ＳＴＡＰ細胞の国際特許出願

【拡散希望】STAP細胞についてひどく誤った記事が拡散されているという話を聞いたので書いておきます。その記事はSTAP細胞については特許が出願されていないので、小保方さんを潰して他の人が特許を出願するための、巨大な利権が絡んだ陰謀だ、という趣旨のようです。しかし、STAP細胞の特許は既に出願され、公開されています。国際公開の番号はWO2013163296です。

最初の出願は２０１２年のようです。

ハーバード大学のバカンティ教授が筆頭発明者ですが、早稲田、理研の研究者、小保方さんも出願人に入っています。ですから、この特許がある以上、ノウハウを公開して実用化が進み、皆が使えば使うほど儲かります。特許権者である小保方さんにも当然持ち分の収入は得られるでしょう。

そういう意味で、ノウハウを隠す意味は何もありません。また、ノウハウを隠すのは、科学者としての姿勢ではありません。少なくとも、直接聞かれたら教えるべきです。でなければ論文を取り下げる必要があります。

それをノートを公開したら小保方さんのノウハウが丸裸になるから利権を守るために彼女や理研が隠している、とかいうデマがあります。小保方さんはSTAP細胞は作れても、STAP幹細胞は作れないと思います。それができるのは山梨大学の若山先生ですが、その部分を担当した若山先生がＳＴＡＰ細胞に疑いを持っているのですから、論文取り下げが妥当でしょう。そういう意味で彼女のノウハウを全部公開しても実用化はできないので隠す意味はありません。

巨大な利権とかいう話があるとすれば、ハーバード大学側の問題ですが、既に特許が出願され、バカンティ教授もプロトコルを公開していますから利権を活用するには、実用化するのが早道で、そのためには、ノウハウを公表することでしょう。なので、再生医療で数百兆円の利権のために小保方さんが潰された、とかいうのは全くデマですね。

しかも、ヒトでできていないのは確実ですし、マウスでも追試した６～８位の研究機関でどこも作れない以上、利権どころではなく、その技術の再現性（完成度）そのものが疑われています。間違っているけど面白い記事が拡散されるのがネットの問題ですね。

個人のブログとはいえ、少しは調べてから書いて欲しいです。STAP細胞というのは、細い管を通したり、酸に浸す等してストレスをかけて、細胞の容積を小さくしたり細胞内小器官を取り除いたりすることで、未分化細胞の指標である、Oct4またはnanog遺伝子を発現するようになった細胞です。刺激を与えたら遺伝子発現が乱れたり、ストレス応答や細胞死等の方向に変化し、Oct4またはnanog遺伝子のスイッチが入って光っても何も不思議ではありません。

特に、生後６日目のマウスなら分化全能性を持つ細胞がより多く混入していてもおかしくありませんからよりOct4遺伝子を発現している細胞が多い可能性があります。ＳＴＡＰ細胞はそうした微量に含まれる分化全能性を持つ細胞を拾っただけ、という説があり、小保方さんもその可能性は認めています。つまりもともと含まれていた微量の未分化細胞を拾っただけで、体細胞が刺激により変化したのではない可能性もあります。もしそうならセルソーターを使えば簡単に取り出せるでしょう。

小保方さんが２００回見た、というのは、このOct4遺伝子が発現している細胞です。そこから何にでも再分化できる細胞にするには、STAP幹細胞という増殖能を持った細胞にする必要があります。この過程は小保方さんは苦手で、山梨大の若山先生がやっていたそうです。STAP細胞→STAP幹細胞→様々な細胞に分化まで行って初めて成功と言えます。小保方さんが確認したのはこの最初のSTAP細胞までです。

ＳＴＡＰ細胞特許ですが、国際公開されていました。チャールズ・バカンティ教授が筆頭出願人です。

WO2013163296

GENERATING PLURIPOTENT CELLS DE NOVO

発明者
VACANTI CHARLES A [US]; VACANTI MARTIN P [US]; KOJIMA KOJI [US]; OBOKATA HARUKO [JP]; WAKAYAMA TERUHIKO [JP]; SASAI YOSHIKI [JP]; YAMATO MASAYUKI [JP] +

