Increased Reprogramming for Generating Pluripotent Stem Cells

Inventors: Teruhisa Kawamura, Jotaro Suzuki, Geoffrey M. Wahl & Juan Carlos Izpisua Belmonte
Potential Uses: Drug Discovery, Gene Expression, Research Tool, Stem Cells

Induced pluripotent stem (iPS) cell generation using two factors and p53 inactivation

The p53 pathway limits reprogramming efficiency. Reprogramming factors/stimuli produce a "reprogramming stress" in somatic cells that activates the p53 pathway. This stress is caused by proto-oncogene over-expression. Eliminating or inhibiting p53 significantly increases reprogramming.

Reprogramming somatic cells to induced pluripotent stem (iPS) cells has been accomplished by expressing pluripotency factors and oncogenes, but 1) the low frequency and 2) the tendency to induce malignant transformation compromise the clinical utility of this powerful approach.

We have developed a method that increases the frequency of reprogramming and reduces the possibility of malignancy in the transformed cells. To do this, we reduce signaling to p53 (either by expressing a mutated version of one of its negative regulators, or by deleting or knocking down p53 or its target gene, p21) or we antagonize reprogramming-induced apoptosis. Quite notably, decreasing p53 protein levels enabled fibroblasts to give rise to iPS cells capable of generating germline-transmitting chimaeric mice using only Oct4 and Sox2. Furthermore, silencing of p53 significantly increased the reprogramming efficiency of human somatic cells. These results provide insights into reprogramming mechanisms and suggest new routes to more efficient reprogramming while minimizing the use of oncogenes.