Dual-isoform hUBE3A gene transfer improves behavioral and seizure outcomes in Angelman syndrome model mice
Matthew C. Judson, Charles Shyng, Jeremy M. Simon, Courtney R. Davis, A. Mattijs Punt, Mirabel T. Salmon, Noah W. Miller, Kimberly D. Ritola, Ype Elgersma, David G. Amaral, Steven J. Gray, Benjamin D. Philpot
Matthew C. Judson, Charles Shyng, Jeremy M. Simon, Courtney R. Davis, A. Mattijs Punt, Mirabel T. Salmon, Noah W. Miller, Kimberly D. Ritola, Ype Elgersma, David G. Amaral, Steven J. Gray, Benjamin D. Philpot
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Research Article Neuroscience Therapeutics

Dual-isoform hUBE3A gene transfer improves behavioral and seizure outcomes in Angelman syndrome model mice

Abstract

Loss of the maternal UBE3A allele causes Angelman syndrome (AS), a debilitating neurodevelopmental disorder. Here, we devised an AS treatment strategy based on reinstating dual-isoform expression of human UBE3A (hUBE3A) in the developing brain. Kozak sequence engineering of our codon-optimized vector (hUBE3Aopt) enabled translation of both short and long hUBE3A protein isoforms at a near-endogenous 3:1 (short/long) ratio, a feature that could help to support optimal therapeutic outcomes. To model widespread brain delivery and early postnatal onset of hUBE3A expression, we packaged the hUBE3Aopt vector into PHP.B capsids and performed intracerebroventricular injections in neonates. This treatment significantly improved motor learning and innate behaviors in AS mice, and it rendered them resilient to epileptogenesis and associated hippocampal neuropathologies induced by seizure kindling. hUBE3A overexpression occurred frequently in the hippocampus but was uncommon in the neocortex and other major brain structures; furthermore, it did not correlate with behavioral performance. Our results demonstrate the feasibility, tolerability, and therapeutic potential for dual-isoform hUBE3A gene transfer in the treatment of AS.

Authors

Matthew C. Judson, Charles Shyng, Jeremy M. Simon, Courtney R. Davis, A. Mattijs Punt, Mirabel T. Salmon, Noah W. Miller, Kimberly D. Ritola, Ype Elgersma, David G. Amaral, Steven J. Gray, Benjamin D. Philpot

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Figure 8

PHP. B/hUBE3Aopt-mediated UBE3A overexpression is largely restricted to the hippocampus and does not correlate with phenotypic outcome.

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PHP. B/hUBE3Aopt-mediated UBE3A overexpression is largely restricted to ...
(A–E) Mean ± SEM UBE3A immunofluorescence (IF) by brain region. *P < 0.05, ***P < 0.001, ****P < 0.0001. Welch’s ANOVA, Dunnett’s post hoc. (A) Left: neocortical UBE3A IF, counterstaining for NeuN and GABA. Arrowheads indicate putative excitatory neurons (white) and GABAergic interneurons (yellow). Middle: representative NeuN-based segmentation of neurons with (white arrowheads) or without (yellow arrowheads) UBE3A IF. Graphed distribution of UBE3A IF levels within individual neurons for each group (>44,000 neurons analyzed per animal, from a subset of animals corresponding to filled circles in mean IF graph). Red line depicts the distribution resulting from the simulated addition (n = 50,000 per simulation; 10 simulations) of randomly selected WT + vehicle and AS + AAV values. Right: quantification of the percentage of NeuN+ neurons costained for UBE3A. (B) Thalamic UBE3A IF, counterstaining for NeuN and parvalbumin (PV). (C) Cerebellar UBE3A IF, counterstaining for NeuN and PV. (D) Striatal UBE3A IF, counterstaining for NeuN and DARPP-32. Arrowheads indicate medium spiny neurons with (white) or without (yellow) UBE3A labeling. (E) Left: hippocampal UBE3A IF, counterstaining for NeuN and PV. Right: representative NeuN-based segmentation of neurons with (white arrowheads) or without (yellow arrowheads) UBE3A IF. Graphed distribution of UBE3A IF levels within individual neurons for each group (>2300 neurons analyzed per animal, from a subset of animals corresponding to filled circles in mean IF graph). (F) Pearson’s r correlation of normalized mean hippocampal UBE3A IF and phenotypic outcome in AS + AAV mice. LV, lateral ventricle; LD, lateral dorsal nucleus; AV, anteroventral nucleus; RT, reticular nucleus; wm, white matter; ic, internal capsule. Scale bars by level of magnification: 100 μm (low), 20 μm (high) (A and E); 250 μm (low), 30 μm (high) (B); 200 μm (low), 30 μm (high) (C); 200 μm (low), 20 μm (high) (D).