Stilla Technologies, Villejuif, France.
Methods Mol Biol. 2021;2279:127-144. doi: 10.1007/978-1-0716-1278-1_10.
The profiling of EGFR mutations, the most common genetic alterations in non-small cell lung cancer (NSCLC) predictive of targeted therapy efficacy, is crucial to anticipate the patient response to EGFR tyrosine kinase inhibitors. Here, we introduce the naica system for 6-color Crystal Digital PCR and describe in detail a standardized workflow for the multiplexed, single-assay detection of the 19 most prevalent sensitizing and resistance EGFR mutations in both tumor and circulating tumor DNA (ctDNA) samples. Two major advantages of the 6-color multiplexing system over current digital PCR systems are the rapid time to results, and the large quantity of mutational information obtained per patient sample, rendering the 6-color system highly cost effective. The 6-color Crystal Digital PCR technology enables highly sensitive and efficient therapeutic monitoring through liquid biopsy, resulting in the early detection of treatment resistance. While the assay presented here specifically addresses EGFR mutation status monitoring in NSCLC patients, 6-color Crystal Digital PCR assays are flexible and evolutive in design. As such, 6-color detection assays can be optimized to monitor mutations associated with a range of cancers and other genetic diseases, as well as to detect genetic changes beyond the oncology and human health domains.
EGFR 突变谱分析是预测非小细胞肺癌（NSCLC）靶向治疗疗效的最常见遗传改变，对于预测患者对 EGFR 酪氨酸激酶抑制剂的反应至关重要。在这里，我们介绍了用于 6 色 Crystal Digital PCR 的 naica 系统，并详细描述了一种标准化工作流程，用于在肿瘤和循环肿瘤 DNA（ctDNA）样本中同时检测最常见的 19 种敏化和耐药性 EGFR 突变，该工作流程采用多重单测定法。与当前的数字 PCR 系统相比，6 色多重系统具有两个主要优势，即快速获得结果，以及每个患者样本获得大量的突变信息，从而使 6 色系统具有很高的成本效益。6 色 Crystal Digital PCR 技术通过液体活检实现了高度敏感和高效的治疗监测，从而早期检测治疗耐药性。虽然这里介绍的检测方法专门针对 NSCLC 患者的 EGFR 突变状态监测，但 6 色 Crystal Digital PCR 检测方法在设计上具有灵活性和可扩展性。因此，6 色检测方法可以进行优化，以监测与一系列癌症和其他遗传疾病相关的突变，以及检测肿瘤学和人类健康领域以外的遗传变化。
