Partial Restoration of Brain Dystrophin and Behavioral Deficits by Exon Skipping in the Muscular Dystrophy X‐Linked (<i>mdx</i>) Mouse

Zarrouki, Faouzi; Relizani, Karima; Bizot, Flavien; Tensorer, Thomas; Garcı́a, Luis; Vaillend, Cyrille; Goyenvalle, Aurélie

doi:10.1002/ana.26409

Cited by 11 publications

(13 citation statements)

References 38 publications

(110 reference statements)

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“…Considering that ASO biodistribution in the CNS may be affected by various factors, such as ASO chemistry and charge, we used two different chemistries of ASO in this study: the Phosphorodiamidate Morpholino Oligomer (PMO) chemistry and the tricyclo-DNA (tcDNA) chemistry. PMOs have a neutral charge and are already approved by the FDA for the systemic treatment of DMD [ 8 ], while tcDNA is a charged, lipid-conjugated ASO that has previously shown therapeutic potential in mouse models of DMD [ 24 , 25 ]. Overall, our detailed comparative study provides useful insights into the local delivery and associated efficacy of ASOs in the CNS of mouse models of DMD.…”

Section: Introductionmentioning

confidence: 99%

Investigating the Impact of Delivery Routes for Exon Skipping Therapies in the CNS of DMD Mouse Models

Fergus

Gileadi²,

Montanaro³

et al. 2023

Cells

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Nucleic acid-based therapies have demonstrated great potential for the treatment of monogenetic diseases, including neurologic disorders. To date, regulatory approval has been received for a dozen antisense oligonucleotides (ASOs); however, these chemistries cannot readily cross the blood–brain barrier when administered systemically. Therefore, an investigation of their potential effects within the central nervous system (CNS) requires local delivery. Here, we studied the brain distribution and exon-skipping efficacy of two ASO chemistries, PMO and tcDNA, when delivered to the cerebrospinal fluid (CSF) of mice carrying a deletion in exon 52 of the dystrophin gene, a model of Duchenne muscular dystrophy (DMD). Following intracerebroventricular (ICV) delivery (unilateral, bilateral, bolus vs. slow rate, repeated via cannula or very slow via osmotic pumps), ASO levels were quantified across brain regions and exon 51 skipping was evaluated, revealing that tcDNA treatment invariably generates comparable or more skipping relative to that with PMO, even when the PMO was administered at higher doses. We also performed intra-cisterna magna (ICM) delivery as an alternative route for CSF delivery and found a biased distribution of the ASOs towards posterior brain regions, including the cerebellum, hindbrain, and the cervical part of the spinal cord. Finally, we combined both ICV and ICM injection methods to assess the potential of an additive effect of this methodology in inducing efficient exon skipping across different brain regions. Our results provide useful insights into the local delivery and associated efficacy of ASOs in the CNS in mouse models of DMD. These findings pave the way for further ASO-based therapy application to the CNS for neurological disease.

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Section: Introductionmentioning

confidence: 99%

Investigating the Impact of Delivery Routes for Exon Skipping Therapies in the CNS of DMD Mouse Models

Fergus

Gileadi²,

Montanaro³

et al. 2023

Cells

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“…Dp427 loss particularly affects the number, size, and distribution of GABA A R clusters containing the α1 and/or α2 subunits within certain brain structures [ 5 , 39 ]. Preclinical studies have shown that Dp427 rescue by exon-skipping strategies in the adult brain of mdx mice has the potential to restore GABA A R clustering, as well as the associated excitatory-synapse plasticity and emotional disturbances [ 19 , 20 , 22 , 30 , 31 ]. This suggests that alterations of local inhibitory networks at least partly underlie the behavioral and cognitive deficits associated with DMD, and that changes in specific GABA A R subunits may serve as key readouts to evaluate the efficacy of genetic therapies.…”

Section: Discussionmentioning

confidence: 99%

“…The resulting electropherograms were inspected to check whether automatic peak detection required any manual correction. A 6-point calibration curve was loaded, made of a mix of WT and dystrophin-deficient control lysates to obtain defined percentages of dystrophin (0, 5, 10, 15% or 0, 10, 15, 30% of corresponding WT tissues) as previously described for our Western blot analyses [ 30 ].…”

Section: Methodsmentioning

confidence: 99%

“…However, the molecular studies in mdx mice have to date been restricted to the α1 and α2 subunits. This should be extended to the expression and distribution of several synaptic and extrasynaptic subunits in various brain structures to better understand the impact of Dp427 loss on inhibitory networks and to identify robust and specific molecular outcome measures for preclinical rescue experiments [ 20 , 30 , 31 ].…”

