced 7 e rd 13 HA MARBRI1 2RU3 Tr cd1 rcd2 r cd 3 p d J Briard RPE
268. 8 h ES hES RP E ES R PE I V EB 146 h E S 8 2 3 5 1 2 MDBK GM Sigma OptiPRO SFM Gibco RPE W O 2009 05167 1 1
269. 1 a b c 23 6 1 2 4 2 2 7 1 3 2 5 2 7 1 3 2 5 2 7
270. 10 1 4 8 1 9 1 1 1 0 1 2 1 4 0 8 1 9 30 3 2 4 FRR 20107168577 1 2010 016910 2010 8 25 16 1
271. 6 1 7 9 b 6 8 1 7 1 0 b 3 5 1 7 1 1 b 130 1 1 1 0 1 2 1 1 1 R PE 1 3 1 1 1 RPE 1 4
272. 1 1 1 a Tb Td II x CD56 MyoD Myf 5 myogenin 1 1 1 2 5 09 1 1 1 3 a Tb Td I x CD56 MyoD Myf5 myogenin 1 1 4 IIa IIb IId Il x CD56 MyoD Myf 5 myogenin 5 09 1 3 1 5 Ila IIb
273. 1 21 34 42 3 2 3 ME F E S ES ES 10 50 P A 6 RPE 1580 RAS 3 1581 1 1 2 3 h ES ES
274. 2 RCE 3 14 lt 16 3 17 3 18 7 2010 12 23 Hf CA 2011 9 9 3 2012 3 8 2012 10 5 3 RCE 51 1 to 5 Canceled 6 Previously Presented A process for producing a three dimensional structure of myocardium like cells in which a myocardium ike cell sheet is produced by the steps of culturing said cells on a cell culture suppor
275. 2 RCE 1 Currently Amended A three dimensional structure applicable to heart comprising a cell derived from a part other than myocardium of an adult wherein the cell is an embryonic stem cell a mesenchymal stem cell induced pluripotent stem cells or synovial cells the synovial cells derives from internal surface of the joint and contains stem cells and the three dimensional structure comprises a cell sheet having biological connection between cell sheets is free from scaffold and is obtained by the process comprising a culturing the cell derived from the part other than myocardium of an adult on a cell culture support grafted with a temperature responsive macromolecule having an upper limit critical solution temperature or lower limit critical solution temperature to in an aqueous solution of from 0 C to 80 C b setting a culture medium temperature to the upper limit critical 99 solution temperature or more or the lower limit critical solution temperature or less and c detaching the cultured cell as a three dimensional structure 2 8 Cancelled 9 Previously Presented The structure according to claim 1 wherein the cell is derived from a subject the structure being applied to the sub ject 10 Previously Presented The structure according
276. 3 EO Rem at TER E nu xe 3 3 4 3 1 9 6 3 13 6 2 2009 8 20 2010 2 22 2010 9 22
277. ES 3 PA6 3 3 2 D 2 4 ES MEF 135 R PE 3 P A6 ES PA6 6 6 RPE
278. 2011 3 30 1 Oct KIf MWyc Sox 2 Oct KIf Sox 3 1 2 1 Oct KIf MWyc Sox 2 1 4 1 2 1 Oct KIf Sox
279. ES RPE RPE B 2 E S R PE ES 3 R PE 3 P a x 6 RAR PE Pa x 6 1 2 C 2 3 PA 6
280. 1 9 1 1 8 2 0 1 4 2 1 2 0 2 2 RPE RPE Oct 4 Sox2 TDGF 1 RPE RPE R PE 5 0 MEF 50 E B MEF DMEM 2mM Glu taMAX II 500 g ml 500 g ml1 1 6 FCS EE DMEM 500 g ml 500g ml
281. 18 1 HIRE ZIBEEXIOEM SS 57ikc oc ILD MES MIC ARS S R PE R PE RPE RPE 1 2 1 1 3 1 4 3 5 2 6 r d RPE 6 5 t ubby RCS Abyssinian c at c d Tp r c d e r d 1 2 3 rcd1 rcd 2 r
282. 5 3 7 5 6 3 8 N 5 6 3 9 m 5 6 3 1 0 9 1
283. Diacrin 3 ES 3 3 3 1 2 3 4 5 0007
284. 1 MEF 3 a hES a hES b 1 1 1 1 1 1 10 22 4ng ml _ bFGF 1ng ml LIF 8 4 9 8 496PLASMANATE EB
285. 6 8 2 5 27 RPE 3 2 b 3 5 2 2 7 RPE 3 3 b 1 2 5 2 7 133 RPE 3 4 1 1 1 RPE 3 5 1 1 1 RPE 3 6 Starg
286. 1 7 1 0 b 3 5 1 7 1 1 b 1 1 1 1 1 2 1 1 1 R PE 1 3 1 1 1 R PE 1 4 1 1 1 RP E
287. 2 3 4 iPS VEGF IiPS
288. 2 3 2 2 2 4 a 0 8 0 b c 2 5 2 4 2 6 N
289. 24 2 2 7 3 3 1 7 2 5 1 2 6 5 5 3 1 3 29 2 3 150 3 E S R PE Abstract 1583 2 1584 RAS 2 MEF ES E S ES 10 5 0 PA6
290. Opt iPRO SFM MDBK GM 150 g mL WO 22009 0 51671 hES RPE TH 6 8 100 2 0 ESC RPE 9 R PE MEF ES
291. Supplementary Table 1 online MSC 1 3B Effect of monolayered MSC on cardiac function 1 Kaplan Meier 4 MSC Fig 5h MC MSC 4 MSC 72 1009 719 long rank test P lt 0 05 1 3 Survival analysis 1 MSC 1
292. ii 6 6 2012 7 23 6 1 1 1 3 3 0 80
293. 1 8 10 1 3 LV UCG T C I ADE LV 3 3 A D CM 3 3B
294. 1 2 Journal of Artificial Organs 2002 vol 5 pp 216 222 3 W002 08387 1 1 2 3 ii 2 2
295. 1 2 4 Sox 2 3 5 Myc Myc 1 4 1 6 bFGF SCF 5 7 TERT 1 6 1 8 SV40 Large T antigen HPV16E6 HPV16E7 Bmil 1 1 7 1 9 Fbx15 Nanog Eras ECAT15 2 Tcl 1 catenin 1 1 8 1 1 0 ECAT 1 Esg 1 Dnmt 3L ECAT 8 Gdf 3 Sox15 ECAT15 1 FthI17 Sall 4 Rex 1 UTF 1 Stella Stat3 Grb2
296. 2 5 1 7H x Xm Wm gt HH cH m BE XE tt HE Fr d dd E IX Ht n 24 4 2 4 nul CL I iPS iPS HI 1 iPS EP IPS
297. P a x RP E Pax R PE Irina Klimanskaya Nature Review Kawasaki E S RPE R PE 178 3 Irina Klimanskaya RATE
298. 2003 8 1 6 6 PCT JP2004 001034 2004 2 2 1 FY 184 RO 14
299. 3 Abstract 1 1793 1794 1 3 R PE 1 4 3 Fig 5 6 1 1794 RAIS 2 RPE 1 3 ES RPE R PE
300. c R PE R PE 2 8 2 5 2 7 R PE 2 9 b h ES FGF L IF PLASMANATE 2 2 7 RPE 3 0 119 b 6 gt 25 27 RPE 3 1 b 6 8 2 5 27 RPE 3 2 b 3
301. 103 1 11 Soejima Li Shimizu Kikuchi Hirose 10325 2006 8 16 112 1 12 13 112 2 myocardium like RU stockinet like 102 D p Eppenberger Akin
302. 3 1 1 2 2 1 3 2012 6 27 1 17 2 3 24 4 6 1 8
303. 9 4 2 2 2009 5 27 4 10 12 13 3 1 4 3 2 5 6 3 7 8 D1 D2 D4 D6 D3 4 7 8 10 1 2 11 3
304. EB b c 12 BY PO d 3 e RPE 8 1 0 1 9 1 1 RPE RPE 8 1 2 RPE R PE b FGF FGF 1 1 1 3 R PE ES RPE
305. c R PE RPE 2 6 2 2 RPE hES RP E a hES b c R PE RPE 2 7 2 2 CR ORHANERPEMMCH IC h E SAAR P E a hES
306. 2003 7 15 2006 2 15 2006 8 15 2006 11 15 non final OA 35 USC 112 1st 2nd 102 a b 103 final OA 35 USC 102 a 103 20 2007 5 H 15H Notice of Appeal 2007 10 A 3H Advisory Action 2007 410 H15 AE RCE 20084 1 9 non final OA 35 USC 101 112 1st 102 b 103 2008 7 9 TIER 2008 7 11 2008 10 15 non final OA 35 USC 112 1st 103 2009 4 15 2009 4 16 2009 8 20 final OA 35 USC 103 2010 2 22 Notice of Appeal 2010 4 6 Advisory Action 2010 9 22 RCE 2010 12 23 non final OA 35 USC 112 1st 103 2011 6 22 2012 3 8 Notice of Appeal 2012 10 5 RCE 3 2 2 PCT JP2001 005722 2001 7 A 2 13 1
307. 5 2 5 2 7 R PE 3 3 b 1 2 5 2 7 RPE 3 4 1 1 1 RPE 3 5 1 1 1 RPE 3 6 Stargard t 3 4 3 7 3 4
308. 8 10 1 8 10 1 8 10 3 4 1 1 Suzuki 6
309. ES RPE p lk 106 kk 106 4 4 3 ACT kk 107 4 5 ACT 20 0 X OTT 108 551392 PCTYUS2005 002273 WO2005 07 T To PM NS 108 0011 OREB t ueri Mee MA nea 108 4 5 1 108 4 5 2 109 4 5 3 kk 111 4 5 3 kk 114 4 5 4 kk 115 4 5 5 AARIIN a aot cv ra cde 122 4 5 6 122 4 5 7 124 2x5 E AAEE ONNE Pe PP Pa ada ee 4 129 4 5 9 kk 135 4 5 10 148 4 5 11 ki 149 4 5 12 152 4 6 ACT 77 3 6 8 9 6 161 4 6 1 161 4 6 2
310. RPE 6 5 tubby RCS Abyssini an cat REE c dq prcd e r dq 1 2 3 rcd1 111 rcd2 r cd 3 p d J Briard RPE 6 5 5 7 1 A 6 8 hES RPE RPE R PE a MEF hES b h ES c hES
311. 1 103 Kawasaki Thomson 1 0 3 Claim Rejections 85 USC 103 The following is a quotation of 35 U S C 103 a which forms the basis for all obviousness rejections set forth 1n this Office action a A patent may not be obtained though the invention is not identically disclosed or described as set forth in section 102 of this title if the differences between the subject matter sought to be patented and the prior art are such that the subject matter as a whole would have been obvious at the time the invention was made to a person having ordinary skill in the art to which said subject matter pertains Patentability shall not be negatived by the manner in which the invention was made Claims 19 20 and 22 are rejected under 35 U S C 103 a as being unpatentable over Kawasaki et al PNAS 99 3 1580 1585 February 5 2002 when taken with Thomson et a7 Science 282 1145 1148 November 6 1998 The breadth of the claims encompass culturing hES cells under any condition to provide RPE like cells Thus the combination of the cited art is proper because it provides conditions
312. 10181984 5 43 3 2 7 10 333473 45 3 2 8 kk 54 3 3 2 kk 55 EEC 55 3 3 2 PCT JP2004 001024 57 3 3 3 2006 519056 3 3 4 2009 223536 pe 75 3 3 5 04707319 2 sees 86 3 3 6 04 567728 96 imi REC I NUT 101 Sd EO NP ANE PRES EN 102 4 RPE H e RU mih Ae A A RR M Zr ec cae toh 102 EO ee a ad 102 4 2 103 dee BT esed iM LL AA cL EE EM 103 De DD NK PS 103 4 3 eee 104 4 4 RPE Leu 105 4 4 1 Lik 105 4 4 2 ACT
313. 104 i PEDF iii iv R PE R PE R PE AMD 2 e 2005 2007 2 4 4
314. 2 3 rcdi rcd 2 r c d3 p d B riard RPE 6 5 1 7 1 9 1 1 8 2 0 131 1 4 2 1 2 0 2 2 RPE RPE Oct4 Sox2 TDGF1 RPE R PE R PE 5096MEF 5 09 E MEF DMEM 2m
315. 2008 11 5 1 29 1 13 13 2 32 11 11 11 3 3 1 3 1 1993 144 4 495 500 1 3 1
316. 26 A method according to claim 24 wherein the temperature responsive macromolecule is poly N isopropylacrylamide 1 2 8 10 17 22 23 Diacrin US6432711 1 5 6 9 17 22 23 Diacrin WO01 07568 1 6 10 16 17 22 23 Kellar Circulation vol 104 no 17 pp 2063 2068 18 21 24 26 Diacrin US6432711 Circulation Research 2002 vol 90 no 3 e40 e48 http www j circ or jp engl ish sessions reports 66th ss okano htm e 2006 1 A 30 2003 3 28 67
317. 4 1 2 0052 h E SARE EB 6 8 EB 1 29 1B EB 3 EB 1D EB 1D 0061 5 R PE BOT bFGF fYAUY TGF B IBMX bmp 2 bmp 4 1 EB ES RPE E CM I IV Matrigel ES RPE Pax6 Pax 2 mitf RPE CRALBP PEDF REPG65
318. ERas ECAT15 2 Tcl1 catenin 13 1 1 8 1 0 ECAT1 Esgl Dnmt3L ECAT8 Gdf3 Sox15 ECAT15 1 FthlI7 Sall4 Rexi UTF1 Stella Stat3 Grb2 1 1 9 11 KIf4 KIf2 1 10 12 Sox2 Sox1 Sox3 Sox15 Sox17 1 10 13 c Myc L Myc N Myc 1 1 0 14 1 13 1 Olswang LLP 2 1 1
319. bFGF FGF 1 1 1 3 R PE ES RPE 112 1 R PE 1 1 1 4 R PE R PE a 109 b PEDF EPC 1 1 5 1 4
320. h ES RPE RPE 000 RPE R PE B 3 1 3 RPE RPE Pa x 6 155 RPE Pax6 RPE Pa x 6 B 4 1
321. 10 18 200845022 7H 2008 07 02 Non Final Rejection 2008 11 17 2008 12 04 2009 02 03 Final Rejection 2009 06 29 2009 08 05 Non Final Rejection 2009 08 17 IDS 2010 O1 20 2010 02 04 IDS 2010 03 24 NOA 2010 04 20 2010 O4 30 2010 05 03 2010 O5 2 6 4 6 2 1 Amethod of treating or preventing retinal degeneration comprising use of a cell selected from the group consisting of at least one of RPE cells RPE like cells RPE or RPE like progenitors derived from mammalian embryon
