Developmental and Molecular Pathways, Novartis Institutes for Biomedical Research, Cambridge, Massachusetts, USA.
Nat Chem Biol. 2014 Aug;10(8):623-5. doi: 10.1038/nchembio.1550. Epub 2014 Jun 15.
Identification and validation of drug-resistant mutations can provide important insights into the mechanism of action of a compound. Here we demonstrate the feasibility of such an approach in mammalian cells using next-generation sequencing of drug-resistant clones and CRISPR-Cas9-mediated gene editing on two drug-target pairs, 6-thioguanine-HPRT1 and triptolide-ERCC3. We showed that disrupting functional HPRT1 allele or introducing ERCC3 point mutations by gene editing can confer drug resistance in cells.
鉴定和验证耐药突变可以为化合物的作用机制提供重要的见解。在这里，我们使用耐药克隆的下一代测序和 CRISPR-Cas9 介导的基因编辑在两种药物靶对 6-硫代鸟嘌呤-HPRT1 和雷公藤甲素-ERCC3 上证明了这种方法在哺乳动物细胞中的可行性。我们表明，通过基因编辑破坏功能性 HPRT1 等位基因或引入 ERCC3 点突变可以赋予细胞耐药性。
