Antisense oligonucleotide (ASO) therapy targeting C9orf72 represents one of the most promising disease-modifying approaches for amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) caused by hexanucleotide repeat expansions. This therapeutic strategy aims to reduce the levels of toxic dipeptide repeat proteins (DPRs) generated by aberrant translation of the expanded GGGGCC repeat in the C9orf72 gene. [1]
C9orf72 repeat expansions are the most common genetic cause of familial ALS and FTD, accounting for approximately 40% of familial ALS cases and 25% of familial FTD cases. The development of ASO therapies specifically targeting this mutation represents a precision medicine approach to neurodegenerative disease treatment. [2]
The C9orf72 gene contains a polymorphic hexanucleotide repeat expansion (GGGGCC) in its first intron that can expand from the normal 2-30 repeats to hundreds or even thousands of repeats in affected individuals. This expansion leads to disease through three primary mechanisms: [3]
RNA Toxicity: The expanded repeat RNA forms nuclear RNA foci that sequester essential RNA-binding proteins, including hnRNPs (heterogeneous nuclear ribonucleoproteins), disrupting normal RNA processing and splicing.
Dipeptide Repeat Protein (DPR) Toxicity: Aberrant translation of the expanded repeat in all three reading frames produces five distinct DPRs: poly-GA, poly-GP, poly-GR, poly-PA, and poly-PR. These aggregating proteins cause proteostasis disruption, nucleocytoplasmic transport defects, and mitochondrial dysfunction.
Loss of Function: The expansion reduces C9orf72 mRNA expression through epigenetic silencing, potentially disrupting normal lysosomal and autophagic functions in neurons and glial cells.
ASOs are short, single-stranded oligonucleotides (typically 12-25 nucleotides) that bind to complementary target RNA via Watson-Crick base pairing. For C9orf72 ASO therapy, the primary targets include: [4]
Repeat-Containing RNA: ASOs can be designed to bind specifically to the expanded repeat RNA, preventing the formation of toxic RNA foci and blocking aberrant DPR translation through a mechanism called repeat-associated non-AUG (RAN) translation.
Splice-Modifying ASOs: Alternative approaches target the C9orf72 pre-mRNA to modulate splicing, potentially increasing the expression of the more functional C9orf72 isoform or reducing toxic transcript variants.
The most advanced ASO candidates target the repeat RNA to suppress DPR production while preserving as much normal C9orf72 function as possible. [5]
Upon binding to target RNA, ASOs exert their effects through two primary mechanisms: [6]
RNase H1-Mediated Degradation: Gapmer ASOs contain a central DNA-like region that recruits RNase H1, an endonuclease that cleaves the RNA strand in DNA-RNA hybrids, leading to target RNA degradation.
Steric Blockade: Steric-blocking ASOs (non-gapmer) bind to target RNA without recruiting RNase H, instead blocking translation initiation, ribosomal assembly, or splice site recognition without degrading the target.
Patient-derived induced pluripotent stem cell (iPSC) models have been instrumental in validating ASO therapy for C9orf72 ALS/FTD: [7]
RNA Foci Reduction: Studies have demonstrated that ASO treatment effectively reduces C9orf72 RNA foci in patient-derived motor neurons and cortical neurons. This reduction correlates with decreased sequestration of RNA-binding proteins like hnRNPA1 and SRSF2. [8]
Dipeptide Repeat Protein Suppression: Multiple studies have shown that ASO treatment significantly reduces DPR protein levels in iPSC-derived neurons. For example, ASOs targeting the repeat region reduce poly-GA, poly-GR, and poly-PR levels by 60-90% depending on the cell model and ASO design. [9]
Rescue of Cellular Phenotypes: Beyond reducing toxic species, ASO treatment has been shown to rescue several disease-relevant cellular phenotypes: [10]
Transgenic mouse models expressing the human C9orf72 repeat expansion have provided critical in vivo validation: [11]
ASO delivery in these models demonstrates: [12]
The most advanced C9orf72 ASO program has been developed through collaboration between Biogen and Bristol Myers Squibb: [13]
BIIB078 (formerly BMS-986369): This ASO targets the C9orf72 repeat expansion and has undergone clinical evaluation: [14]
The clinical trials have investigated multiple dose levels administered via intrathecal injection to achieve direct CNS delivery. [15]
Additional ASO programs targeting C9orf72 have been advanced by other pharmaceutical companies and academic groups, though Biogen's program remains the most clinically advanced. [16]
While full clinical trial results continue to emerge, early data indicate:
Clinical trial data to date have identified several categories of adverse events:
Common (≥10%):
Injection-Related:
Potential Neurological:
The safety profile supports continued development, with risk mitigation strategies including:
ALS/FTD are chronic conditions requiring long-term treatment. Long-term considerations include:
The blood-brain barrier (BBB) presents a significant challenge for CNS therapeutics. Currently approved ASO therapies for neurological diseases utilize intrathecal (IT) administration to bypass the BBB:
Research continues on improving CNS delivery:
Conjugated ASOs: ASOs conjugated to ligands that engage transport receptors on the BBB (e.g., transferrin receptor) may enable intravenous delivery
AAV-Vectored ASO Delivery: Using AAV vectors to deliver ASO expression constructs could provide long-lasting ASO production from a single dose
Exosome-Mediated Delivery: Exploring natural extracellular vesicle platforms for CNS-targeted delivery
Even with direct CNS delivery, achieving uniform distribution throughout the brain remains challenging:
The therapeutic window for C9orf72 ASOs involves balancing:
ASO monotherapy may ultimately be combined with other therapeutic approaches:
Robust biomarkers are needed to guide patient selection and treatment response:
Future development may enable genotype-guided therapy:
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Zheng et al. C9orf72 ASO clinical trials (2023). 2023. ↩︎
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