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.

中文翻译：

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通过 CRISPR 碱基编辑有效纠正致癌 KRAS 和 TP53 突变

KRAS 是人类癌症中最常见的突变癌基因，其激活突变代表了长期寻找的治疗靶点。可编程核酸酶，特别是 CRISPR-Cas9 系统，为基因靶向癌细胞中的 KRAS 突变提供了一种有吸引力的工具。在这里，我们展示了一组 KRAS 驱动突变的裂解抑制了各种人类癌细胞系的生长，揭示了它们对突变 KRAS 的依赖。然而，对长期 Cas9 表达后剩余细胞群的分析揭示了对 Cas9 裂解具有抗性的致癌 KRAS 逃逸变体的出现。相比之下，使用腺嘌呤碱基编辑器来纠正致癌的 KRAS 突变会逐渐耗尽目标细胞，而不会出现逃逸变异，并且可以有效地同时纠正与癌症相关的 TP53 突变。致癌 KRAS 和 TP53 碱基编辑在患者来源的癌症类器官中是可能的，这表明可以开发纠正致癌突变的碱基编辑方法，以个性化的方式对漏洞进行功能询问，以用于未来的精准肿瘤学应用。意义：使用碱基编辑器修复 KRAS 突变可用于更好地了解 RAS 生物学，并可能为改进 KRAS 突变癌症的治疗奠定基础。这表明可以开发纠正致癌突变的碱基编辑器方法，以个性化的方式对漏洞进行功能询问，以用于未来的精准肿瘤学应用。意义：使用碱基编辑器修复 KRAS 突变可用于更好地了解 RAS 生物学，并可能为改进 KRAS 突变癌症的治疗奠定基础。这表明可以开发纠正致癌突变的碱基编辑器方法，以个性化的方式对漏洞进行功能询问，以用于未来的精准肿瘤学应用。意义：使用碱基编辑器修复 KRAS 突变可用于更好地了解 RAS 生物学，并可能为改进 KRAS 突变癌症的治疗奠定基础。