322. 11 A myocardium like tissue produced by a method in which the myocardium like cell sheet or three dimensional structure of myocardium like cells according to claim 1 is buried in the living body 46 so that tubular cavities are formed of vascular endothelial cells endogenous in said myocardium like cell sheet or three dimensional structure of myocardium like cells and or the vascular endothelial cells in the tissue around the buried graft are allowed to grow inward and form tubular cavities thereby forming blood vessels 12 The myocardium like cell sheet or three dimensional structure of myocardium l ike cells according to claim 1 which are suitable for use in the treatment of heart disease and other circulatory organ related diseases or digestive organ related diseases 13 A method of treating heart disease and other circulatory organ related diseases or digestive organ related diseases using the myocardium like cell sheet or three dimensional structure of myocardium like cells according to claim 1 2 2006 2 15 12 13 112 1 1
323. 13 112 2 1 12 Eppenberger FASEB vol 13 Supplement 1999 p S83 89 Oyamada Exp Cell Res 1994 p 351 357 102 b 2 Akins 5 Tissue Engineering 1999 vol 5 p 103 115 1 10 12 13 Shimizu Cardiac tissue engineer ing 2000 10 21 A 19525 35 p 203 204 102 3 1 10 12 13 Kikuchi J Biomater Sci Polymer Edn 1998 vol 9 No 12 p 1331 1348 Hirose Biomecromolecules vol 1 2000 p 377 381 Eppenberger Akins 103 1 10 12 13 Shimizu Kikuchi Hirose Eppenberger 103 1 11 Soejima Plastic and Reconstructive Surgery 1998 p 1552 1560 Li D Circulation Research vol 78 1996 p 283 288 Kikuchi Hirose Eppenberger Akins 47
324. 15 2000 227 2 271 8 cited by other Thomson et al Embryonic stem cell lines derived from human blastocysts Science Nov 6 1998 282 5391 1145 7 Erratum in Science Dec 4 1998 282 5395 1827 cited by other Schuldiner et al Effects of eight growth factors on the differentiation of cells derived from human embryonic stem cells Proc Natl Acad Sci U S A Oct 10 2000 97 21 11307 12 cited by other Kehat et al Human embryonic stem cells can differentiate into myocytes with structural and functional properties of cardiomyocytes J Clin Invest Aug 2001 108 3 407 14 cited by other Carpenter et al Enrichment of neurons and neural precursors from human embryonic stem cells Exp Neurol Dec 2001 172 2 383 97 cited by other Binder et al Transplantation of autologous retinal pigment epithelium in eyes with foveal neovascularization resulting from age related macular degeneration a pilot study Am J Ophthalmol Feb 2002 133 2 215 25 cited by other http www answers com topic embryonic stem cell 1 2008 cited by other Klimanskaya et al Derivation and Comparative Assessment of Retinal Pigment Epithelium from human Embryonic Stem Cells Using Transcriptomics Cloning and Stem Cells 6 217 245 2004 cited by other Lund et al Human Embryonic Stem Cell Derived Cells Rescue Visual 176 Function in Dystrophic RCS Rats Cloning and Stem Cells 8 3 189 199 2006 ci
325. ii ii ES R PE 3 2 6 1 2 1 4 4 2 4 2 2 7 4 5 11
326. 1791 1795 1 21 Kawasaki H et al PNAS 20024 V ol 99 No 83 pp 1580 1585 2 3 Haruta M et al Invest Ophthalmol Vis Sci 2004 Mar 45 3 1020 1025 4 2 p75 SP12 2 2003 2 28 5 Invest Ophthalmol Vis Sci 200 3 44 E Abstract 381 2003 ARVO 20034 5 1 6 Thomson JA etal Biol Reprod 5 5 254 259 1996 7 2003 523766 85 Kawamorita M et al Hum Cell 2002 Sep 15 3 178 182 9 Subramanian T Semin Neurol 20 OT 227 X eg 9915 10 WO2011 063005 1 4 2 200 3 3 12 2 2ODF i g5D Pa x 6 ii 1 1 4 2 1 9 1 2 2 9 1 3
327. 2 3 2 50 5 6 3 0 80 1 2
328. 6 7 7 6 2003 6 p 50 1 4 7 7 6 5 2 3 iv Suzuki 6 Ahmad 4
329. 6 5 1 9 2 1 113 15 NILEREOTHMMR RMB AMARA KURR A 2 0 4 5 3 3 7 1 2 9 1 3 H2 3 29 2 1 3 7 2 2 9 1 3 3
330. 7 19 21 1 2 1 1793 1794 RAIS 1 ERIT IX Wi RPE ES ES RCS 2 Abstract 1583 RAHI 2 1584 2 ES 1 2
331. R PE 4 5 7 2 9 1 3 1 29 2 2 3 6 6 1 3 6 6 2 A 1 2 9 1 3 2
332. SDIA stromal cell derived inducing activity see Abstract Claim 19 is drawn to a method involving spontaneous differentiation of hES cells to pigmented epithelial cells Since the combined publications do not anticipate every limitation of claim 19 claim 19 and its dependent claims are not rendered obvious by the cited publications Next we turn to the differences between the cells produced by the claimed methods and the cells produced by Kawasaki et al There is no indication that the method of Kawasaki et al produced non human RPE65 bestrophin Pax6 pigmented epithelial cells similar to the cells produced by claim 19 Kawasaki et al state Consistently pigmented epithelial cells from primate ES cells were positive for the optic cup marker Pax6 page 1584 column 1 As the instant specification states substantial Pax6 expression is characteristic of early RPE progenitors rather than mature RPE or RPE like cells Pax6 at earlier stages acts as an activator of proneural genes and is downregulated in the RPE in further development paragraph 63 emphasis added In contrast to Kawasaki et al claim 19 requires that the RPE like culture comprise RPE65 bestrophin Pax6 pigmented epithelial cells Thus the cells of Kawasaki et al are not the equivalents of the cells of claim 19 Consequently one of skill in the art attempting to combine Thompson et al and Kawasaki et al would not have produced the cells described in c
333. after being peeled off from the substrate 3 A process for producing a myocardium like cell sheet which comprises culturing cells which have been obtained from a heart on a cell culture support having a substrate surface coated with a temperature responsive polymer whose upper or lower critical solution temperature in water is 0 SIMILAR 80 DEG C and subsequently 1 bringing the temperature of the culture solution to above the upper critical solution temperature or below the lower critical solution temperature 2 peeling the cell sheet off from the culture support wherein the cell sheet peeled off fro the culture support shringks and has no contamination by a third substance 4 The process for producing a myocardium l ike cell sheet according to claim 3 wherein the step of peeling off does not involve treatment with a proteolytic enzyme 5 The process for producing a myocardium l ike cell sheet according to claim 3 wherein the temperature responsive polymer is poly N isopropylacrylamide 6 The process for producing a myocardium l ike cell sheet according to claim 3 wherein the cell sheet is peeled off from the culture support together with a polymer membrane selected from the group consisting of a hydrophilized polyvinylidene difluoride membrane polyurethane a Spandex mesh and a stockinet like material 7 The myocardium like cell sheet according to claim 1 or 2 for use in a 44 method of treatment of heart disease and o
334. by other Hirano M et al Generation of Structures Formed by Lens and Retinal Cells Differentiating From Embryonic Stem Cells Developmental Dynamics Wiley Liss Inc New York NY vol 228 No 4 Dec 2003 pp 664 671 cited by other Ooto S etal Induction of the Differentiation of Lentoids from Primate Embryonic Stem Cells Investigative Opthamology amp Visual Science Association for Research in Vision and Opthamology vol 44 No 6 Jun 2003 pp 2689 2693 cited by other Zhao X et al Differentiation of Embryonic Stem Cells Into Retinal Neurons Biochemical and Biophysical Research Communications vol 297 No 2 Sep 2002 pp 177 184 cited by other Ying Q L et al Conversion of Embryonic Stem Cells Into Neuroctoderma Precursors in Adherent Monoculture Nature Biotechnology Nature Publishing Group New York NY vol 21 No 2 Feb 2003 pp 183 186 cited by other Reubinoff B et al Embryonic Stem Cell Lines From Human Blastocysts Somatic Differentiation In Vitro Nature Biotechnology Nature Publ ishing Group New York NY vol 18 No 4 Apr 2000 pp 399 404 cited by other Lund R D et al Cell Transplantation As A Treatment for Retinal Disease Progress in Retinal and Eye Research vol 20 No 4 Jul 2001 pp 415 449 cited by other Supplementary European Search Report In application No EP05711960 4 mailed Jun 3 2009 cited by other Marmorstein et al Be
335. c d3 p d B riard RPE 65 5 7 1 A 6 8 109 h ES RPE RPE RPE a MEF hES b h ES c hES d RPE RPE R PE 9 R PE a hES
336. critical solution temperature 2 bringing the cultured cell sheet into close contact with a polymer membrane or mesh 3 peeling the cell sheet off together with the polymer membrane or mesh without using a protelystic enzyme to obtain a myocardium l ike cell sheet and 4 producing a three dimensional structural of myocardium like cell sheet by allowing the thus obtained myocardium like cell sheet to adhere to another myocardium like cell sheet which is adhered to a cell culture support not coated with a temperature responsive polymer or to a cell culture support coated with a temperature responsive polymer or to a cell culture support coated with a temperature responsive polymer whose upper or lower critical solution temperature in water is 0 80 C and repeating this process as appropriate thus piling up two or more cell sheets 2 he process for producing a three dimensional structure according to claim 1 wherein the three dimensional structure has been oriented in a specified direction by stretching them in the specified direction after it was peeled off from the substrate 3 The process for producing a three dimensional structure of myocardium like cells according to claim 1 wherein the 42 temperature responsive polymer is poly N isopropylacrylamide 4 The process for producing a three dimensional structure of myocardium like cells according to claim 1 wherein the polymer membrane is selected from among a hydrophi ized pol
