Efficient Correction of Oncogenic KRAS and TP53 Mutations through CRISPR Base Editing
Shady Sayed1,2, Olga A Sidorova1, Alexander Hennig2,3
1Medical Systems Biology, Medical Faculty and University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany.
Abstract:
KRAS is the most frequently mutated oncogene in human cancer, and its activating mutations represent long-sought therapeutic targets. Programmable nucleases, particularly the CRISPR-Cas9 system, provide an attractive tool for genetically targeting KRAS mutations in cancer cells. Here, we show that cleavage of a panel of KRAS driver mutations suppresses growth in various human cancer cell lines, revealing their dependence on mutant KRAS. However, analysis of the remaining cell population after long-term Cas9 expression unmasked the occurence of oncogenic KRAS escape variants that were resistant to Cas9-cleavage. In contrast, the use of an adenine base editor to correct oncogenic KRAS mutations progressively depleted the targeted cells without the appearance of escape variants and allowed efficient and simultaneous correction of a cancer-associated TP53 mutation. Oncogenic KRAS and TP53 base editing was possible in patient-derived cancer organoids, suggesting that base editor approaches to correct oncogenic mutations could be developed for functional interrogation of vulnerabilities in a personalized manner for future precision oncology applications.
Significance:
Repairing KRAS mutations with base editors can be used for providing a better understanding of RAS biology and may lay the foundation for improved treatments for KRAS-mutant cancers.
Insights
Base editors offer a promising strategy for targeting KRAS mutations in cancer, unlike CRISPR-Cas9 which can lead to resistance. This approach enables precise correction of oncogenic KRAS and TP53 mutations, paving the way for personalized cancer therapies.
Area of Science:
- Oncology
- Molecular Biology
- Gene Editing
Background:
- KRAS mutations are prevalent in human cancers, driving tumor growth and representing key therapeutic targets.
- CRISPR-Cas9 has emerged as a tool for targeting KRAS mutations, but can lead to the development of resistant cancer cell variants.
Purpose of the Study:
- To investigate the efficacy of base editors in correcting oncogenic KRAS mutations and assess their potential for cancer therapy.
- To compare base editing with CRISPR-Cas9 in targeting KRAS mutations and evaluate the emergence of resistance.
Main Methods:
- Utilized CRISPR-Cas9 to cleave KRAS driver mutations in cancer cell lines, observing growth suppression and resistance.
- Employed adenine base editors to correct oncogenic KRAS mutations, monitoring for cell depletion and escape variants.
- Simultaneously corrected KRAS and TP53 mutations using base editing in patient-derived cancer organoids.
Main Results:
- CRISPR-Cas9-mediated cleavage of KRAS mutations suppressed cancer cell growth but resulted in the emergence of Cas9-resistant KRAS escape variants.
- Adenine base editing progressively depleted KRAS-mutant cells without generating escape variants.
- Simultaneous base editing of oncogenic KRAS and TP53 mutations was achieved in cancer organoids.
Conclusions:
- Base editors provide a more robust approach than CRISPR-Cas9 for targeting KRAS mutations, avoiding resistance.
- Base editing facilitates simultaneous correction of multiple oncogenic mutations, offering potential for personalized precision oncology.
- This study highlights the therapeutic potential of base editors for understanding RAS biology and developing novel treatments for KRAS-mutant cancers.
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