Enhancing the Efficiency of CRISPR/Cas9 Precise Gene Editing for Cystic Fibrosis Gene Therapy
Year: 2018
Type: dissertation
Abstract: Cystic fibrosis (CF) is the most common cause of chronic obstructive lung disease in children and young adults, yet there is no cure for this disease. Intimations of curative strategies have emerged from gene therapy, but these approaches have failed in clinical trials due to insufficient safety and efficacy profiles. The molecular technology necessary to overcome preeminent challenges in gene therapy may reside in programmable nucleases – namely CRISPR/Cas9 – which allow for site-specific genomic integration of therapeutic transgenes. However, the clinical utility of these technologies is limited by inherently low efficiencies. Here, we use flow cytometry and PCR assays to demonstrate the efficacy of perturbing DNA repair to overcome such limitations. We demonstrate that overexpressing CtIP and EXO1-4D can enhance the efficiency of CRISPR/Cas9-directed transgene integration by ~3- and ~6-fold, respectively. These discoveries may provide necessary impetus for translating gene therapies into clinical realities for genetic diseases such as CF.
Source: TSpace (University of Toronto)
Author: Kyle Seigel
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
Funder: Hospital for Sick Children
OpenAlex ID: W2907243755