337. culturing said multilayer population of hES cells under conditions that do not maintain the undifferentiated state of said hES cells for a sufficient time to al low for the appearance of putative human RPE cells wherein said putative human RPE cells comprise brown pigment dispersed within their cytoplasm c 169 selecting one or more of said putative human RPE cells from the culture of step b to obtain human RPE cells and d culturing said human RPE cells obtained in step c to form a cell monolayer containing cells that are Pax6 and bestrophin and exhibit a characteristic cobblestone polygonal epithelial like appearance and compr ise brown pigment dispersed within their cytoplasm thereby producing an enriched population of human RPE cells 2 he method of claim 1 wherein said culturing in step b comprises culturing the multilayer population of hES cells in media lacking exogenously added FGF 3 The method of claim 1 wherein said culturing in step b comprises culturing the multilayer population of hES cells in media lacking exogenously added FGF and lacking exogenously added LIF 4 The method of claim 1 wherein said culturing in step b comprises culturing the hES cells in media lacking exogenously added FGF and lacking exogenously added LIF and lacking exogenously added PLASMANATE RTM 5 The method of claim 1 wherein the duration of culturing in step b is about 6 weeks 6 The method of claim 1 wherein the dura
338. exogenously added PLASMANATE RTM 4 6 12 U S Patent Documents 6642048 November 2003 Xu et al 7247479 July 2007 Kochanek et al 2002 0022268 February 2002 Xu et al 2003 0087859 May 2003 Kochanek et al 2004 0018617 January 2004 Hwang et al 2006 0031951 February 2006 KI imanskaya 2007 0031386 February 2007 Kl imanskaya Foreign Patent Documents W0 98 30679 Jul 1998 WO Other References Gepstein Cir Res 91 866 876 2002 cited by examiner Motohashi Pigment Cell Res 19 284 289 2006 cited by examiner Zhou Genome Res 12 1716 1722 2002 Abstract cited by examiner Kawasaki et al PNAS 99 3 1580 1585 Feb 5 2002 cited by examiner Thomson et al Science 282 1145 1147 Nov 6 1998 cited by examiner Plasmanate RTM product information Accessed online at http www bdipharma com Products Albumin P asmanate PPF aspx on Mar 19 2010 cited by examiner Hori et al Growth inhibitors promote differentation of insulin producing tissue from embryonic stem cells PNAS vol 99 No 25 pp 16105 116110 2002 cited by other Lu et al Generation of functional hemangioblasts from human embryonic stem cells Nature Methods vol 4 No 6 pp 501 509 2007 cited by other 174 Ohno Matsui et al In vitro and in vivo characterization of iris pigment epithelial cells cultured on amniotic membranes Molecular Vision vol 12 pp 1022 1032 2006 cited
339. plurality of the assemblies are piled up wherein the step of peeling off does not involve treatment with a proteolytic enzyme 8 The process for producing a myocardium like cell sheet according to a process for producing a three dimensional structure of myocardium l ike cells in which the myocardium like cell sheet as obtained in claim 5 is again allowed to adhere to a cell culture support a cell culture support coated with a temperature responsive polymer a polymer membrane or a cellular sheet and a plurality of the assemblies are piled up wherein the temperature responsive polymer is poly N isopropylacrylamide 9 The process for producing a myocardium like cell sheet according to claim 5 or the process for producing a three dimensional structure of myocardium like cells according to a process for producing a three dimensional structure of myocardium like cells in which the myocardium l ike cell sheet as obtained in claim 5 is again allowed to adhere to a cell culture support a cell culture support coated with a temperature responsive polymer a polymer membrane or a cellular sheet and a plurality of the assemblies are piled up wherein the polymer membrane is selected from among a hydrophi ized polyvinylidene difluoride membrane polyurethane a Spandex mesh and a stockinet like material 10 A myocardium like cell sheet or a three dimensional structure of myocardium like cells that are produced by the process according to claim 5
340. that result in RPE like cells Kawasaki teach culturing monkey ES cell lines with PA6 cells to produce RPE cells With regard to step c of claim 19 although Kawasaki teach mechanically isolating pigmented epithelium patches p 1581 col 2 Culture of Pigmented Epithelium they also teach methods of dissociating by trypsin p 1581 col 2 Sorting Neural Cells and these methods would be well known to the skilled artisan to employ in isolating pigmented epithelial cells 165 With regard to the limitation in claim 20 Kawasaki teach that the pigmented cells could be replated at least twice p 1581 col 2 Culture of Pigmented Epithelium With regard to the limitation in claim 22 Kawasaki teach that the cells were cultured in 10 knockout serum replacement p 1581 col 1 1st 4 Kawasaki do not specifically teach using their methods for human ES cells However prior to the time of the claimed invention Thomson teach the generation of human ES cells and suggest the need in the art for directing the differentiation of hES cells to cell types of interest See p 1146 col 3 last f Accordingly in view of the combined art it would have been obvious for one of ordinary skill in the art to modify the techniques as taught by Kawasaki to produce human RPE cells from human ES cells with a reasonable expectation of success One of ordinary skill in the art would have been motivated to make this modification in view of Thom
341. to claim 1 wherein the cell is not derived from a subject the structure being applied to the sub ject 11 15 Canceled 16 Previously Presented The structure according to claim 1 wherein the cell derived from a part other than myocardium is a cell not derived from heart 17 Canceled 18 Previously Presented The structure according to claim 1 comprising a monolayer cell sheet 19 Previously Presented The structure according to claim 1 comprising a multilayer cell sheet 20 Previously Presented The structure according to claim 19 wherein the multilayer cell sheet has biological connection between cell sheets 21 Previously Presented The structure according to claim 20 wherein the biological connection is selected from the group consisting of connection via extracel lular matrix electrical connection and connection without scaffold 22 Canceled 23 Currently Amended A medicament according to claim 1 which is used for treatment of a heart and wherein the heart has a disease or disorder selected from the group consisting of ischemic heart disease myocardial infarct and dilated cardiomyopathy 24 Withdrawn A method for producing a three dimensional structure applicable to heart comprising a cell derived from a part other than myocardium of an adult the method comprising the steps of a culturing the cell derived from the part other than myocardium of an adult on a cell culture support grafted with a temperature
342. 05 0 0 8 P CTYJP2008 050305 2008 1 1 1 WO2008 2087917 2008 7 2 7 4 4 2 ACT ES RPE ACT R PE 1 US7736896 amp 2010 6 15 US7794704 2010 9 14 US7795025 2010 9 14 1 2006 551392 2005 1 2 4 2007 522131 2007 8 9 ACTH
343. 1 12 16 Notice of Appeal 2012 7 16 RCE e PCT JP2004 001024 2004 2 2 26 1 A three dimensional structure applicable to heart comprising a cell derived from a part other than myocardium of an adult 2 A structure according to claim 1 wherein the cell is a stem cell or a differentiated cell 3 A structure according to claim 1 wherein the cell is a mesenchymal cell 4 A structure according to claim 1 wherein the cell is derived from a myoblast 5 A structure according to claim 4 wherein the myoblast is a skeletal myob last 6 A structure according to claim 1 wherein the cell is a fibroblast A A structure according to claim 1 wherein the cell is a synovial cell 8 A structure according to claim 1 wherein the cell is derived from a stem cell 9 A structure according to claim 1 wherein the cell is derived from a subject the structure being applied to the subject 10 A structure according to claim 1 wherein the cell is not derived from a subject the structure being applied to the subject 11 A structure according to claim 1 wherein the structure expresses at least one non adult heart marker selected from the group consisting of myosin heavy chain Ia myosin heavy chain Ib myosin heavy chain IId I 57 x CD56 MyoD Myf5 and m
344. 10 The structure according to claim 4 wherein the cell derived from a part other than myocardium is a cell not derived from heart 11 The structure according to claim 4 wherein the applicability to heart includes applicability to myocardium 12 The structure according to claim 4 comprising a monolayer cell sheet 13 The structure according to claim 4 comprising a multilayer cell sheet 14 The structure according to claim 13 wherein the multilayer cell sheet has biological connection 15 The structure according to claim 14 wherein the biological connection is selected from the group consisting of connection via extracellular matrix electrical connection and connection without scaffold 16 A medicament for use in a method of treating dilated phase hypertrophic cardiomyopathy or ischemic heart disease comprising a three dimensional structure according to any one of claims 4 to 15 10183361 4 1 Athree dimensional structure for use in a method of implanting into a heart of human to treat dilated phase hypertrophic cardiomyopathy or ischemic heart disease comprising a cell derived from a part other than myocardium of an adult wherein the three dimensional structure is free from a scaffold and has at least 6 cm area and at least 50 um thickness wherein the three dimensional structure shrinks when being detached from an cell culture support and wherein the cell is a myoblast 2 A structu
