Methods and products for transfection
The present invention relates in part to methods for producing tissue-specific cells from patient samples, and to tissue-specific cells produced using these methods. Methods for reprogramming cells using RNA are disclosed. Therapeutics comprising cells produced using these methods are also disclosed.
1. A method for reprogramming a differentiated cell to a less differentiated state, comprising:
(a) providing a differentiated cell;
(b) culturing the differentiated cell; and
(c) transfecting the differentiated cell with one or more synthetic RNA molecules,
wherein the one or more synthetic RNA molecules include at least one RNA molecule encoding one or more reprogramming factors selected from the group consisting of Oct4 protein, Sox2 protein, Klf4 protein, c-Myc protein, 1-Myc protein, Tert protein, Nanog protein, and Lin28 protein;
wherein the transfecting results in the cell expressing the one or more reprogramming factors to result in the cell being reprogrammed to a less differentiated state; and
wherein step (c) is performed not in the presence of irradiated fibroblast feeder cells and occurs in the presence of a medium containing ingredients that support reprogramming of the differentiated cell to a less differentiated state.
2. The method of claim 1 , wherein the differentiated cell is derived from a biopsy.
3. The method of claim 2 , wherein the differentiated cell is derived from a dermal punch biopsy sample.
4. The method of claim 1 , wherein the differentiated cell is from a human subject.
5. The method of claim 1 , wherein the differentiated cell is a skin cell.
6. The method of claim 1 , further comprising contacting the cell with at least one member of the group: poly-L-lysine, poly-L-ornithine, RGD peptide, fibronectin, vitronectin, collagen, and laminin.
7. The method of claim 1 , wherein the one or more synthetic RNA molecules contain at least one of a pseudouridine or a 5-methylcytidine residue.
8. The method of claim 1 , wherein the medium is substantially free of immunosuppressants.
9. A method for reprogramming a non-pluripotent cell, comprising:
(a) providing a non-pluripotent cell;
(b) culturing the non-pluripotent cell; and
(c) transfecting the non-pluripotent cell with one or more synthetic RNA molecules,
wherein the one or more synthetic RNA molecules include at least one RNA molecule encoding one or more reprogramming factors selected from the group consisting of Oct4 protein, Sox2 protein, Klf4 protein, c-Myc protein, 1-Myc protein, Tert protein, Nanog protein, and Lin28 protein;
wherein the transfecting results in the cell expressing the one or more reprogramming factors to result in the cell being reprogrammed; and
wherein step (c) is performed not in the presence of irradiated fibroblast feeder cells and occurs in the presence of a medium containing ingredients that support reprogramming of the non-pluripotent cell.
10. The method of claim 9 , wherein the non-pluripotent cell is derived from a biopsy.
11. The method of claim 10 , wherein the non-pluripotent cell is derived from a dermal punch biopsy sample.
12. The method of claim 9 , wherein the non-pluripotent cell is from a human subject.
13. The method of claim 9 , wherein the non-pluripotent cell is a skin cell.
14. The method of claim 9 , further comprising contacting the cell with at least one member of the group: poly-L-lysine, poly-L-ornithine, RGD peptide, fibronectin, vitronectin, collagen, and laminin.
15. The method of claim 9 , wherein the one or more synthetic RNA molecules contain at least one of a pseudouridine or a 5-methylcytidine residue.
16. The method of claim 9 , wherein the medium is substantially free of immunosuppressants.