Methods and products for transfection

Patent metadata

Field	Value
Publication number	US-10443045-B2
Application number	US-201916374482-A
Country	US
Kind code	B2
Filing date	Apr 3, 2019
Priority date	Dec 5, 2011
Publication date	Oct 15, 2019
Grant date	Oct 15, 2019

Abstract

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.

First claim

What is claimed is:

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;
wherein the transfecting results in the cell expressing the one or more reprogramming factors; and
wherein step (c) is performed at least twice and the amount of one or more synthetic RNA molecules transfected in one or more later transfections is greater than the amount transfected in one or more earlier transfections to result in the cell being reprogrammed to a less differentiated state 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;
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 at least twice and the amount of one or more synthetic RNA molecules transfected in one or more later transfections is greater than the amount transfected in one or more earlier transfections to result in the non-pluripotent cell being reprogrammed 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.

Assignees

• Factor Bioscience Inc

Inventors

• Angel Matthew
• Rohde Christopher

Classifications

• A61P37/04

  Immunostimulants · CPC title

• A61P9/10

  for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis · CPC title

• A61P7/06

  Antianaemics · CPC title

• A61P3/10

  for hyperglycaemia, e.g. antidiabetics · CPC title

• A61P35/00

  Antineoplastic agents · CPC title