345. 5 months 172 27 The method of claim 22 wherein said media containing exogenously added FGF in step a further comprises exogenously added LIF 28 The method of claim 22 wherein said media containing exogenously added FGF in step a further comprises exogenously added PLASMANATE RTM 29 A method for producing an enriched population of human retinal pigment epithelium RPE cells the method comprising a providing a culture of human embryonic stem hES cells b culturing the hES cells to produce one or more embryoid bodies wherein said one or more embryoid bodies or the cells from which said one or more embryoid bodies are formed have been cultured in media containing exogenously added FGF c culturing said one or more embryoid bodies for a sufficient time for the appearance of putative human RPE cells within at least one of said one or more embryoid bodies wherein said putative human RPE cells comprise brown pigment dispersed within their cytoplasm wherein the one or more embryoid bodies are cultured in a media lacking exogenously added FGF and the duration of culturing is at least 6 weeks whereby one or more embryoid bodies containing putative human RPE cells are formed d selecting and dissociating one or more of said embryoid bodies containing putative human RPE cells from the culture of step c to obtain human RPE cells and e culturing said human RPE cells obtained in step d to form a cell monolayer containing cell
346. 6 7 1 9 27 8 RPEG65 2 7 PE DF 2 9 3 0 1 18 Canceled 19 Currently Amended A method for producing a population of cells enriched for RPE65 bestrophin Pax6 pigmented epithelial cells comprising a culturing hES cells in medium that supports spontaneous differentiation of hES cells to pigmented epithelial cells b selecting the cells of step a that exhibit pigmented epithelial morphology c passaging for a sufficient time or number passages the cells selected in step b using an enzyme selected from the group consisting of trypsin collagenase IV collagenase I and dispase to enrich for RPE65 bestrophin Pax6 pigmented epithelial cells 20 Previously presented The method of Claim 19 wherein the passaging of cells in step c is repeated at least twice 168 26 Canceled 27 Currently Amended The method of claim 19 wherein the RPE65 bestrophin Pax6 pigmented epithelial cells are enriched for CRALBP cells and PEDF 28 Currently Amended The method of claim 19 wherein the RPE65 bestrophin Pax6 pigmented epithelial cells have an absence of at least one ES cell marker selected from the group consisting of Oct4 or Sox2 29 New The method of claim 19 wher
347. 6 weeks c selecting one or more of said putative human RPE cells from the culture of step b to obtain human RPE cells and d culturing said human RPE cells selected in step c to form a cel monolayer containing cells that are Pax6 bestrophin CRALBP PEDF and express RPE65 have the absence of at least one ES cell marker selected from the group consisting of Oct4 and Sox2 and exhibit a characteristic cobblestone polygonal epithelial like appearance and compr ise brown pigment dispersed within their cytoplasm wherein during said culturing the cultured cells temporarily lose their epithelial appearance and pigmentation after plating and then regain their epithelial appearance and pigmentation upon further culturing thereby producing an enriched population of human RPE cells 23 The method of claim 22 wherein said culturing in step b comprises culturing the multilayer population of hES cells in media lacking exogenously added FGF and lacking exogenously added LIF 24 The method of claim 22 wherein said culturing in step b comprises culturing the multilayer population of hES cells in media lacking exogenously added FGF and lacking exogenously added LIF and lacking exogenously added PLASMANATE RTM 25 The method of claim 22 wherein the duration of culturing in step b is between about 6 weeks and about 8 weeks 26 The method of claim 22 wherein the duration of culturing in step b is between about 3 months and about
348. M GlutaMAX 1I 500g ml 500g ml 1 6 FCS EB DMEM 500 gm1 500 g ml 19 2 mM GlutaMAX 1 0 1mM8 RPE 2 3 RPE P a x 6 CRALBP PEDF RPE65 2 2 RPE 2 4 RPE cobbleston e 2 2 RPE 2 5 2 2 RPE hES R PE a MEF hES h ES b
349. PS iZUMI iPIERIAN iPIERIAN iPS 179
350. and a plurality of the assemblies are piled up 7 The process for producing a myocardium like cell sheet according to claim 5 or the process for producing a three dimensional structure of myocardium like cells according to claim 6 wherein the step of peeling off does not involve treatment with a proteolytic enzyme 8 The process for producing a myocardium like cell sheet according to claim 5 or the process for producing a three dimensional structure of myocardium like cells according to claim 6 wherein the temperature responsive polymer is poly N isopropylacrylamide 9 The process for producing a myocardium like cell sheet according to claim 5 or the process for producing a three dimensional structure of 35 myocardium like cells according to claim 6 wherein the polymer membrane is selected from among a hydrophi ized polyvinylidene difluoride membrane polyurethane a Spandex mesh and a stockinet like material 10 A myocardium like cell sheet or a three dimensional structure of myocardium like cells that are produced by the process according to any one of claims 5 to 9 11 A myocardium like tissue produced by a method in which the myocardium like cell sheet or three dimensional structure of myocardium l ike cells according to any one of claims 1 to 4 and 10 is buried in the living body so that tubular cavities are formed of vascular endothelial cells endogenous in said myocardium like cell sheet or three dimensional structure of myo
351. ardt 3 4 3 7 3 4 3 8 r d RPE 65 tu bby RCS Abyssinian c a t MABE lod RX TTA RE Tp r c d er dg 1 2 3 rcd1 rcd 2 r cd3 p d B riard RPE 65 3 7 3 9 1 3 8 4 0 3 4 4 1
352. avitreal y aF GF P BS aFGF PBS bFGF ONL 5 FGF ONL 3 FGF 1 6 RCS 1 2 8 RCS
353. bout 6 weeks 18 The method of claim 13 wherein the duration of culturing in step c is between about 6 weeks and about 8 weeks 19 The method of claim 13 wherein the duration of culturing in step c is between about 3 months and about 5 months 20 The method of claim 13 wherein the cell monolayer of step e contains cells that are Pax6 bestrophin CRALBP PEDF and express RPE65 21 The method of claim 13 wherein the cell monolayer of step e contains cells that are Pax6 bestrophin CRALBP PEDF and express RPE65 and 171 have the absence of at least one ES cell marker selected from the group consisting of Oct4 and Sox2 22 A method for producing an enriched population of human retinal pigment epithelium RPE cells the method comprising a providing a multilayer population of human embryonic stem hES cells wherein said multilayer population of hES cells have been cultured in media containing exogenously added FGF and a fibroblast feeder layer b culturing said multilayer population of hES cells under conditions that do not maintain the undifferentiated state of said hES cells for a sufficient time for the appearance of putative human RPE cells wherein said putative human RPE cells comprise brown pigment dispersed within their cytoplasm wherein said conditions that do not maintain the undifferentiated state of said hES cells comprise media lacking exogenously added FGF and the duration of culturing is at least
354. bular cavities of vascular endothelial cells and or having a single epicardium like outer cell layer 3 The myocardium l ike cultured cell sheet according to claim 1 or the three dimensional structure according to claim 2 which have been peeled off from a substrate without being treated with a proteolytic enzyme 4 The myocardium like cultured cell sheet or three dimensional structure according to claim 1 2 or 3 which have been oriented in a specified direction by stretching them in the specified direction after they were peeled off from the substrate 5 A process for producing a myocardium like cell sheet which comprises culturing cells on a cell culture support having a substrate surface coated with a temperature responsive polymer whose upper or lower critical solution temperature in water is 0 to 80 C and subsequently 1 bringing the temperature of the culture solution to above the upper critical solution temperature or below the lower critical solution temperature and optionally 2 bringing the cultured cell sheet into close contact with a polymer membrane and 3 peeling the cell sheet off together with the polymer membrane 6 A process for producing a three dimensional structure of myocardium like cells in which the myocardium like cell sheet as obtained in claim 5 is again allowed to adhere to a cell culture support a cell culture support coated with a temperature responsive polymer a polymer membrane or a cellular sheet
355. cardium like cells and or the vascular endothelial cells in the tissue around the buried graft are allowed to grow inward and form tubular cavities thereby forming blood vessels 12 The myocardium like cell sheet or three dimensional structure of myocardium like cells according to any one of claims 1 to 4 and 10 or the myocardium like tissue according to claim 11 which are suitable for use in the treatment of heart disease and other circulatory organ related diseases or digestive organ related diseases 13 Use of a myocardium like cell sheet or three dimensional structure of myocardium like cells according to any one of claims 1 4 and 10 or the myocardium like tissue according to claim 11 for the treatment of heart disease and other circulatory organ related diseases or digestive organ related diseases 2 1 2007 7 31 1 lt 3 D1 1993 1425 4 495 500 A 5 7 8 10 D2 1998 1925 173 1748 12 D1 D2
