Introduction: Dacomitinib was superior to gefitinib in terms of progression‐free survival in patients with EGFR‐mutant lung cancer in a recent ARCHER 1050 trial. However, despite a marked initial response, lung cancers eventually acquire resistance to these inhibitors. This study aimed to elucidate the mechanisms of acquired resistance to dacomitinib in vitro. Methods: Dacomitinib‐resistant clones were established by exposure to fixed concentrations of dacomitinib by using N‐ethyl‐N‐nitrosourea (ENU) mutagenesis or by chronic exposure to increasing concentrations of dacomitinib without ENU. EGFR secondary mutations were analyzed by Sanger sequencing. Time to resistance in each clone was compared according to the mutational status. EGFR Del19, L858R, and G719A mutations were introduced into Ba/F3 cells by using retroviral vectors. Results: Chronic exposure to dacomitinib without ENU induced T790M in Ba/F3 cells expressing Del19. ENU mutagenesis resulted in 171 dacomitinib‐resistant clones. Among these clones, 90% acquired T790M. However, C797S occurred in 11% of L858R‐mutant clones (four of 35) and in 24% of G719A‐mutant clones (12 of 38) established by using low‐dose dacomitinib. Time to resistance was not significantly different between T790M‐ and C797S‐mutant clones in both of L858R clones (p = 0.93) and G719A clones (p = 0.86). Cells expressing Del19 that acquired T790M were sensitive to osimertinib, whereas cells with L858R plus C797S mutations were sensitive to gefitinib or erlotinib. Conclusions: These in vitro data demonstrate that dacomitinib can directly induce T790M or C797S secondary mutations. Our data suggest the importance of analyzing these secondary mutations because appropriate selection of EGFR inhibitors could overcome acquired resistance to dacomitinib in a subset of lung cancers.

中文翻译：

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EGFR T790M 和 C797S 突变作为达克替尼获得性耐药机制

简介：在最近的 ARCHER 1050 试验中，达克替尼在 EGFR 突变肺癌患者的无进展生存期方面优于吉非替尼。然而，尽管有明显的初始反应，但肺癌最终会对这些抑制剂产生耐药性。本研究旨在阐明体外达克替尼获得性耐药的机制。方法：通过使用 N-乙基-N-亚硝基脲 (ENU) 诱变暴露于固定浓度的达克替尼或通过长期暴露于没有 ENU 的增加浓度的达克替尼来建立达克替尼耐药克隆。通过 Sanger 测序分析 EGFR 二次突变。根据突变状态比较每个克隆的抗性时间。通过使用逆转录病毒载体将EGFR Del19、L858R和G719A突变引入Ba/F3细胞。结果：在表达 Del19 的 Ba/F3 细胞中，长期暴露于无 ENU 的达克替尼诱导 T790M。ENU 突变导致 171 个达克替尼耐药克隆。在这些克隆中，90% 获得了 T790M。然而，C797S 发生在 11% 的 L858R 突变克隆（35 个中的 4 个）和使用低剂量达克替尼建立的 G719A 突变克隆（38 个中的 12 个）中的 24%。在 L858R 克隆 (p = 0.93) 和 G719A 克隆 ​​(p = 0.86) 中，T790M 和 C797S 突变克隆之间的抗性时间没有显着差异。获得 T790M 的表达 Del19 的细胞对奥希替尼敏感，而具有 L858R 加 C797S 突变的细胞对吉非替尼或厄洛替尼敏感。结论：这些体外数据表明达克替尼可以直接诱导 T790M 或 C797S 二次突变。