Generation of iPSCs from Genetically Corrected Brca2 Hypomorphic Cells: Implications in Cell Reprogramming and Stem Cell Therapy. (February 2014)

• Record Type: Journal Article
• Authors: Navarro, S.; Moleiro, V.; Molina‐Estevez, F.J.; Lozano, M.L.; Chinchon, R.; Almarza, E.; Quintana‐Bustamante, O.; Mostoslavsky, G.; Maetzig, T.; Galla, M.; Heinz, N.; Schiedlmeier, B.; Torres, Y.; Modlich, U.; Samper, E.; Río, P.; Segovia, J.C.; Raya, A.; Güenechea, G.; Izpisua‐Belmonte, J.C.; Bueren, J.A.
• Is Part Of: Stem cells. Volume 32:Number 2(2014:Feb.)
• Page Start: 436
• Page End: 446
• Publication Date: 2014-02
• Subjects: Cloning -- Periodicals; Clone cells -- Periodicals; Stem cells -- Periodicals; Cell Differentiation -- Periodicals; Cell Division -- Periodicals; Clone Cells -- Periodicals; Hematopoietic Stem Cells -- Periodicals; Stem Cells -- Periodicals; 571.84
• DOI: 10.1002/stem.1586
• Languages: English
• ISSNs: 1066-5099

Abstract

<abstract abstract-type="main"> <title>A<sc>bstract</sc></title> <p>Fanconi anemia (FA) is a complex genetic disease associated with a defective DNA repair pathway known as the FA pathway. In contrast to many other FA proteins, BRCA<sub>2</sub> participates downstream in this pathway and has a critical role in homology‐directed recombination (HDR). In our current studies, we have observed an extremely low reprogramming efficiency in cells with a hypomorphic mutation in <italic>Brca2</italic> (<italic>Brca2<sup>Δ</sup><sup>27/</sup></italic><sup>Δ27</sup>), that was associated with increased apoptosis and defective generation of nuclear RAD51 foci during the reprogramming process. Gene complementation facilitated the generation of <italic>Brca2<sup>Δ</sup><sup>27/</sup></italic><sup>Δ27</sup> induced pluripotent stem cells (iPSCs) with a disease‐free FA phenotype. Karyotype analyses and comparative genome hybridization arrays of complemented <italic>Brca2<sup>Δ</sup><sup>27/</sup></italic><sup>Δ27</sup> iPSCs showed, however, the presence of different genetic alterations in these cells, most of which were not evident in their parental <italic>Brca2<sup>Δ</sup><sup>27/</sup></italic><sup>Δ27</sup> mouse embryonic fibroblasts. Gene‐corrected <italic>Brca2<sup>Δ</sup><sup>27/</sup></italic><sup>Δ27</sup> iPSCs could be differentiated in vitro toward the hematopoietic lineage, although with a more limited efficacy than WT iPSCs or mouse embryonic stem cells, and did not engraft in irradiated <italic>Brca2<sup>Δ</sup><sup>27/</sup></italic><sup>Δ27</sup> recipients. Our results are consistent with previous studies proposing that HDR is critical for cell reprogramming and demonstrate that reprogramming defects characteristic of <italic>Brca2</italic> mutant cells can be efficiently overcome by gene complementation. Finally, based on analysis of the phenotype, genetic stability, and hematopoietic differentiation potential of gene‐corrected <italic>Brca2<sup>Δ</sup><sup>27/</sup><sup>Δ</sup><sup>27</sup></italic> iPSCs, achievements and limitations in the application of current reprogramming approaches in hematopoietic stem cell therapy are also discussed. S<sc>tem</sc> C<sc>ells</sc><italic>2014;32:436–446</italic></p> </abstract>