356. ch have been oriented in a specified direction by stretching them in the specified direction after they were peeled off from the substrate 5 A process for producing a myocardium like cell sheet which comprises culturing cells on a cell culture support having a substrate surface coated with a temperature responsive polymer whose upper or lower critical solution temperature in water is 0 to 80 C and subsequently 1 bringing the temperature of the culture solution to above the upper critical solution temperature or below the lower critical solution temperature and optionally 2 bringing the cultured cell sheet into close contact with a polymer membrane and 3 peeling the cell sheet off together with the polymer membrane 6 A process for producing a three dimensional structure of 45 myocardium like cells in which the myocardium ike cell sheet as obtained in claim 5 is again allowed to adhere to a cell culture support a cell culture support coated with a temperature responsive polymer a polymer membrane or a cellular sheet and a plurality of the assemblies are piled up 7 The process for producing a myocardium like cell sheet according to claim 5 or the process for producing a three dimensional structure of myocardium like cells in which the myocardium like cell sheet as obtained in claim 5 is again allowed to adhere to a cell culture support coated with a temperature responsive polymer a polymer membrane or a cellular sheet and a
357. culation 2001 vol 104 Suppl 1 pp 1213 1217 1 1 80 4 29 2 i 2 3
358. e of putative human RPE cells within at least one of said one or more embryoid bodies wherein said putative human RPE cells comprise brown pigment dispersed within their cytoplasm whereby one or more embryoid bodies containing putative human RPE cells are formed d selecting and dissociating one or more of said embryoid bodies containing putative human RPE cells from the culture of step c to obtain human RPE cells and e culturing said human RPE cells obtained in step d to form a cel monolayer containing cells that are Pax6 and bestrophin and exhibit a characteristic cobblestone polygonal epithelial like appearance and comprise brown pigment dispersed within their cytoplasm thereby producing an enriched population of human RPE cells 14 The method of claim 13 wherein the culturing of one of more embryoid bodies in step c comprises culturing the embryoid bodies in media lacking exogenously added FGF 15 The method of claim 13 wherein the culturing of one of more embryoid bodies in step c comprises culturing the embryoid bodies in media lacking exogenously added FGF and lacking exogenously added LIF 16 The method of claim 13 wherein the culturing of one or more embryoid bodies in step c comprises culturing the embryoid bodies in media lacking exogenously added FGF and lacking exogenously added LIF and lacking exogenously added PLASMANATE RTM 17 The method of claim 13 wherein the duration of culturing in step c is a
359. eet in intimate contact with the polymer membrane of 3 is allowed to adhere to another el cut rue support such that the cell sheet contacts to the Celle culture support b a medium is added to peel off the polymer membrane from the cell she 52 t c another myocardium like cell sheet or a three dimensional structure of myocardium like cells in intimate contact with an another polymer membrane is allowed to adhere to the cell sheet of b to form a three dimensional structure of myocardium like cells and wherein the process further comprises d repeating b and c 15 Currently Amended The process according to Claim 6 wherein 4 comprises e the myocardium like cell sheet in intimate contact with the polymer membrane of 3 is turned over and fixed on the cell culture support such that the polymer membrane contacts the cell culture support f another myocardium like sheet or a three dimensional structure of myocardium like cells which is peeled off together with another polymer membrane is allowed to adhere to adhere to the cell sheet of e to form a three dimensional structure of myocardium like cells g a medium is added to peel off the another polymer membrane from the another cell sheet or the three dimensional structure of f h another cell sheet or a three dimensional structure of myocardium l ike cells in intimate contact with another polymer membrane is allowed to adhere to the cell sheet or the three dim
360. ein the bestrophin Pax6 pigmented epithelial cells are enriched for PEDF cells 30 New The method of claim 19 wherein the bestrophin Pax6 pigmented epithelial cells are enriched for RPE65 cells 4 6 8 4 6 9 Non Final Rejection 1 9 2 5 27 30 1 12 1 2 3 25 1 12 2 19 23 No 11490953 8 1 3 4 6 10 1 3 0 31 70 4 6 11 1 A method for producing an enriched population of human retinal pigment epithelium RPE cells the method comprising a providing a multilayer population of human embryonic stem hES cells b
361. ensional structure of g and wherein the process further comprises i repeating g and h 16 Previously Presented The process according to Claim 6 wherein 4 comprises j the myocardium like cell sheet in intimate contact with the polymer membrane of 3 is turned over and k another myocardium like sheet or a three dimensional structure of myocardium like cells which is peeled off together with another polymer membrane is allowed to adhere to the cell sheet of k to form a three dimensional structure of myocardium like cells 17 Previously Presented Athree dimensional structure obtained by the process according to claim 6 wherein the three dimensional structure has tubular cavities formed by vascular endothelial cells and or a single epicardium like outer cell layer 18 Previously Presented A method of treating heart disease 58 comprising grafting the three dimensional structure obtained by the process according to claim 6 to the site of weakened contractile force in a heart of a subject wherein the heart disease is myocardial infarction 19 Withdrawn Currently Amended A method according to claim 18 compr ising grafting the three dimensional structure obtained by the process according to claim 6 to the surrounding of blood vessels in a subject thereby occurring regenesis of blood vessels in the grafted part 3 2 8 1
362. erein the cell derived from a part other than myocardium is a cell not derived from heart 10 The structure according to claim 1 wherein the applicability to heart includes applicability to myocardium 11 The structure according to claim 1 comprising a monolayer cell sheet 12 The structure according to claim 1 comprising a multilayer cell sheet 13 The structure according to claim 12 wherein the multilayer cell sheet has biological connection 14 The structure according to claim 13 wherein the biological connection is selected from the group consisting of connection via extracellular matrix electrical connection and connection without scaffold 15 The structure according to any one of claims 12 to 14 wherein the multilayer cell sheet comprises a cell sheet of a myoblast 95 16 The structure according to claim 15 wherein the myoblast is a skeletal myoblast 17 The medicament comprising a three dimensional structure according to any one of claims 1 to 16 18 The medicament according to claim 17 wherein the heart has a disease or disorder selected from the group consisting of heart failure ischemic heart disease myocardial infarct cardiomyopathy myocarditis hypertrophic cardiomyopathy dilated phase hypertrophic cardiomyopathy and dilated cardiomyopathy 19 A method for producing a three dimensional structure applicable to heart comprising a cell derived from a part other than myocardium of an adult whe
363. from a subject the structure being applied to the subject 94 3 The structure according to claim 1 wherein the cell is not derived from a subject the structure being applied to the subject 4 The structure according to claim 1 wherein the structure expresses at least one non adult heart marker selected from the group consisting of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin 5 The structure according to claim 114 wherein an expression level of the non adult heart marker in the structure is at least 50 of an expression level of the non adult heart marker in skeletal myoblasts 6 The structure according to claim 1 wherein the three dimensional structure expresses all of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain IId ID CD56 MyoD Myf5 and myogenin 7 The structure according to claim 6 wherein an expression level of each of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin in the structure is at least about 50 of an expression level thereof in skeletal myoblasts 8 The structure according to claim 6 wherein an expression level of each of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin in the structure is at least about 100 of an expression level thereof in skeletal myoblasts 9 The structure according to claim 1 wh
