X-linked Severe Combined Immunodeficiency (SCID-X1) is an immunopathy caused by a mutation of the common gamma chain (γc) gene, IL2RG, which results in a lack of T cells, NK cells and with dysfunctional B cells. Current gene therapy methods involve the addition of a correct γc gene via integrating viral vectors. However, these current non-targeting gene addition strategies can result in transformation of the cell. A novel solution to this problem is met by targeted gene correction via homologous recombination stimulated by a site specific cleavage event caused by zinc finger nucleases (ZFN) within the disease gene. A γc deficient mouse has been created by replacing the murine il2rg locus with a mutated human IL2RG containing a point mutation frequently seen in SCID-X1 patients. The mutant human IL2RG is transcribed and initial analysis of this new SCID-X1 model has revealed a phenotype mirroring γc gene knockout mice. Lineage negative bone marrow cells from these mice, transduced with integrating lentiviral vector encoding functional IL2RG can reconstitute the immune cells in the Rag2-/-γc-/- double knockout SCID mouse model. Therefore the humanised mouse model of SCID-X1 can be corrected and is an appropriate platform to assess the efficiency of various gene targeting and correction strategies for the human mutation including ZFN induced homologous recombination. We have successfully achieved targeted homologous recombination in both a human T cell SCID-X1 cell line model and the humanised mouse embryonic stem cells with IL2RG specific ZFN.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:626335 |
| Date | January 2013 |
| Creators | Pallant, C. E. |
| Contributors | Thrasher, A. |
| Publisher | University College London (University of London) |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://discovery.ucl.ac.uk/1403328/ |
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