Transcription factors can exert a dominant effect in specifying gene expression program and impart unique cellular property. Important regulators of pluripotency, Oct4, Sox2 and Nanog, are transcription factors. Of these three proteins, Oct4 and Nanog have roles that are specific to pluripotent cells. Pluripotency is the ability of a cell to give rise to a cell of any desired tissue of an organism. Further transcription factors such as Stat3, P53 and others are thought to play a role in a regulatory network that controls pluripotency.
The multitude of cells such as cells of the human or animal body is generated through the process of differentiation. It was previously surmised that as stem cells differentiate, they lose their ability to make cell fate decision and become more restricted in their potential. However, the developmentally restricted state of differentiated somatic cells can also be reversed to a pluripotent state through several strategies of reprogramming (for an introduction see Jaenisch, R., & Young, R., Cell (2008) 132, 567-582; cf. also FIG. 1). Among these methods are somatic cell nuclear transfer using enucleated unfertilized oocyte, fusion of differentiated cells with pluripotent cells and treatment of differentiated cells using extracts derived from pluripotent cells (Lewitzky, M. & Yamanaka, S., Curr. Opin. Biotechnol. (2007) 18, 467-473). The somatic cell nuclear transfer has in the meantime also been applied to a fertilized embryo as a recipient (WO 2008103462).
Reprogramming of somatic cells to pluripotent cells can also be achieved by the retrovirus mediated transduction of defined transcription factors. Conversion of murine and human fibroblasts to pluripotent cells known as induced pluripotent stem (iPS) cells can be achieved using the four transcription factors Oct4, Sox2, c-Myc and Klf4 (see e.g. Takahashi, K. & Yamanaka, S., Cell (2006) 126, 663-676; Lowry, W. E., et al., Proc. Natl. Acad. Sci. (2008) 105, 8, 2883-2888). The obtained iPS cells are developmentally and epigenetically indistinguishable from embryonic stem (ES) cells (ibid.) and have expression profiles that are highly similar to wild-type embryonic stem cells (Mikkelsen, T. S., et al., Nature (2008) 454, 49-55, corrigendum in Nature (2008) 454, 794-794). By overexpressing these transcription factors, differentiated fibroblasts from human ES cells, primary fetal tissues, neonatal skin fibroblasts, adult fibroblasts and adult mesenchymal stem cells can be reprogrammed to iPS cells (Park, I-H, et al., Nature (2007) 451, 141-147). Successful reprogramming of fibroblasts to iPS cells requires the heterologous expression of these four transcription factors for at least 14 days (Brambrink, T., et al., Cell Stem Cell (2008) 2, 151-159).
Mature, fully differentiated B lymphocytes, pancreatic β cells, hepatocytes, keratinocytes and gastric epithelial cells can also be reprogrammed into iPS cells by expressing heterologous Oct4, Sox2, c-Myc and Klf4 using inducible lentiviral vectors or pMXs-based retroviruses (Takahashi, K., et al., Cell (2007) 131, 861-872; Hanna, J.; et al., Cell (2008) 133, 250-264; Stadtfeld, M., et al., Current Biology (2008) 18, 12, 890-894; Maherali, N., et al., Cell Stem Cell (2008) 3, 340-345; for an overview see e.g. Welstead, G. G., et al., Current Opinion in Genetics & Development (2008) 18, doi:10.1016/j.gde.2008.01.013, or Durcova-Hills, G., et al., Differentiation (2008) 76, 323-325; Aoi, T., et al., Science (2008) 321, 699-702). Mesenchymal cells and myeloid cells from human ES cells, as well as primary fibroblasts and newborn foreskin fibroblasts can also be reprogrammed into iPS cells by expressing heterologous Oct4, Sox2, Nanog and Lin28 using a lentiviral vector (Yu, J., et al., Science (2007) 318, 191-920). After the priority date of the present application even adult neural stem cells have been reprogrammed to induced pluripotent stem cells with heterologous Oct4 and either Klf4 or c-Myc (Kim, J. B., et al., Nature (2008) 454, 646-650).
The ability of embryonic stem cells to readily differentiate furthermore continues to pose a major practical challenge. In order to maintain embryonic stem cells in a pluripotent state, their differentiating during handling and growing in culture has to be prevented. For this reason they are traditionally cultured in the presence of fetal calf serum on a layer of feeder cells (see e.g. U.S. Pat. No. 5,843,780 and U.S. Pat. No. 6,090,622) or in fibroblast-conditioned medium (CM). Nevertheless, even under carefully controlled conditions embryonic stem cells may undergo spontaneous differentiation during in-vitro propagation. Leukaemia inhibitory factor, a factor mediating self-renewal in mouse embryonic stem cells, has also been found to inhibit differentiation of mouse embryonic stem cells, but it does not replace the role of feeder cells in preventing differentiation of human embryonic stem cells. Therefore, means of maintaining pluripotency and/or self-renewing characteristics of embryonic stem cells would be a substantial achievement towards realizing the full commercial potential of stem cell therapy.
It is an object of the present invention to offer an alternative method of reprogramming somatic cells and of maintaining pluripotency of an undifferentiated cell.
This object is solved by increasing the amount or the activity of an Err protein, or a functional fragment thereof.