364. ic stem cells 2 The method of claim 1 wherein the condition of retinal degeneration is selected from the group consisting of at least one of retinitis pigmentosa and macular degeneration 3 The method of claim 1 further comprising transplantation of the cell by vitrectomy surgery into the subretinal space of the eye 4 The method of claim 3 wherein the cells are transplanted in a suspension matrix or substrate 5 The method of claim 2 wherein the retinitis pigmentosa is associated with an animal model 6 The method of claim 5 wherein the animal model is selected from the group consisting of rd mouse RPE 65 knockout mouse tubby like mouse 162 RCS rat Abyssinian cat cone degeneration cd dog progressive rod cone degeneration prcd dog early retinal degeneration erd dog rod cone dysplasia 1 2 amp 3 rcdl rcd2 and rcd3 dogs photoreceptor dysplasia pd dog and Briard RPE 65 dog 7 The method of claim 6 wherein the outcome of the therapy in the animal model is evaluated using one or more of behavioral tests fluorescent angiography histology and functional testing such as measuring the ability of the cells to perform phagocytosis photoreceptor fragments vitamin A metabolism tight junctions conductivity or evaluation using electron microscopy 8 A method for the spontaneous differentiation of hES cells or embryoid bodies into RPE cells RPE like cells or RPE progenitor cells said method compris
365. in in the structure is at least about 100 of an expression level thereof in skeletal myoblasts 10 The structure according to claim 1 wherein the cell derived from a part other than myocardium is a cell not derived from heart 11 The structure according to claim 1 wherein the heart includes myocardium 12 The structure according to claim 1 comprising a monolayer cel sheet 13 The structure according to claim 1 comprising a multilayer cel sheet 14 The structure according to claim 12 wherein the multilayer cell sheet has biological connection 15 The structure according to claim 13 wherein the biological connection is selected from the group consisting of connection via extracellular matrix electrical connection and connection without scaffold 16 A medicament comprising a three dimensional structure according to any one of claims 1 to 15 17 The medicament according to claim 16 for use in a method for 93 treating dilated phase hypertrophic cardiomyopathy or ischemic heart disease 18 A method for producing a three dimensional structure applicable to heart of a human to treat dilated phase hypertrophic cardiomyopathy or ischemic heart disease in which a scaffold is not used which shrinks and has at least 6 cm area and at least 50 um thickness and which comprises a cell derived from a part other than myocardium of an adult wherein the cell is a myoblast and the method comprising the steps of a cultur
366. ing a allowing hES cell cultures to overgrow on MEF b allowing the hES cell cultures to form a thick multilayer of cells c culturing the hES cells d isolating and culturing the pigmented RPE RPE ike and or RPE progenitor cells from the resultant cell cultures 9 The method of claim 8 wherein the isolating and culturing of PRE I ike cells in step d comprises a digesting the cultured hES cells or embryoid bodies with an enzyme b selectively isolating the pigmented cells c plating the isolated cells on gelatin or laminin for 1 2 days to form primary cultures P0 d continued culturing the primary culture for a period of up to 3 weeks and e isolating the RPE like cells 10 The method of claim 9 wherein the enzyme is selected from the group consisting of one or more of trypsin collagenase and dispase 11 The method of claim 8 wherein the RPE cells are grown to establish a new RPE cell line 12 The method of claim 11 wherein the RPE cell line is differentiated into alternate ineages comprising treatment of the RPE cell line in culture with bFGF or FGF 13 The method of claim 11 wherein the new RPE cell lines varies from the already established RPE cell lines in at least one of the characteristics selected from the group consisting of growth rate expression of pigment de differentiation in culture and re differentiation in culture of RPE like cells when they are derived from different ES cell lines 14 A method f
367. ing the cell derived from the part other than myocardium of an adult on a cell culture support grafted with a temperature responsive macromolecule having an upper limit critical solution temperature or lower limit critical solution temperature to water of from 0 C to 80 C wherein the cell is a myoblast and the temperature responsive macromolecule is poly N isopropylacrylamide b setting a culture medium temperature to the upper limit critical solution temperature or more or the lower limit critical solution temperature or less and c detaching the cultured cell as a three dimensional structure 19 The method according to claim 18 wherein the culture medium temperature in b is set to 20 C to lower limit critical solution temperature to water 20 The method according to claim 19 wherein ascorbic acid or a derivative thereof is added before the detaching step 21 The method according to claim 18 or 19 wherein a treatment using a protein degrading enzyme is not performed in or before the detaching step 10183373 9 1 A three dimensional structure applicable to heart comprising a cell derived from a part other than myocardium of an adult wherein the three dimensional structure is free from a scaffold and wherein the cell is selected from the group consisting of a mesenchymal stem cell a synovial cell and an embryonic stem cell 2 The structure according to claim 1 wherein the cell is derived
368. lagenase I and dispase until pigmented epithelial islands appear or multiply in number and d selecting pigmented or non pigmented cells passaged in step c for establishment of high purity RPE l ike cultures 20 The method of claim 19 wherein the passaging of cells in step c is repeated at least twice 21 The method of claim 19 wherein the selection of cells in step b is a selection of cells that express a nestin or Pax6 neural lineage specific marker 22 The method of claim 19 wherein said medium contains Serum Replacement 23 The method of claim 22 wherein said medium comprises knockout high glucose DMEM supplemented with 500 u ml Penicillin 500 mu g ml streptomycin 1 non essential amino acids solution 2 mM GlutaMAX I O 1 mM beta mercaptoethanol 4 80 ng ml bFGF and 8 4 20 Serum Replacement 24 The method of claim 23 wherein said medium further comprises 10 100 ng ml human LIF 25 The method of claim 23 wherein said medium further comprises Plasmanate 164 26 An isolated RPE or RPE like cell line which varies from established RPE cell lines in at least one of the characteristics selected from the group consisting of growth rate expression of pigment de differentiation in culture and re differentiation in culture 4 6 3 Non Final Rejection Election Restriction gt 19 2 5 1 2 2
369. laim 19 and would thus not have arrived at the method recited in claim 19 Well differentiated cells are clearly preferable to partially differentiated cells in certain applications such as therapeutic transplantations In conclusion claim 19 is non obvious in view of Kawasaki et al and Thompson et al Based on the above arguments and amendments at least one of the legal requirements to establish a prima facie case of obviousness is not met Reconsideration and withdrawal of this rejection on ground of 35 USC 103 a is respectfully requested 4 6 5 1 1 8 19 RPE65 P a x 2 7 28 167 4 6 6 Final Rejection 1 1 2 1 0 3 1 9 20 22 Kawasaki P a x 6 4
370. n 21 A structure according to claim 20 wherein the biological connection is selected from the group consisting of connection via extracellular matrix electrical connection and connection without scaffold 22 A medicament comprising a three dimensional structure according to any one of claims 1 to 21 23 A medicament according to claim 22 wherein the heart has a disease or disorder selected from the group consisting of heart failure ischemic heart disease myocardial infarct cardiomyopathy myocarditis hypertrophic cardiomyopathy dilated phase hypertrophic cardiomyopathy and dilated cardiomyopathy 24 A method for producing a three dimensional structure applicable to heart comprising a cell derived from a part other than myocardium of an 58 adult the method comprising the steps of a culturing the cell derived from the part other than myocardium of an adult on a cell culture support grafted with a temperature responsive macromolecule having an upper limit critical solution temperature or lower limit critical solution temperature to water of from 0 C to 80 C b setting a cul ture medium temperature to the upper limit critical solution temperature or more or the lower limit critical solution temperature or less and c detaching the cultured cell as a three dimensional structure 25 A method according to claim 24 wherein a treatment using a protein degrading enzyme is not performed in or before the detaching step
371. or the derivation of RPE lines or precursors to RPE cells that have an increased ability to prevent neovascularization said method 163 comprising a aging a somatic cell from an animal such that telomerase is shortened wherein at least 10 of the normal replicative lifespan of the cell has been passed and b using the somatic cell as a nuclear transfer donor cell to create cells that overexpress angiogenesis inhibitors wherein the angiogenesis inhibitors can be Pigment Epithel ium Derived Factor PEDF EPC 1 15 The method of claim 14 wherein the somatic cells are genetically modified with exogenous genes that inhibit neovascular ization 16 The method of claim 8 wherein the RPE like cells are derived from a bank of ES or embryo der ived cells with homozygosity in the HLA region such that ES derived cells have reduced complexity of their HLA antigens 17 The method of claim 8 wherein the ES cells are derived from a human 18 A method for the treatment of Parkinson s disease comprising the transplantation of RPE like cells or progenitor cells 19 A method for isolating RPE like cells comprising a culturing hES cells in medium that supports proliferation and transdifferentiation of hES cells to RPE like cells b selecting the cells of step a that exhibit the signs of differentiation along the neural lineage c passaging the cells selected in step b using an enzyme selected from the group consisting of trypsin collagenase IV col
