Mokrane, Omar;
(2024)
Delivery of CRISPR-Cas9 to the brain for the treatment of CNS disease.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
CRISPR-Cas9 is a pioneering gene-editing technology with the potential to permanently correct gene mutations involved in disease pathogenesis. It’s application in CNS disorders is currently limited due to the BBB, which restricts the access of large biomolecules into CNS. Current CNS delivery strategies for CRISPR-Cas9 rely on intracranial injections to bypass the BBB which are invasive and have limited applicability in clinical settings. The nose-to-brain delivery route is a non-invasive pathway to the CNS which can bypass the BBB via perivascular and perineural pathways connecting the nasal epithelia and CNS. Polymeric delivery carriers can be utilised to overcome the barriers associated with the N2B route but are yet to be explored for CRISPR-Cas9 cargo. Therefore, this thesis aimed to develop a polymeric delivery system for CRISPR-Cas9 which could target the CNS via the N2B route. In the first instance, glycol chitosan polymers of different chain length were studied for their ability to compact CRISPR-pDNA into cationic, nanosized polyplex structures which are amenable to N2B delivery. A lead formulation (CRISPR-GC37) was identified and was assessed via in vitro cellular studies which identified a pH specific transfection mechanism which could be utilised to transfer the CRISPR-pDNA cargo via the N2B route. Novel methods were identified to concentrate polyplexes to clinically relevant concentrations whilst minimising aggregation. In vivo N2B studies demonstrated the transfer of the CRISPR-GC37 delivery system into the CNS via fluorescence labelling. Variable CNS expression of Cas9 was found, requiring further study.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Delivery of CRISPR-Cas9 to the brain for the treatment of CNS disease |
| Language: | English |
| Additional information: | Copyright © The Author 2024. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > UCL School of Pharmacy |
| URI: | https://discovery-pp.ucl.ac.uk/id/eprint/10193588 |
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