US2010075421A1PendingUtilityA1

Efficient method for nuclear reprogramming

58
Assignee: UNIV KYOTOPriority: Dec 10, 2007Filed: Nov 25, 2008Published: Mar 25, 2010
Est. expiryDec 10, 2027(~1.4 yrs left)· nominal 20-yr term from priority
Inventors:Shinya YamanakaMichiyo Koyanagi
C12N 5/0607C12N 5/0696C12N 15/113C12N 2501/606C12N 2501/65C12N 2501/604C12N 2510/00C12N 2310/141C12N 2501/602C12N 2501/605C12N 2501/603C12N 5/10C12N 2501/608C12N 2330/10
58
PatentIndex Score
0
Cited by
0
References
0
Claims

Abstract

A method of preparing induced pluripotent stem cells, comprising a nuclear reprogramming step with a nuclear reprogramming factor in the presence of miRNA, wherein said miRNA has a property of providing a higher nuclear reprogramming efficiency in the presence of said miRNA than in the absence thereof.

Claims

exact text as granted — not AI-modified
1 . A method of preparing induced pluripotent stem cells, comprising nuclear reprogramming at least one somatic cell with nuclear reprogramming factor and at least one miRNA, wherein the at least one miRNA increases efficiency of the nuclear reprogramming of the at least one somatic cell compared to nuclear reprogramming of the at least one somatic cell with the nuclear reprogramming factor in the absence of the at least one miRNA. 
     
     
         2 . The method according to  claim 1 , wherein the at least one miRNA is expressed in embryonic stem cells at a higher level than in somatic cells. 
     
     
         3 . The method according to  claim 1 , wherein a gene encoding the nuclear reprogramming factor and/or the at least one miRNA is introduced into the at least one somatic cell. 
     
     
         4 . The method according to  claim 3 , wherein a vector comprising the gene and/or a vector encoding the at least one miRNA is introduced into the at least one somatic cell. 
     
     
         5 . The method according to  claim 4 , wherein the vector comprising the gene or encoding the at least one miRNA is a retroviral vector. 
     
     
         6 . The method according to  claim 3 , wherein the gene is selected from an Oct family gene, a Klf family gene, and a Sox family gene. 
     
     
         7 . The method according to  claim 3 , wherein the gene is selected from Oct3/4, Klf4, and Sox2. 
     
     
         8 . The method according to  claim 3 , wherein the nuclear reprogramming factor comprises Oct3/4, Klf4, and Sox2. 
     
     
         9 . The method according to  claim 3 , wherein the at least one miRNA is introduced into the at least one somatic cell as primary miRNA. 
     
     
         10 . The method according to  claim 3 , wherein the at least one miRNA is introduced into the at least one somatic cell as pre-miRNA. 
     
     
         11 . The method according to  claim 1 , wherein the at least one miRNA comprises at least one miRNA represented by SEQ ID NOs: 1 to 14. 
     
     
         12 . The method according to  claim 1 , wherein the at least one miRNA comprises at least one miRNA contained in miRNA cluster hsa-miR-302-367 cluster, hsa-miR-371-373 cluster and hsa-miR-520c. 
     
     
         13 . The method according to  claim 1 , wherein the at least one miRNA regulates DNA methylation. 
     
     
         14 . The method according to  claim 11 , wherein the at least one miRNA regulates de novo DNA methylation. 
     
     
         15 . The method according to  claim 11 , wherein the at least one miRNA down-regulates DNA methylation. 
     
     
         16 . The method according to  claim 1 , wherein the at least one miRNA comprises at least 10 contiguous nucleotides of at least one miRNA represented by SEQ ID NOs: 1 to 14. 
     
     
         17 . The method according to  claim 1 , wherein the at least one miRNA comprises at least 30 contiguous nucleotides of at least one miRNA represented by SEQ ID NOs: 1 to 14. 
     
     
         18 . The method according to  claim 1 , wherein the at least one miRNA comprises at least 60 contiguous nucleotides of at least one miRNA represented by SEQ ID NOs: 1 to 14. 
     
     
         19 . The method according to  claim 1 , wherein the nuclear reprogramming factor does not include c-Myc and/or Sox2. 
     
     
         20 . The method according to  claim 1 , wherein the at least one miRNA comprises hsa-miR-302-367 cluster, hsa-miR-371-373 cluster and hsa-miR-520C miRNA. 
     
     
         21 . The method according to  claim 20 , wherein the nuclear reprogramming factor comprises an Oct family gene member, a Sox family gene member, and a Klf family gene member. 
     
     
         22 . The method according to  claim 1 , wherein the at least one miRNA comprises mmu-miR-295/295* and 294/294*. 
     
     
         23 . The method according to  claim 1 , wherein the at least one miRNA comprises hsa-miR-302-367 cluster and hsa-miR-371-373 cluster miRNA. 
     
     
         24 . The method according to  claim 1 , wherein the nuclear reprogramming factor comprises a Klf family gene, and an Oct family gene. 
     
     
         25 . The method according to  claim 24 , wherein the nuclear reprogramming factor further comprises a Myc family gene. 
     
     
         26 . The method according to  claim 24 , wherein the nuclear reprogramming factor further comprises a Sox family gene. 
     
     
         27 . The method according to  claim 25 , wherein the nuclear reprogramming factor further comprises a Sox family gene. 
     
     
         28 . The method according to  claim 24 , wherein the nuclear reprogramming factor comprises KLF4 and OCT3/4. 
     
     
         29 . A method of increasing the efficiency of nuclear reprogramming comprising: adding a nuclear reprogramming factor and at least one miRNA to at least one somatic cell so that the number of induced pluripotent stem cells produced is greater than in the absence of the added miRNA. 
     
     
         30 . An induced pluripotent stem cell induced by reprogramming a somatic cell, wherein the reprogramming is performed by adding at least one miRNA and in the absence of eggs, embryos, or embryonic stem (ES) cells. 
     
     
         31 . The induced pluripotent stem cell according to  claim 30 , wherein the induced pluripotent stem cell is a human cell. 
     
     
         32 . An induced pluripotent stem cell obtained by the method of  claim 1 . 
     
     
         33 . An induced pluripotent stem cell obtained by the method of  claim 29 . 
     
     
         34 . A somatic cell derived by inducing differentiation of the pluripotent stem cell according to  claim 32 . 
     
     
         35 . A somatic cell derived by inducing differentiation of the pluripotent stem cell of  claim 33 . 
     
     
         36 . The method according to  claim 25 , wherein the nuclear reprogramming factor excludes a Sox family gene. 
     
     
         37 . The method according to  claim 26 , wherein the nuclear reprogramming factor excludes a Myc family gene. 
     
     
         38 . The method according to  claim 1 , where the at least one somatic cell comprises a plurality of somatic cells.

Join the waitlist — get patent alerts

Track US2010075421A1 — get alerts on status changes and closely related new filings.

We store only your email — no account needed. See our privacy policy.