372. poly N isopropylacrylamide 4 A three dimensional structure for use in a method of implanting into a heart of human to treat dilated phase hypertrophic cardiomyopathy or ischemic heart disease which is produced by the method according to any one of claims 1 to 3 5 The structure according to claim 4 wherein the structure is produced by culturing the cell derived from a subject by the method according to any one of claims 1 to 3 and is suitable to be applied to the sub ject 6 The structure according to claim 4 wherein the structure is produced by culturing the cell not derived from a subject by the method according to any one of claims 1 to 3 and is suitable to be applied to the sub ject 7 The structure according to claim 4 wherein the structure expresses at least one non adult heart marker selected from the group consisting of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin 8 The structure according to claim 4 wherein the three dimensional structure expresses all of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain IId ID CD56 MyoD Myf5 and myogenin 9 The structure according to claim 8 wherein an expression level 91 of each of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin in the structure is at least about 100 of an expression level thereof in skeletal myoblasts of a human
373. r Surgery 2003 7 A vol 126 p 114 122 Suzuki BOX 4 5 1 1 Saito Suzuki 3 4
374. re according to claim 1 wherein the myoblast is a skeletal myob last 3 The structure according to claim 1 wherein the cell is derived from a subject the structure being applied to the subject 4 The structure according to claim 1 wherein the cell is not derived 92 from a subject the structure being applied to the subject 5 The structure according to claim 1 wherein the structure expresses at least one non adult heart marker selected from the group consisting of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin 6 The structure according to claim 11 wherein an expression level of the non adult heart marker in the structure is at least 50 of an expression level of the non adult heart marker in skeletal myoblasts 7 The structure according to claim 1 wherein the three dimensional structure expresses all of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain IId ID CD56 MyoD Myf5 and myogenin 8 The structure according to claim 6 wherein an expression level of each of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogenin in the structure is at least about 50 of an expression level thereof in skeletal myoblasts 9 The structure according to claim 6 wherein an expression level of each of myosin heavy chain Ila myosin heavy chain lib myosin heavy chain I d IIx CD56 MyoD Myf5 and myogen
375. rein the cell is selected from the group consisting of a mesenchymal stem cell a synovial cell and an embryonic stem cell and the method comprising the steps of a culturing the cell derived from the part other than myocardium of an adult on a cell culture support grafted with a temperature responsive macromolecule having an upper limit critical solution temperature or lower limit critical solution temperature to water of from 0 C to 80 C wherein the cell is selected from the group consisting of a mesenchymal stem cel l a synovial cell and an embryonic stem cell b setting a culture medium temperature to the upper limit critical solution temperature or more or the lower limit critical solution temperature or less and c detaching the cultured cell as a three dimensional structure 20 The method according to claim 19 wherein ascorbic acid or a derivative thereof is added before the detaching step 21 The method according to claim 19 or 20 wherein a treatment using a protein degrading enzyme is not performed in or before the detaching step 22 The method according to claim 19 or 20 wherein the temperature responsive macromolecule is poly N isopropylacrylamide 3 3 6 04 567728 1 2006 1 31 2009 3 30
376. responsive macromolecule having an upper limit critical solution temperature or lower 100 limit critical solution temperature to water of from 0 C to 80 C b setting a culture medium temperature to the upper limit critical solution temperature or more or the lower limit critical solution temperature or less and c detaching the cultured cell as a three dimensional structure 25 Withdrawn A method according to claim 24 wherein a treatment using a protein degrading enzyme is not performed in or before the detaching step 26 Withdrawn A method according to claim 24 wherein the temperature responsive macromolecule is poly N isopropylacrylamide 2 Currently Amended A method or treating the disease or disorder selected from the group consisting of ischemic heart disease myocardial infarct and di lated cardiomyopathy which comprises the steps of disposing the three dimensional structure according to claim 1 to cover a portion of a heart to be treated and holding the structure for a time sufficient to the portion 28 Previously Presented The method according to claim 27 wherein a surface of the heart is treated with HGF 3 3 7 1 2003 285476 2003 3 28 67
377. rgan related diseases 3 2 6 10181984 5 1 01945762 1 2010 9 29 2 1 2010 5 11 43 2011 7 19 7 1 A myocardium like cultured cell sheet made of myocardial tissue cells that retain contracting and relaxing functions intercellular electrical coupling and orientation wherein the cell sheet is obtainable by peeling off from a cell culture support having a substrate surface coated with a temperature responsive polymer whose upper or lower critical solution temperature in water is 0 80 degrees C without being treated with a proteolytic enzyme and wherein the cell have been obtained from a heart 2 The myocardium l ike cultured cell sheet according to claim 1 which can obtain orientation in a specified direction by being stretched in the specified direction
378. rin D1 D1 Shimizu D4 Okano D5 68 B 21 24 069 27 29 Mid 71 12 13 47
379. s that are Pax6 bestrophin CRALBP PEDF and express RPE65 have the absence of at least one ES cell marker selected from the group consisting of Oct4 and Sox2 and exhibit a characteristic cobblestone polygonal epithelial like appearance and comprise brown pigment dispersed within their cytoplasm wherein during said culturing the cultured cells temporarily lose their epithelial appearance and pigmentation after plating and then regain their epithelial appearance and pigmentation upon further culturing thereby producing an enriched population of human RPE cells 30 The method of claim 29 wherein said culturing in step c comprises culturing the embryoid bodies in media lacking exogenously added FGF and lacking exogenously added LIF 31 The method of claim 29 wherein said culturing in step c comprises culturing the embryoid bodies in media lacking exogenously added FGF and lacking exogenously added LIF and lacking exogenously added PLASMANATE RTM 173 32 The method of claim 29 wherein the duration of culturing in step c is between about 6 weeks and about 8 weeks 33 The method of claim 29 wherein the duration of culturing in step c is between about 3 months and about 5 months 34 The method of claim 29 wherein said media containing exogenously added FGF in step b further comprises exogenously added LIF 35 The method of claim 29 wherein said media containing exogenously added FGF in step b further comprises
380. se hypertrophic cardi omyophthy or ischemic heart disease in which a scaffold is not used and which comprises a cell derived from a part other than myocardium of an adult the method comprising the steps of a culturing the cell derived from the part other than myocardium of an adult on a cell culture support grafted with a temperature responsive macromolecule having an upper limit critical solution temperature or lower limit critical solution temperature to water of from 0 C to 80 C wherein the temperature responsive macromolecule is selected from the group 90 consisting of poly N isopropylacrylamide poly N isopropylacrylamide acrylic acid copolymer poly N i sopropylacrylamidemethylmethacrylate copolymer poly N isopropylacrylamide sodium acrylate copolymer poly N isopropylacrylamide vinyl ferrocene copolymer gannma ray exposed poly vinylmethylether PVME poly oxyethylene and a gel obtained by cross linking the above described macromolecules with a cross linking agent b setting a culture medium temperature to the upper limit critical solution temperature or more or the lower limit critical solution temperature or less and c detaching the cultured cell as a three dimensional structure 2 A method according to claim 1 wherein a treatment using a protein degrading enzyme is not performed in or before the detaching step 3 A method according to claim 24 wherein the temperature responsive macromolecule is
381. son s suggestion of the art recognized need to produce large purified populations of differentiated cells for drug discovery and therapeutic purposes see p 1147 col 1 and Kawasaki s suggesting that the availability of primate pigmented epithelia generated in vitro should facilitate the pathogenic and therapeutic studies of retinal diseases p 1584 col 1 2nd 4 4 6 4 R PE RPE65 P a x 1 0 3 Kawasaki 2 kawasaki RPE Kawasaki R PE Kawasaki ES SD I A PA6
