Application of amplicon-based targeted sequencing with the molecular barcoding system to detect uncommon minor EGFR mutations in patients with treatment-naive lung adenocarcinoma
Application of amplicon-based targeted sequencing with the molecular barcoding system to detect uncommon minor EGFR mutations in patients with treatment-naive lung adenocarcinoma
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DOI：
10.1186/s12885-019-5374-1
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发表时间：
2019-02-26
期刊：
影响因子：
3.8
通讯作者：
Toyooka, Shinichi
中科院分区：
文献类型：
--
作者：
Namba, Kei;Tomida, Shuta;Toyooka, Shinichi
背景在肺癌中，表皮生长因子受体(EGFR)酪氨酸激酶抑制物致敏突变与罕见的EGFR微小突变共存，称为复合突变。这些微小的EGFR突变可导致EGFR酪氨酸激酶抑制剂治疗后的获得性耐药，因此确定患者的突变状态非常重要。然而，基于下一代测序的基于扩增子的靶向深度测序来表征突变，容易出现测序错误。因此，我们评估了分子条形码和高通量测序相结合的益处，以调查接受非小细胞肺癌切除的未接受治疗的患者的基因组异质性。方法我们利用分子条形码系统(MBS)进行基于扩增子的靶向测序，检测新鲜冷冻肺癌标本中常见的主要EGFR突变和罕见的微小突变，等位基因频率为0.5%。结果在63例(98.4%)病例中，MBS检测到的常见的EGFR突变与临床检测结果一致。EGFR异常突变7例(10.9%)。在三种主要的表皮生长因子受体突变中，G719X突变患者的复合突变发生率显著高于L858R突变或外显子19缺失的患者(p=0.0052)。这在癌症基因组图谱数据集的独立队列中得到了验证(p=0.018)。结论我们的研究结果表明，使用MBS来建立准确的非小细胞肺癌患者基因是可行的。这项工作将有助于了解非小细胞肺癌中EGFR复合突变的分子基础，并可能有助于开发新的治疗方式。
BackgroundIn lung cancer, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor sensitizing mutations co-existing with rare minor EGFR mutations are known as compound mutations. These minor EGFR mutations can lead to acquired resistance after EGFR tyrosine kinase inhibitor treatment, so determining the mutation status of patients is important. However, using amplicon-based targeted deep sequencing based on next-generation sequencing to characterize mutations is prone to sequencing error. We therefore assessed the benefit of incorporating molecular barcoding with high-throughput sequencing to investigate genomic heterogeneity in treatment-naive patients who have undergone resection of their non-small cell lung cancer (NSCLC) EGFR mutations.MethodsWe performed amplicon-based targeted sequencing with the molecular barcoding system (MBS) to detect major common EGFR mutations and uncommon minor mutations at a 0.5% allele frequency in fresh-frozen lung cancer samples.ResultsProfiles of the common mutations of EGFR identified by MBS corresponded with the results of clinical testing in 63 (98.4%) out of 64 cases. Uncommon mutations of EGFR were detected in seven cases (10.9%). Among the three types of major EGFR mutations, patients with the G719X mutation had a significantly higher incidence of compound mutations than those with the L858R mutation or exon 19 deletion (p=0.0052). This was validated in an independent cohort from the Cancer Genome Atlas dataset (p=0.018).ConclusionsOur findings demonstrate the feasibility of using the MBS to establish an accurate NSCLC patient genotype. This work will help understand the molecular basis of EGFR compound mutations in NSCLC, and could aid the development of new treatment modalities.