CNS-targeted base editing of the major late-onset Tay-Sachs mutation alleviates disease in mice
Maria L. Allende, Mari Kono, Y. Terry Lee, Samantha M. Olmsted, Vienna Huso, Jenna Y. Bakir, Florencia Pratto, Cuiling Li, Colleen Byrnes, Galina Tuymetova, Hongling Zhu, Cynthia J. Tifft, Richard L. Proia
Maria L. Allende, Mari Kono, Y. Terry Lee, Samantha M. Olmsted, Vienna Huso, Jenna Y. Bakir, Florencia Pratto, Cuiling Li, Colleen Byrnes, Galina Tuymetova, Hongling Zhu, Cynthia J. Tifft, Richard L. Proia
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Research Article Genetics Neuroscience

CNS-targeted base editing of the major late-onset Tay-Sachs mutation alleviates disease in mice

Abstract

Late-onset Tay-Sachs (LOTS) disease is a lysosomal storage disorder most commonly caused by a point mutation (c.805G>A) in the HEXA gene encoding the α subunit of the lysosomal enzyme β-hexosaminidase A. LOTS manifests as a range of gradually worsening neurological symptoms beginning in young adulthood. Here, we explored the efficacy of an adenine base editor (ABE) programmed with an sgRNA to correct the HEXA c.805G>A mutation. Base editing in fibroblasts from a patient with LOTS successfully converted the pathogenic HEXA c.805A to G and partially restored β-hexosaminidase activity, with minimal genome-wide off-target editing. We generated a LOTS mouse model in which the mice exhibited decreased β-hexosaminidase activity, accumulation of GM2 ganglioside in the brain, progressive neurological manifestations, and reduced lifespan. Treatment of LOTS mice with the neurotropic virus AAV-PHP.eB carrying the ABE and an sgRNA targeting the LOTS point mutation partially corrected the c.805G>A mutation in the CNS, significantly increased brain β-hexosaminidase activity, and substantially reduced GM2 ganglioside accumulation in the brain. Moreover, the therapy delayed symptom onset and significantly extended median lifespan. These findings highlight the potential of base editing as an effective treatment for LOTS and its broader applicability to other lysosomal storage disorders.

Authors

Maria L. Allende, Mari Kono, Y. Terry Lee, Samantha M. Olmsted, Vienna Huso, Jenna Y. Bakir, Florencia Pratto, Cuiling Li, Colleen Byrnes, Galina Tuymetova, Hongling Zhu, Cynthia J. Tifft, Richard L. Proia

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Figure 4

Base-editor treatment reduces brain GM2 ganglioside accumulation in LOTS mice.

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Base-editor treatment reduces brain GM2 ganglioside accumulation in LOTS...
WT, control-treated LOTS, and ABE-treated LOTS mice were euthanized at 21 weeks of age, and sagittal brain sections were prepared (n = 4 per group; mixed males and females). The AAV-treated mice each received 2.4 × 1012 vg. (A) Representative sagittal brain sections stained with anti-GM2 ganglioside antibody (red) and counterstained with DAPI (blue); scale bar: 1 mm. (B and C) Representative 40× images of the cerebral cortex (B) and brain stem (C) stained with anti-GM2 ganglioside antibody (red) and counterstained with DAPI (blue); scale bar: 20 μm. (D) Quantification of GM2 fluorescence intensity in cortex and brain stem. Small gray symbols represent image-level measurements (technical replicates); large colored symbols indicate per-mouse means (biological replicates). Statistical analysis was performed using a mixed-effects model with Tukey’s correction. ***P < 0.001, ****P < 0.0001. n = 3 for WT; n = 4 for ABE-treated; n = 3 for control-treated. (E) Representative 40× images of the cerebral cortex from control- and ABE-treated LOTS mice stained with anti-NeuN (green), anti-GM2 ganglioside (red), and counterstained with DAPI (blue). Merged images (right panels) show colocalization of GM2 and NeuN; scale bar: 20 μm. (F) Representative 40× images of the cerebral cortex from control- and ABE-treated LOTS mice stained with anti-LAMP1 (green, pseudocolored) and anti-GM2 ganglioside (red). Merged images (right panels) show colocalization of LAMP1 and GM2; scale bar: 10 μm.