382. strophin the product of the Best vitelliform macular dystrophy gene VMD2 localizes to the basolateral plasma membrane of the retinal pigment epithelium Proc Nat Acad Sci U S A Nov 7 2000 97 23 12758 63 cited by other Fuhrmann et al Extraocular mesenchyme patterns the optic vesicle during early eye development in the embryonic chick Development Nov 2000 127 21 4599 609 cited by other Zaghloul et al Step wise specification of retinal stem cells during normal embryogenesis Biol Cell May 2005 97 5 321 37 cited by other Dunn et al ARPE 19 a human retinal pigment epithelial cell line with 175 differentiated properties Exp Eye Res Feb 1996 62 2 155 69 cited by other Davis et al A human retinal pigment epithelial cell line that retains epithelial characteristics after prolonged culture Invest Ophthalmol Vis Sci Apr 1995 36 5 955 64 cited by other Hu and Bok A cell culture medium that supports the differentiation of human retinal pigment epithelium into functionally polarized monolayers Mol Vis Feb 7 2001 7 14 9 cited by other Zhang et al Invitrodifferentiation of transplantable neural precursors from human embryonic stem cells Nat Biotechnol Dec 2001 19 12 1129 33 cited by other Amit et al Clonally derived human embryonic stem cell lines maintain pluripotency and proliferative potential for prolonged periods of culture Dev Biol Nov
383. t having a substrate surface coated with a temperature responsive polymer whose lower critical solution temperature in water is 0 to 80 C and subsequently 1 bringing the temperature of the culture solution to below the lower critical solution temperature and optionally 2 bringing the cultured cell sheet into close contact with a polymer membr ane wherein the polymer membrane is selected from the group consisting of cellulose and its derivative collagen Japanese paper polyurethane Spandex mesh net like material and stockinet like material and 3 peeling the cell sheet off together with the polymer membrane without being treated with proteolytic enzyme and then 4 the myocardium like cell sheet thus obtained is allowed to adhere to another myocardium like cell sheet or a three dimensional structure of myocardium l ike cells 1 Canceled 8 Previously Presented The process for producing a three dimensional structure of myocardium like cells according to claim 6 in which the myocardium like cell sheet is again allowed to adhere to a cell culture support a cell culture support coated with a temperature responsive polymer a polymer membrane or a cellular sheet and a plurality of the assemblies are piled up wherein the temperature responsive polymer is poly N isopropylacrylamide 9 to 13 Canceled 14 Previously Presented The process according to Claim 6 wherein 4 compr ises a the myocardium like cell sh
384. ted by other Opas et al Formation of retinal pigment epithelium in vitro by transdifferentiation of neural retina cells Int J Dev Biol 45 633 642 2001 cited by other Sauve et al Preservation of visual responsiveness in the superior colliculus of RCS rats after retinal pigment epithelium cell transplantation Neuroscience 2002 114 2 389 401 cited by other Schraermeyer et al Subretinally transplanted embryonic stem cell rescue photoreceptor cells from degeneration in the RCS rats Cell Transplantation 10 673 680 2001 cited by other Van Meurs et al Br J Ophthalmol Jan 2004 88 1 110 3 Autologous peripheral retinal pigment epithelium translocation in patients with subfoveal neovascular membranes cited by other Gouras et al Invest Ophthalmol Vis Sci Oct 2002 43 10 3307 11 Retinal degeneration and RPE transplantation in Rpe65 mice cited by other 4 7 8 ACT PCT US2005 002273 W 02005070011 US7736896 2005 551392 E S RPE Kaw asaki 2002 ES
385. ther circulatory organ related diseases or digestive organ related diseases 2011 12 7 2012 5 10 3 2 7 10 333473 1 2003 7 15 13 4 7 13 1 A myocardium like cultured cell sheet made of myocardial tissue cells that retain contracting and relaxing functions intercellular electrical coupling and orientation 2 A three dimensional structure of myocardium like cultured cells that retain contracting and relaxing functions as well as three dimensional intercellular electrical coupling and orientation said structure forming tubular cavities of vascular endothelial cells and or having a single epicardium like outer cell layer 3 The myocardium l ike cultured cell sheet according to claim 1 or the three dimensional structure according to claim 2 which have been peeled off from a substrate without being treated with a proteolytic enzyme 4 The myocardium like cultured cell sheet or three dimensional structure according to claim 1 whi
386. tion of culturing in step b is between about 6 weeks and about 8 weeks 7 The method of claim 1 wherein the duration of culturing in step b is between about 3 months and about 5 months 8 The method of claim 1 wherein the cell monolayer of step d contains cells that are Pax6 bestrophin CRALBP PEDF and express RPE65 9 The method of claim 1 wherein the cell monolayer of step d contains cells that are Pax6 bestrophin CRALBP PEDF and express RPE65 and have the absence of at least one ES cell marker selected from the group consisting of Oct4 and Sox2 10 The method of claim 1 wherein prior to step b said multilayer population of hES cells are cultured in the presence of exogenously added FGF and a fibroblast feeder layer 11 The method of claim 1 wherein prior to step b said multilayer population of hES cells are cultured the presence of exogenously added FGF and LIF and a fibroblast feeder layer 12 The method of claim 1 wherein prior to step b said multilayer population of hES cells are cultured the presence of exogenously added FGF 170 PLASMANATE RTM and a fibroblast feeder layer 13 A method for producing an enriched population of human retinal pigment epithelium RPE cells the method comprising a providing a culture of human ES hES cells b culturing the hES cells to produce one or more embryoid bodies c culturing said one or more embryoid bodies for a sufficient time for the appearanc
387. ving heart which have been 41 obtained from the living heart D5 2010 5 20 2010 7 10 2010 12 23 9 1 A process for producing a three dimensional structure of myocardium ike cells having contracting and relaxing functions intercellular electrical coupling and orientation which comprises culturing cells which have been obtained from the living heart on a cell culture support having a substrate surface coated with a temperature responsive polymer whose upper or lower critical solution temperature in water is 0 80 C and subsequently 1 bringing the temperature of the culture solution to above the upper critical solution temperature or below the lower
388. yogenin 12 A structure according to claim 11 wherein an expression level of the non adult heart marker in the structure is at least 50 of an expression level of the non adult heart marker in skeletal myoblasts 13 A structure according to claim 1 wherein the three dimensional structure expresses all of myosin heavy chain IIa myosin heavy chain II b myosin heavy chain Id Ix CD56 MyoD Myf5 and myogenin 14 A structure according to claim 13 wherein an expression level of each of myosin heavy chain IIa myosin heavy chain lib myosin heavy chain IId IIx CD56 MyoD Myf5 and myogenin in the structure is at least about 50 of an expression level thereof in skeletal myoblasts 15 A structure according to claim 13 wherein an expression level of each of myosin heavy chain IIa myosin heavy chain lib myosin heavy chain IId IIx CD56 MyoD Myf5 and myogenin in the structure is at least about 100 of an expression level thereof in skeletal myoblasts 16 A structure according to claim 1 wherein the cell derived from a part other than myocardium is a cell not derived from heart 17 A structure according to claim 1 wherein the applicability to heart includes applicability to myocardium 18 A structure according to claim 1 comprising a monolayer cell sheet 19 A structure according to claim 1 comprising a multilayer cell sheet 20 A structure according to claim 19 wherein the multilayer cell sheet has biological connectio
389. yvinylidene difluoride membrane polyurethane a Spandex mesh and a stockinet like material 5 A three dimensional structure of myocardium like cells having contracting and relaxing functions intercellular electrical coupling and orientation that is produced by the process according to any one of claims 1 4 6 The three dimensional structure of myocardium l ike cells according to claim 5 which is a three dimensional structure of myocardium l ike cells in which tubular cavities are formed of vascular endothelial cells endogenous in said three dimensional structure of myocardium like cel and the vascular endothelial cells in the tissue around the buried graft are allowed to group inward and form tubular cavities thereby forming blood vessels 7 The three dimensional structure of myocardium like cells according to claim 5 which is suitable for use in the treatment of heart disease and other circulatory organ related diseases or digestive organ related diseases 8 Use of cells which have been obtained from the living heart for the manufacture of three dimensional structure of myocardium like cells according to any one of claim 5 6 and 7 for use in the treatment of heart disease and other circulatory organ related diseases or digestive organ related diseases 9 The three dimensional structure of myocardium like cells according to claim 5 for us in a method of treating heart disease and other circulatory organ related diseases or digestive o

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