Section: Introductionmentioning

confidence: 99%

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Abnormal Expression of Synaptic and Extrasynaptic GABAA Receptor Subunits in the Dystrophin-Deficient mdx Mouse

Zarrouki

Goutal

Vacca

et al. 2022

IJMS

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Duchenne muscular dystrophy (DMD) is a neurodevelopmental disorder primarily caused by the loss of the full-length Dp427 dystrophin in both muscle and brain. The basis of the central comorbidities in DMD is unclear. Brain dystrophin plays a role in the clustering of central gamma-aminobutyric acid A receptors (GABAARs), and its loss in the mdx mouse alters the clustering of some synaptic subunits in central inhibitory synapses. However, the diversity of GABAergic alterations in this model is still fragmentary. In this study, the analysis of in vivo PET imaging of a benzodiazepine-binding site radioligand revealed that the global density of central GABAARs is unaffected in mdx compared with WT mice. In contrast, semi-quantitative immunoblots and immunofluorescence confocal imaging in tissue sections revealed complex and differential patterns of alterations of the expression levels and/or clustered distribution of a variety of synaptic and extrasynaptic GABAAR subunits in the hippocampus, cerebellum, cortex, and spinal cord. Hence, dystrophin loss not only affects the stabilization of synaptic GABAARs but also influences the subunit composition of GABAARs subtypes at both synaptic and extrasynaptic sites. This study provides new molecular outcome measures and new routes to evaluate the impact of treatments aimed at compensating alterations of the nervous system in DMD.

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“…They found that in particular the number and position of guanine nucleotides in the 3′-end of the ASO increased toxicity, while there was a decrease when adenine nucleotides were substituted [ 27 ]. Recent studies demonstrated that reducing the PS content of ONDs leads to increased tolerability in the CNS [ 138 , 139 ].…”

Section: Toxicities Associated With Cns Local Deliverymentioning

confidence: 99%

Considerations in the Preclinical Assessment of the Safety of Antisense Oligonucleotides

Goyenvalle

Jiménez-Mallebrera

Roon-Mom

et al. 2023

Nucleic Acid Therapeutics

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The nucleic acid therapeutics field has made tremendous progress in the past decades. Continuous advances in chemistry and design have led to many successful clinical applications, eliciting even more interest from researchers including both academic groups and drug development companies. Many preclinical studies in the field focus on improving the delivery of antisense oligonucleotide drugs (ONDs) and/or assessing their efficacy in target tissues, often neglecting the evaluation of toxicity, at least in early phases of development. A series of consensus recommendations regarding regulatory considerations and expectations have been generated by the Oligonucleotide Safety Working Group and the Japanese Research Working Group for the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use S6 and Related Issues (WGS6) in several white papers. However, safety aspects should also be kept in sight in earlier phases while screening and designing OND to avoid subsequent failure in the development phase. Experts and members of the network “DARTER,” a COST Action funded by the Cooperation in Science and Technology of the EU, have utilized their collective experience working with OND, as well as their insights into OND-mediated toxicities, to generate a series of consensus recommendations to assess OND toxicity in early stages of preclinical research. In the past few years, several publications have described predictive assays, which can be used to assess OND-mediated toxicity in vitro or ex vivo to filter out potential toxic candidates before moving to in vivo phases of preclinical development, that is, animal toxicity studies. These assays also have the potential to provide translational insight since they allow a safety evaluation in human in vitro systems. Yet, small preliminary in vivo studies should also be considered to complement this early assessment. In this study, we summarize the state of the art and provide guidelines and recommendations on the different tests available for these early stage preclinical assessments.

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Partial Restoration of Brain Dystrophin and Behavioral Deficits by Exon Skipping in the Muscular Dystrophy X‐Linked (mdx) Mouse

Cited by 11 publications

References 38 publications

Investigating the Impact of Delivery Routes for Exon Skipping Therapies in the CNS of DMD Mouse Models

Investigating the Impact of Delivery Routes for Exon Skipping Therapies in the CNS of DMD Mouse Models

Abnormal Expression of Synaptic and Extrasynaptic GABAA Receptor Subunits in the Dystrophin-Deficient mdx Mouse

Considerations in the Preclinical Assessment of the Safety of Antisense Oligonucleotides

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