Antisense oligonucleotide therapy for neurodegenerative disease
Richard A. Smith, Timothy M. Miller, Koji Yamanaka, Brett P. Monia, Thomas P. Condon, Gene Hung, Christian S. Lobsiger, Chris M. Ward, Melissa McAlonis-Downes, Hongbing Wei, Ed V. Wancewicz, C. Frank Bennett, Don W. Cleveland
Richard A. Smith, Timothy M. Miller, Koji Yamanaka, Brett P. Monia, Thomas P. Condon, Gene Hung, Christian S. Lobsiger, Chris M. Ward, Melissa McAlonis-Downes, Hongbing Wei, Ed V. Wancewicz, C. Frank Bennett, Don W. Cleveland
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Research Article Neuroscience

Antisense oligonucleotide therapy for neurodegenerative disease

Abstract

Neurotoxicity from accumulation of misfolded/mutant proteins is thought to drive pathogenesis in neurodegenerative diseases. Since decreasing levels of proteins responsible for such accumulations is likely to ameliorate disease, a therapeutic strategy has been developed to downregulate almost any gene in the CNS. Modified antisense oligonucleotides, continuously infused intraventricularly, have been demonstrated to distribute widely throughout the CNS of rodents and primates, including the regions affected in the major neurodegenerative diseases. Using this route of administration, we found that antisense oligonucleotides to superoxide dismutase 1 (SOD1), one of the most abundant brain proteins, reduced both SOD1 protein and mRNA levels throughout the brain and spinal cord. Treatment initiated near onset significantly slowed disease progression in a model of amyotrophic lateral sclerosis (ALS) caused by a mutation in SOD1. This suggests that direct delivery of antisense oligonucleotides could be an effective, dosage-regulatable means of treating neurodegenerative diseases, including ALS, where appropriate target proteins are known.

Authors

Richard A. Smith, Timothy M. Miller, Koji Yamanaka, Brett P. Monia, Thomas P. Condon, Gene Hung, Christian S. Lobsiger, Chris M. Ward, Melissa McAlonis-Downes, Hongbing Wei, Ed V. Wancewicz, C. Frank Bennett, Don W. Cleveland

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Figure 2

Identifying antisense oligonucleotides that reduce rat SOD1 in vitro and in vivo.

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Identifying antisense oligonucleotides that reduce ra...
(A) We synthesized seventy-eight 24-mer, modified oligonucleotides complementary to rat SOD1 mRNA and transfected them at 150 nM into primary rat A10 cells. RNA was prepared 24 hours after transfection, and SOD1 mRNA levels were measured by quantitative RT-PCR. Oligonucleotides are displayed relative to their positions on the 462-nucleotide SOD1 coding sequence. Mean ± SD are shown (n = 4). UTR, untranslated region; ASOs, antisense oligonucleotides. (B) Oligonucleotides identified by the in vitro screen in A were evaluated in a similar transfection paradigm again using rat A10 cells and transfection of increasing concentrations of oligonucleotide to produce a dose-response curve. (C) Oligonucleotides SODr/h146144, SODr/h146145, SODr146192, and SODscrambled (a control oligonucleotide) were injected (37.5 mg/kg) 3 times per week intraperitoneally into adult rats for 3 weeks, after which time mRNA levels were measured in the liver, kidney, and brain. Mean ± SD are shown (n = 6). (D) SOD1 protein levels in liver extracts from animals treated with oligonucleotides SODr/h146144, SODr/h146145, SODr146192, and SODscrambled were measured by immunoblotting with an antibody to SOD1 (18). An immunoblot for tubulin was performed to verify protein loading.