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Updated: May 23, 2025
A Nonsequencing Approach for the Rapid Detection of RNA Editing
Published on: April 21, 2022
RNA Editing-Mediated Correction of TP53 Nonsense Mutations via Lipid Nanoparticle-Delivered Circular ADAR-Recruiting
Jinjin Wang1, Wenjing Zhang1, Shuguang Li1
1School of Pharmaceutical Sciences, Tianjian Laboratory of Advanced Biomedical Sciences, Zhengzhou University, Zhengzhou 450001, P. R. China.
Abstract:
Nonsense mutations account for over 20% of disease-associated mutations, which refer to the occurrence of premature termination codons (PTCs) in gene sequences, resulting in truncated and dysfunctional proteins. Nonetheless, due to poor accessibility of precise target sites and the limitations of gene editing tools, there is still a lack of safe, effective, and site-specific approach for correction of nonsense mutations. Here, we designed a circular ADAR-recruiting RNA (Circ-arRNA) for the in vivo RNA editing-mediated repair of the TP53-W53X nonsense mutation. Compared with linear arRNA, Circ-RNA demonstrates strong intracellular stability and high efficiency for site-specific correction of the TP53-W53X nonsense mutant, with no detectable off-target effects on bystander bases. In triple-negative breast cancer TP53-W53X 4T1 cells and tumor-bearing mouse models, we used lipid nanoparticles (LNPs) to encapsulate and deliver Circ-arRNA, which achieved mutation correction efficiencies of 73.32 and 48.48%, respectively. Furthermore, Circ-arRNA LNPs effectively restored full-length p53 protein expression and its functional activity, significantly enhancing the sensitivity of tumor-bearing mice to paclitaxel chemotherapy. Our research demonstrated the safety and efficacy of LNP-based circular arRNA for specifically the repair of nonsense mutations in vivo, highlighting the immense potential of ADAR-mediated editing for correcting such mutations.
Insights
Circular ADAR-recruiting RNA (Circ-arRNA) offers a safe and effective method for correcting nonsense mutations in vivo. This novel RNA editing approach successfully repaired the TP53-W53X mutation, restoring protein function and enhancing cancer treatment sensitivity.
Area of Science:
- Molecular Biology
- Gene Editing
- RNA Therapeutics
Background:
- Nonsense mutations, causing over 20% of genetic diseases, lead to truncated proteins due to premature termination codons (PTCs).
- Current gene editing tools face limitations in precise targeting and accessibility, hindering effective correction of these mutations.
- Developing safe, site-specific, and efficient methods for in vivo nonsense mutation repair remains a critical challenge.
Purpose of the Study:
- To design and evaluate a circular ADAR-recruiting RNA (Circ-arRNA) for in vivo RNA editing-mediated repair of the TP53-W53X nonsense mutation.
- To assess the stability, efficiency, and specificity of Circ-arRNA compared to linear constructs.
- To investigate the therapeutic potential of Circ-arRNA delivered via lipid nanoparticles (LNPs) in cancer models.
Main Methods:
- Design of a circular ADAR-recruiting RNA (Circ-arRNA) targeting the TP53-W53X mutation.
- In vitro and in vivo evaluation of Circ-arRNA stability and editing efficiency.
- Delivery of Circ-arRNA using lipid nanoparticles (LNPs) in triple-negative breast cancer 4T1 cells and tumor-bearing mouse models.
- Assessment of p53 protein restoration, functional activity, and impact on chemotherapy sensitivity.
Main Results:
- Circ-arRNA demonstrated superior intracellular stability and high efficiency for site-specific correction of the TP53-W53X mutation.
- No detectable off-target editing effects were observed on bystander bases.
- In 4T1 cells and mouse models, Circ-arRNA LNP delivery achieved mutation correction efficiencies of 73.32% and 48.48%, respectively.
- Restoration of full-length p53 protein expression and functional activity was achieved, enhancing sensitivity to paclitaxel chemotherapy.
Conclusions:
- LNP-based Circ-arRNA is a safe and effective strategy for in vivo site-specific repair of nonsense mutations.
- ADAR-mediated RNA editing holds significant potential for correcting disease-causing nonsense mutations.
- This approach offers a promising therapeutic avenue for cancers harboring TP53 nonsense mutations and potentially other genetic disorders.
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