Self-regulating gene therapy ameliorates phenotypes and overcomes gene dosage sensitivity in a mouse model of Rett syndrome

Ross, PD, Gadalla, KKE ORCID: https://orcid.org/0000-0002-1657-8953, Thomson, SR, Selfridge, J, Bahey, NG ORCID: https://orcid.org/0000-0002-3819-0108, Benito, J, Burstein, SR, McMinn, R, Bolon, B, Hector, RD and Cobb, SR, 2025. Self-regulating gene therapy ameliorates phenotypes and overcomes gene dosage sensitivity in a mouse model of Rett syndrome. Science Translational Medicine, 17 (792). ISSN 1946-6234

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Abstract

Conventional methods of gene transfer lead to inconsistent transgene expression within cells. This variability can be problematic, particularly in conditions like Rett syndrome (RTT), a neurological disorder caused by mutations in the MECP2 (methyl-CpG binding protein 2) gene, because overexpression of MECP2 can also cause adverse effects. To address these challenges, we devised a gene regulation system called Expression Attenuation via Construct Tuning (EXACT), which uses a self-contained, microRNA-based feed-forward loop that not only ensures more consistent transgene expression but also protects against excessive expression. Through cell-based screening assays, we demonstrated the ability of the EXACT circuit to modulate the expression of full-length human MeCP2. Compared with a conventional construct, an EXACT-MECP2 construct exhibited a narrower range of cellular protein abundance. Furthermore, the degree of regulation by the EXACT circuit increased with higher transgene doses in vitro and in wild-type mice and mice modeling RTT. On the basis of cellular and in vivo testing, we identified an optimal configuration for the adeno-associated virus serotype 9 (AAV9) construct for self-regulated MECP2 gene therapy, designated NGN-401. Delivery of NGN-401 to neonatal male Mecp2−/y hemizygous mice via intracerebroventricular injection resulted in prolonged survival and amelioration of RTT-like phenotypes compared with vehicle-treated animals. NGN-401 was also well tolerated by female Mecp2+/− mice and healthy juvenile nonhuman primates, in contrast with a conventional construct, which caused toxicity. The results from these studies underpin a first-in-human pediatric trial of NGN-401 in RTT (ClinicalTrials.gov, NCT05898620).

Item Type:	Journal article
Publication Title:	Science Translational Medicine
Creators:	Ross, P.D., Gadalla, K.K.E., Thomson, S.R., Selfridge, J., Bahey, N.G., Benito, J., Burstein, S.R., McMinn, R., Bolon, B., Hector, R.D. and Cobb, S.R.
Publisher:	American Association for the Advancement of Science (AAAS)
Date:	2 April 2025
Volume:	17
Number:	792
ISSN:	1946-6234
Identifiers:	Number Type 10.1126/scitranslmed.adq3614 DOI 2422591 Other
Rights:	This is the author’s version of the work. It is posted here by permission of the AAAS for personal use, not for redistribution. The definitive version was published in Science Translational Medicin on 2 April 2025; DOI: https://doi.org/10.1126/scitranslmed.adq3614.
Divisions:	Schools > School of Science and Technology
Record created by:	Jonathan Gallacher
Date Added:	08 Jul 2025 13:34
Last Modified:	08 Jul 2025 13:34
URI:	https://irep.ntu.ac.uk/id/eprint/53899

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