請求項は以下です。こんな請求項は見たことがないですね。構成がほとんど書かれていないし、おそらく実施可能要件も満たさないでしょう。

What is claimed herein is:
1. A method to generate a pluripotent cell, comprising subjecting a cell to a stress.
2. The method according to claim 1, wherein the pluripotent cell is generated without introduction of an exogenous gene, a transcript, a protein, a nuclear component or cytoplasm, or without cell fusion.
3. The method of any of claims 1-2, further comprising selecting a cell exhibiting
pluripotency.
4. The method of any of claims 1-3, wherein the cell is not present as part of a tissue.
5. The method of any of claims 1-4, wherein the cell is a somatic cell, a stem cell, a progenitor cell or an embryonic cell.
6. The method of any of claims 1-5, wherein the cell is an isolated cell.
7. The method of any of claims 1-6, wherein the cell is present in a heterogeneous
population of cells.
8. The method of any of claims 1-7, wherein the cell is present in a homogenous
population of cells.
9. The method of any of claims 1-8, wherein selecting the cell exhibiting pluripotency comprises selecting a cell expressing a stem cell marker.
10. The method of any of claim 9, wherein the stem cell marker is selected from the group consisting of :
Oct4; Nanog; E-cadherin, and SSEA4.
11. The method of any of claims 1-10, wherein selecting the cell exhibiting pluripotency comprises selecting a cell which is not adherent.
12. The method of any of claims 1-11, wherein the stress comprises unphysiological stress in tissue or cell culture.
13. The method of any of claims 1-12, wherein the stress comprises exposure of the cell to at least one environmental stimulus selected from: trauma, mechanical stimuli, chemical exposure, ultrasonic stimulation, oxygen-deprivation, radiation, exposure to extreme temperatures, dissociation, trituration, physical stress, hyperosmosis, hypoosmosis, membrane damage, toxin, extreme ion concentration, active oxygen, UV exposure, strong visible light, deprivation of essential nutrition, or unphysiolosically acidic environment.
14. The method of any of claims 1-13, wherein the stress comprises exposing the cell to a pH of from about 3.0 to about 6.8.
15. The method of any of claims 1-4, wherein the stress comprises exposing the cell to a pH of from about 4.5 to about 6.0.
16. The method of claim 15, wherein the stress comprises exposing the cell to a pH of from about 5.4 to about 5.8.
17. The method of any of claims 12-16, wherein the cell is exposed for 2-3 days.
18. The method of any of claims 12-17, wherein the cell is exposed for 1 day or less.
19. The method of any of claims 12-18, wherein the cell is exposed for 1 hour or less.
20. The method of any of claims 12-19, wherein the cell is exposed for about 30 minutes.
21. The method of claim 13, wherein the exposure to extreme temperatures comprises exposing the cell to temperatures below 35 [deg.]C or above 42[deg.]C.
22. The method of claim 21, wherein the exposure to extreme temperatures comprises exposing the cell to temperatures at, or below freezing or exposure of the cell to temperatures at least about 85[deg.]C.
23. The method of claim 13, wherein the mechanical stimulus comprises exposing the cell to shear stress or/and high pressure.
24. The method of claim 23, wherein the mechanical stimulus comprises passing the cell through at least one device with a smaller aperture than the size of the cell.
25. The method of claim 23, wherein the mechanical stimulus comprises passing the cell through several devices having progressively smaller apertures.
26. The method of any of claims 1-25, further comprising culturing the pluripotent cell to allow propagation of the pluripotent cell.
27. The method of any of claims 1-26, wherein the pluripotent cell expresses a stem cell marker.
28. The method of claim 27, wherein the stem cell marker is selected from the group consisting of:
Oct4; Nanog; E-cadherin, and SSEA4.
29. The method of any of claims 1-28, wherein the cell is a mammalian cell.
30. The method of any of claims 1-29, wherein the cell is a human cell.
31. The method of any of claims 1-30, wherein the cell is an adult cell, a neonatal cell, a fetal cell, amniotic cell, or cord blood cell.
32. The method of any of claims 1-31, further comprising maintaining the pluripotent cell in vitro.
33. The method of any of claims 1-32, wherein the epigenetic state of the cell is altered to more closely resemble the epigenetic state of an embryonic stem cell.
34. The method of claim 33, wherein the epigenetic state comprises methylation patterns.
35. The method of any of claims 1-34, wherein the stress comprises removing at least about 40% of the cytoplasm from the cell.
36. The method of claim 35, wherein at least about 50% of the cytoplasm is removed from the cell.
37. The method of claim 36, wherein at least about 60% of the cytoplasm is removed from the cell.
38. The method of claim 37, wherein between 60-80% of the cytoplasm is removed from the cell.
39. The method of claim 37, wherein at least about 80% of the cytoplasm is removed from the cell.
40. The method of claim 39, wherein at least about 90% of the cytoplasm is removed from the cell.
