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Deimmunization for gene therapy: host matching of synthetic zinc finger constructs enables long-term mutant Huntingtin repression in mice

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dc.contributor.author Agustín Pavón, Carmen
dc.contributor.author Mielcarek, Michael
dc.contributor.author Garriga-Canut, Mireia
dc.contributor.author Isalan, Mark
dc.date.accessioned 2016-11-14T16:40:40Z
dc.date.available 2016-11-14T16:40:40Z
dc.date.issued 2016
dc.identifier.citation Agustín-Pavón C, Mielcarek M, Garriga-Canut M, Isalan M. Deimmunization for gene therapy: host matching of synthetic zinc finger constructs enables long-term mutant Huntingtin repression in mice. Molecular Neurodegeneration. 2016;11(1):64. DOI: 10.1186/s13024-016-0128-x
dc.identifier.issn 1750-1326
dc.identifier.uri http://hdl.handle.net/10230/27505
dc.description.abstract BACKGROUND: Synthetic zinc finger (ZF) proteins can be targeted to desired DNA sequences and are useful tools for gene therapy. We recently developed a ZF transcription repressor (ZF-KOX1) able to bind to expanded DNA CAG-repeats in the huntingtin (HTT) gene, which are found in Huntington's disease (HD). This ZF acutely repressed mutant HTT expression in a mouse model of HD and delayed neurological symptoms (clasping) for up to 3 weeks. In the present work, we sought to develop a long-term single-injection gene therapy approach in the brain. METHOD: Since non-self proteins can elicit immune and inflammatory responses, we designed a host-matched analogue of ZF-KOX1 (called mZF-KRAB), to treat mice more safely in combination with rAAV vector delivery. We also tested a neuron-specific enolase promoter (pNSE), which has been reported as enabling long-term transgene expression, to see whether HTT repression could be observed for up to 6 months after AAV injection in the brain. RESULTS: After rAAV vector delivery, we found that non-self proteins induce significant inflammatory responses in the brain, in agreement with previous studies. Specifically, microglial cells were activated at 4 and 6 weeks after treatment with non-host-matched ZF-KOX1 or GFP, respectively, and this was accompanied by a moderate neuronal loss. In contrast, the host-matched mZF-KRAB did not provoke these effects. Nonetheless, we found that using a pCAG promoter (CMV early enhancer element and the chicken β-actin promoter) led to a strong reduction in ZF expression by 6 weeks after injection. We therefore tested a new non-viral promoter to see whether the host-adapted ZF expression could be sustained for a longer time. Vectorising mZF-KRAB with a promoter-enhancer from neuron-specific enolase (Eno2, rat) resulted in up to 77 % repression of mutant HTT in whole brain, 3 weeks after bilateral intraventricular injection of 10(10) virions. Importantly, repressions of 48 % and 23 % were still detected after 12 and 24 weeks, respectively, indicating that longer term effects are possible. CONCLUSION: Host-adapted ZF-AAV constructs displayed a reduced toxicity and a non-viral pNSE promoter improved long-term ZF protein expression and target gene repression. The optimized constructs presented here have potential for treating HD.
dc.description.sponsorship Authors were funded by the European Research Council grants: FP7 ERC 201249 ZINC-HUBS and H2020 - ERC-2014-PoC 641232 - Fingers4Cure.
dc.format.mimetype application/pdf
dc.language.iso eng
dc.publisher BioMed Central
dc.relation.ispartof Molecular Neurodegeneration. 2016;11(1):64
dc.rights © 2016 The Author(s). Open Access. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.
dc.rights.uri http://creativecommons.org/licenses/by/4.0/
dc.title Deimmunization for gene therapy: host matching of synthetic zinc finger constructs enables long-term mutant Huntingtin repression in mice
dc.type info:eu-repo/semantics/article
dc.identifier.doi http://dx.doi.org/10.1186/s13024-016-0128-x
dc.subject.keyword Monogenetic disease
dc.subject.keyword Gene therapy
dc.subject.keyword Huntington’s disease
dc.subject.keyword Neurodegenerative disorder
dc.subject.keyword Immune response
dc.subject.keyword Synthetic transcription factors
dc.subject.keyword rAAV
dc.subject.keyword Host optimization
dc.relation.projectID info:eu-repo/grantAgreement/EC/FP7/201249
dc.relation.projectID info:eu-repo/grantAgreement/EC/H2020/641232
dc.rights.accessRights info:eu-repo/semantics/openAccess
dc.type.version info:eu-repo/semantics/publishedVersion

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