Department of Neurology
Molecular and Cellular Neuroscience | Nervous System | Nervous System Diseases | Neurology
BACKGROUND: Frontotemporal lobar degeneration (FTLD) is a devastating and progressive disorder, and a common cause of early onset dementia. Progranulin (PGRN) haploinsufficiency due to autosomal dominant mutations in the progranulin gene (GRN) is an important cause of FTLD (FTLD-GRN), and nearly a quarter of these genetic cases are due to a nonsense mutation. Premature termination codons (PTC) can be therapeutically targeted by compounds allowing readthrough, and aminoglycoside antibiotics are known to be potent PTC readthrough drugs. Restoring endogenous PGRN through PTC readthrough has not previously been explored as a therapeutic intervention in FTLD.
METHODS: We studied whether the aminoglycoside G418 could increase PGRN expression in HEK293 and human induced pluripotent stem cell (hiPSC)-derived neurons bearing the heterozygous S116X, R418X, and R493X pathogenic GRN nonsense mutations. We further tested a novel substituted phthalimide PTC readthrough enhancer in combination with G418 in our cellular models. We next generated a homozygous R493X knock-in hiPSC isogenic line (R493X(-/-) KI), assessing whether combination treatment in hiPSC-derived neurons and astrocytes could increase PGRN and ameliorate lysosomal dysfunction relevant to FTLD-GRN. To provide in vivo proof-of-concept of our approach, we measured brain PGRN after intracerebroventricular administration of G418 in mice expressing the V5-tagged GRN nonsense mutation R493X.
RESULTS: The R418X and R493X mutant GRN cell lines responded to PTC readthrough with G418, and treatments increased PGRN levels in R493X(-/-) KI hiPSC-derived neurons and astrocytes. Combining G418 with a PTC readthrough enhancer increased PGRN levels over G418 treatment alone in vitro. PGRN deficiency has been shown to impair lysosomal function, and the mature form of the lysosomal protease cathepsin D is overexpressed in R493X(-/-) KI neurons. Increasing PGRN through G418-mediated PTC readthrough normalized this abnormal lysosomal phenotype in R493X(-/-) KI neuronal cultures. A single intracerebroventricular injection of G418 induced GRN PTC readthrough in 6-week-old AAV-GRN-R493X-V5 mice.
CONCLUSIONS: Taken together, our findings suggest that PTC readthrough may be a potential therapeutic strategy for FTLD caused by GRN nonsense mutations.
Frontotemporal lobar degeneration, G418, GRN, Induced pluripotent stem cell, Neurons, Nonsense mutation, Premature termination codon, Progranulin, Readthrough
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DOI of Published Version
Frew J, Baradaran-Heravi A, Balgi AD, Wu X, Yan TD, Arns S, Shidmoossavee FS, Tan J, Jaquith JB, Jansen-West KR, Lynn FC, Gao FB, Petrucelli L, Feldman HH, Mackenzie IR, Roberge M, Nygaard HB. Premature termination codon readthrough upregulates progranulin expression and improves lysosomal function in preclinical models of GRN deficiency. Mol Neurodegener. 2020 Mar 16;15(1):21. doi: 10.1186/s13024-020-00369-5. PMID: 32178712; PMCID: PMC7075020. Link to article on publisher's site
Frew J, Gao F, Nygaard HB. (2020). Premature termination codon readthrough upregulates progranulin expression and improves lysosomal function in preclinical models of GRN deficiency. Open Access Articles. https://doi.org/10.1186/s13024-020-00369-5. Retrieved from https://escholarship.umassmed.edu/oapubs/4192
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This work is licensed under a Creative Commons Attribution 4.0 License.