Gynogenetic or androgenetic production of pluripotent cells and cell lines, and use thereof to produce differentiated cells and tissues

• US 8,273,571 B2
• Filed: 05/27/2011
• Issued: 09/25/2012
• Est. Priority Date: 10/28/1999
• Status: Expired due to Fees

• Current Assignee: University Of Massachusetts
• Original Assignee: Advanced Cell Technology Incorporated (Astellas Pharma Incorporated)
• Inventors: Robl, James M., Cibelli, Jose, Burnside, Amy
• Primary Examiner(s): Noble, Marcia S
• Application Number:
  US13/117,184
• Publication Number:
  US 20110293580A1
• Time in Patent Office: 487 Days
• Field of Search: 435/377, 435/366, 435/373, 435/384, 435/455
• US Class Current: 435/377
• CPC Class Codes: A61K 35/12 Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells vaccines or medicinal preparations containing antigens or antibodies A61K39/00
• CPC Class Codes: A61K 35/545 Embryonic stem cells; Pluripotent stem cells; Induced pluripotent stem cells; Uncharacterised stem cells
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C12N 15/873 Techniques for producing new embryos, e.g. nuclear transfer, manipulation of totipotent cells or production of chimeric embryos
• CPC Class Codes: C12N 2500/14 Calcium; Ca chelators; Calcitonin
• CPC Class Codes: C12N 2501/999 Small molecules not provided for elsewhere
• CPC Class Codes: C12N 2502/13 connective tissue cells; generic mesenchyme cells, e.g. so-called "embryonic fibroblasts"
• CPC Class Codes: C12N 2506/04 from germ cells
• CPC Class Codes: C12N 2510/00 Genetically modified cells
• CPC Class Codes: C12N 2517/10 Conditioning of cells for in vitro fecondation or nuclear transfer
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696
• CPC Class Codes: C12N 5/0696 Artificially induced pluripotent stem cells, e.g. iPS
• CPC Class Codes: G01N 2333/475 Assays involving growth factors
• CPC Class Codes: G01N 33/5008 for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
• CPC Class Codes: G01N 33/502 for testing non-proliferative effects
• CPC Class Codes: G01N 33/5044 involving specific cell types
• CPC Class Codes: G01N 33/5073 Stem cells
• CPC Class Codes: G01N 33/5088 of vertebrates

Abstract

Methods for obtaining pluripotent (embryonic stem) cells from parthenogenetic embryos, especially primates, are provided. These cells are useful for producing differentiated cells, tissues and organs, especially human and non-human primate cells, tissues and organs.

Claims

1. A method for producing primate gynogenetic pluripotent cells comprising:

(a) providing a primate oocyte in metaphase II;

(b) activating said primate oocyte using activation conditions selected from the group consisting of;

contacting the oocyte with a calcium ionophore, contacting the oocyte with a protein synthesis inhibitor, contacting the oocyte with phosphorylation inhibitor, and any combination thereof, to produce an activated oocyte,

(c) culturing said activated oocyte to produce a gynogenetic blastocyst having an inner cell mass;

(d) isolating said inner cell mass or isolated cells therefrom and transferring said inner cell mass or isolated cells therefrom into an in vitro culture that inhibits differentiation of said inner cell mass or isolated cells therefrom; and

(e) culturing said inner cell mass or isolated cells therefrom or cells derived therefrom during inner cell mass proliferation in a culture that maintains the inner cell mass cells or cells therefrom in the undifferentiated state thereby producing a primate gynogenetic pluripotent stem cell.

2. The method of claim 1, wherein said calcium ionophore is A23187 or Ionomycin.

3. The method of claim 1, wherein said protein synthesis inhibitor is cycloheximide or puromycin.

4. The method of claim 1, wherein said phosphorylation inhibitor is 6-DMAP (dimethylaminopurine).

5. The method of claim 1, wherein the primate oocyte is human.

6. The method of claim 1, wherein said method further comprises a step of differentiating in vitro said undifferentiated primate gynogenetic pluripotent cells to produce primate gynogenetic differentiated cells.

7. The method of claim 1, wherein said primate oocyte is genetically modified.

8. The method of claim 3, wherein said protein synthesis inhibitor is puromycin.