Zinc finger nuclease gene repair as a treatment for cystinosis

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dc.contributor.advisor Harrison, Patrick en
dc.contributor.advisor Scallan, Martina en
dc.contributor.author Kaschig, Katrin
dc.date.accessioned 2014-01-27T15:46:26Z
dc.date.issued 2013
dc.date.submitted 2013
dc.identifier.citation Kaschig, K. 2013. Zinc finger nuclease gene repair as a treatment for cystinosis. PhD Thesis, University College Cork. en
dc.identifier.uri http://hdl.handle.net/10468/1337
dc.description.abstract Cystinosis is a multi-system autosomal recessive disorder caused by mutations and/or deletions in both alleles of CTNS, a gene encoding for the low pH dependent lysosomal cystine exporter cystinosin. Cystinosis occurs in approximately 1:200,000 newborns worldwide and is characterised by an accumulation of cystine in the lysosomes. The most severe form of the disorder is nephropathic cystinosis presenting Fanconi syndrome and leads without treatment to an end-stage renal failure before the age of ten. The only treatment available so far is cysteamine therapy, which delays disease progression by five years, but does not provide a cure for cystinosis patients. Current gene and cell based therapeutic approaches have not yet provided a suitable alternative. A potentially approach for a long-term treatment could be to generate autologous gene–modified stem cells by repairing the gene. Zinc Finger Nucleases (ZFNs) serve as a tool to increase HDR up to a 200,000-fold by introducing a double-stranded break (DSB). Thus, simple mutations in the CTNS gene could be corrected by introduction of a double-stranded break using ZFNs to boost the process of HDR with a suitable donor DNA sequence. A permanent repair of the most common lesion CTNS, a 57 kb deletion, could be achieved by ZFN-mediated HDR using a minigene CTNS promoter/cDNA construct. The thesis describes the design and testing of seven zinc finger nuclease pairs for their cleavage activity in vitro and in cellulo.. A highly sensitive assay to detect even low levels of ZFN-mediated HDR was also developed. Finally, to further investigate the role of autophagy in tissue injury in cystinotic cells an assay to monitor autophagy levels in the cells was successfully developed. This assay provides the opportunity to demonstrate functional restoration of CTNS after successful ZFN-HDR in cystinotic cells. en
dc.format.mimetype application/pdf en
dc.language.iso en en
dc.publisher University College Cork en
dc.rights © 2013, Katrin Kaschig. en
dc.rights.uri http://creativecommons.org/licenses/by-nc-nd/3.0/ en
dc.subject Zinc finger nucleases en
dc.subject Homology directed repair en
dc.subject.lcsh Cystinosis en
dc.title Zinc finger nuclease gene repair as a treatment for cystinosis en
dc.type Doctoral thesis en
dc.type.qualificationlevel Doctoral en
dc.type.qualificationname PhD (Science) en
dc.internal.availability Full text not available en
dc.check.info Indefinite en
dc.check.date 10000-01-01
dc.description.version Accepted Version
dc.contributor.funder Cystinosis Foundation Ireland en
dc.contributor.funder Health Research Board en
dc.description.status Not peer reviewed en
dc.internal.school Biosciences Institute en
dc.check.reason This thesis is due for publication or the author is actively seeking to publish this material en
dc.check.opt-out Yes en
dc.thesis.opt-out true
dc.check.entireThesis Entire Thesis Restricted
dc.check.embargoformat E-thesis on CORA only en
dc.internal.conferring Autumn Conferring 2013 en


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© 2013, Katrin Kaschig. Except where otherwise noted, this item's license is described as © 2013, Katrin Kaschig.
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