41. The method of any of claims 1-40, wherein the stress comprises removing at least about 40% of the mitochondria from the cell.
42. The method of claim 41, wherein the removal of a portion of the cytoplasm removes at least about 50% of the mitochondria from the cytoplasm.
43. The method of claim 42, wherein the removal of cytoplasm or mitochondria removes about 50%-90% of the mitochondria from the cytoplasm.
44. The method of claim 42, wherein the removal of cytoplasm or mitochondria removes more than 90% of the mitochondria from the cytoplasm.
45. The method of any of claims 1-44, wherein the stress is sufficient to disrupt the cellular membrane of at least 10% of cells exposed to the stress.
46. An assay comprising;
contacting a pluripotent cell produced by the method according to any of claims 1 to 45 with a candidate agent.
47. The assay of claim 46, for use to identify agents which affect one or more of the
viability, differentiation, proliferation of the pluripotent cell.
48. Use of a pluripotent cell produced by the method according to any one of claims 1 to 45 in a method of cell therapy for a subject.
49. A method of preparing a cell or tissue that is compatible with cell therapy to be
administered to a subject, comprising:
generating a pluripotent cell from a cell according to any one of claims 1 to 45; wherein the cell is an autologous cell or HLA-matched allogeneic cell.
50. The method of claim 49, further comprising differentiating the pluripotent cell along a pre-defined cell lineage prior to administering the cell or tissue to the subject.
51. A composition comprising a pluripotent cell, wherein the pluripotent cell is generated from a cell by the methods according any of claims 1 to 45.
52. A method of producing a pluripotent stem cell, the method comprising culturing a cell in the presence of adrenocorticotropic hormone (ACTH), 2i or 3i medium
53. The method of claim 52, wherein the cell is cultured in LIF medium comprising ACTH.
54. The method of claim 52 or 53, wherein the ACTH is present at a concentration of from about 0.1 [mu][Mu] to about 100 [mu][Mu] .
55. The method of any of claims 52-54, wherein the cell is a cell generated by the method of any of claims 1-45.
56. The method of any of claims 52-55, wherein the cell is a totipotent cell.
57. The method of any of claims 52-56, wherein the cell is cultured in the presence of ACTH, 2i or 3i medium for at least 3 days.
58. The method of any of claims 52-57, wherein the cell is cultured in the presence of ACTH, 2i or 3i medium for at least 5 days.
59. The method of any of claims 52-58, wherein the cell is cultured in the presence of ACTH, 21 or 3i medium for at least 7 days.
60. The method of any of claims 52-59, wherein after the culturing step, the cell expresses detectable level of a stem cell marker selected from the group consisting of:
Oct3/4; Nanog; Rexl; Klf4; Sox2; Kl[pound]2; Esrr-beta; Tbx3; and Klf5.
61. A method of increasing the self-renewal ability of a pluripotent cell, the method
comprising culturing the cell in the presence of adrenocorticotropic hormone (ACTH), 2i or 3i medium.
62. The method of claim61, wherein the cell is cultured in LIF medium comprising ACTH.
63. The method of any of claims 61-62, wherein the ACTH is present at a concentration of from about 0.1 [mu][Mu] to about 100 [mu][Mu] .
64. The method of any of claims 61-63, wherein the cell is a cell generated by the method of any of claims 1-45.
65. The method of any of claims 61-64, wherein the cell is a totipotent cell.
66. The method of any of claims 61-65, wherein the cell is cultured in the presence of ACTH, 2i or 3i medium for at least 3 days.
67. The method of any of claims 61-66, wherein the cell is cultured in the presence of ACTH, 2i or 3i medium for at least 5 days.
68. The method of any of claims 61-67, wherein the cell is cultured in the presence of ACTH, 2i or 3i medium for at least 7 days.
69. The method of any of claims 61-68, wherein after the culturing step, the cell expresses detectable level of a stem cell marker selected from the group consisting of: Oct3/4; Nanog; Rexl; Klf4; Sox2; Kl[pound]2; Esrr-beta; Tbx3; and Klf5.
70. A method of autologous cell therapy in a subject in need of cell therapy, comprising a. generating a pluripotent cell from a cell according to any one of claims 1 to 45, wherein the cell is obtained from the subject, and
b. administering a composition comprising the pluripotent cell or a differentiated progeny thereof to the subject.
71. The method of claim 70, further comprising differentiating the pluripotent cell along a pre-defined cell lineage prior to administering the composition to the subject.
72. A method of producing a pluripotent cell capable of differentiating into a placental cell, the method comprising culturing the pluripotent cell generated by the method of any of claims 1-45 in the presence of FGF4.
73. The method of claim 72, wherein the concentration of FGF4 is 1 nM to 1 uM.
74. The method of claim 72 or 73, wherein the pluripotent cell is capable of differentiating into an embryonic stem cell.