Thesis
Reduction of APOC3 Activity by Inducing Exon 2 Removal from APOC3 Gene Transcript Using Antisense Oligonucleotides: A Potential Therapy for Atherosclerosis
Masters by Research, Murdoch University
2024
Abstract
Nucleic acid therapeutics have seen significant progress in recent years, and antisense oligonucleotides (AOs) stand out as the key innovations in this field. AOs are potent tools that can modulate gene expression at the RNA level, providing unique mechanisms to inhibit or restore protein functions. Thus, they are especially useful for treating genetic disorders that were previously difficult to treat.
Apolipoprotein C3 (APOC3), a key regulator of triglyceride metabolism, has emerged as a crucial target for treating cardiovascular diseases (CVDs), particularly atherosclerosis. Elevated levels of APOC3 have been linked to an increased risk of atherosclerotic events, and APOC3 is a validated target for atherosclerosis. Currently, there are two RNase H-dependent antisense oligonucleotides (AO) are being developed to treat cardiovascular diseases by targeting APOC3 mRNA. One of them, Volanesorsen, has already been approved, while the other, Olezarsen, is still in development. However, reports suggest that there have been some toxicity issues with Volanesorsen, while no such issues have been observed with Olezarsen. Additionally, injection site reactions have been observed as the most common adverse events for both AOs.
This study aimed to reduce the activity of APOC3 by utilising a different mechanism of AO, splice modulation. We intend to remove exon 2 from the APOC3 transcript to remove the start codon to prevent APOC3 translation and, hence, APOC3 activity. We designed AOs to induce exon 2 skipping from the APOC3 mRNA transcript after in silico analysis of the APOC3 transcript. These AOs were evaluated in hepatocyte cell lines, HepG2, where APOC3 is expressed. When used individually, all AOs induced low levels (~20-30%) of exon 2 skipping from APOC3 transcripts. To enhance exon 2 skipping efficacy, we tested combinations of AOs in a cocktail approach. This strategy led to a notable improvement, with exon skipping efficiencies increasing to approximately 54%. Despite this enhancement, it is imperative to analyse the level of APOC3 protein further to determine the therapeutic viability of the observed exon-skipping efficiencies.
In conclusion, this study showed that exon 2 splicing may be tightly regulated and that AO-induced exon skipping was not highly efficient even when more than one AO was used. This observation also indicates that not all gene targets may be amenable to exon skipping.
Details
- Title
- Reduction of APOC3 Activity by Inducing Exon 2 Removal from APOC3 Gene Transcript Using Antisense Oligonucleotides: A Potential Therapy for Atherosclerosis
- Authors/Creators
- Shaimaa A Jasim
- Contributors
- May Aung-Htut (Supervisor) - Murdoch University, Centre for Molecular Medicine and Innovative TherapeuticsCraig S. McIntosh (Supervisor) - Murdoch University, Centre for Molecular Medicine and Innovative Therapeutics
- Awarding Institution
- Murdoch University; Masters by Research
- Identifiers
- 991005672070307891
- Murdoch Affiliation
- Centre for Molecular Medicine and Innovative Therapeutics; School of Medical, Molecular and Forensic Sciences
- Resource Type
- Thesis
- Note
- Accelerated Research Masters with Training
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