Department of Medical Oncology, Beijing Tuberculosis and Thoracic Tumor Research Institute/Beijing Chest Hospital, Capital Medical University, Beijing, 101149, China.
Department of Cellular and Molecular Biology, Beijing Tuberculosis and Thoracic Tumor Research Institute/Beijing Chest Hospital, Capital Medical University, Beijing, 101149, China.
J Cancer Res Clin Oncol. 2018 Aug;144(8):1413-1422. doi: 10.1007/s00432-018-2668-7. Epub 2018 May 24.
AZD9291 is an irreversible, small-molecule inhibitor which has potency against mutant EGFR- and T790M-resistant mutation. Despite the encouraging efficacy in clinical, the acquired resistance will finally occur. Further study will need to be done to identify the acquired resistance mechanisms and determine the next treatment.
We established an AZD9291-resistant cell line (HCC827/AZDR) from parental HCC827 cell line through stepwise pulsed selection of AZD9291. The expression of EGFR and its downstream pathways were determined by western blot analysis or immunofluorescence assay. The sensitivity to indicated agents were evaluated by MTS.
Compared with parental HCC827 cells, the HCC827/AZDR cells showed high resistance to AZD9291 and other EGFR-TKIs, and exhibited a mesenchymal-like phenotype. Almost complete loss of EGFR expression was observed in HCC827/AZDR cells. But the activation of downstream pathway, MAPK signaling, was found in HCC827/AZDR cells even in the presence of AZD9291. Inhibition of MAPK signaling had no effect on cell viability of HCC827/AZDR and could not reverse AZD9291 resistance because of the subsequent activation of AKT signaling. When treated with the combination of AKT and MAPK inhibitor, HCC827/AZDR showed remarkable growth inhibition.
Loss of EGFR could be proposed as a potential acquired resistance mechanism of AZD9291 in EGFR-mutant NSCLC cells with an EMT phenotype. Despite the loss of EGFR, the activation of MAPK pathway which had crosstalk with AKT pathway could maintain the proliferation and survival of resistant cells. Blocking MAPK and AKT signaling may be a potential therapeutic strategy following AZD9291 resistance.
AZD9291 是一种针对 EGFR 突变和 T790M 耐药突变的不可逆小分子抑制剂。尽管在临床治疗中疗效令人鼓舞，但最终还是会产生获得性耐药。需要进一步研究以确定获得性耐药机制，并确定下一步的治疗方案。
我们通过逐步脉冲选择 AZD9291，从亲本 HCC827 细胞系中建立了 AZD9291 耐药细胞系（HCC827/AZDR）。通过 Western blot 分析或免疫荧光检测来确定 EGFR 及其下游通路的表达。通过 MTS 评估对指示剂的敏感性。
与亲本 HCC827 细胞相比，HCC827/AZDR 细胞对 AZD9291 和其他 EGFR-TKI 表现出高度耐药性，并表现出间充质样表型。在 HCC827/AZDR 细胞中观察到 EGFR 表达几乎完全丧失。但是，即使在存在 AZD9291 的情况下，下游通路 MAPK 信号通路也被激活。MAPK 信号通路的抑制对 HCC827/AZDR 细胞的活力没有影响，并且不能逆转 AZD9291 耐药性，因为随后 AKT 信号通路被激活。当用 AKT 和 MAPK 抑制剂联合处理时，HCC827/AZDR 显示出明显的生长抑制。
在具有 EMT 表型的 EGFR 突变 NSCLC 细胞中，EGFR 的缺失可被提出作为 AZD9291 获得性耐药的潜在机制。尽管 EGFR 缺失，但与 AKT 通路有串扰的 MAPK 通路的激活可以维持耐药细胞的增殖和存活。阻断 MAPK 和 AKT 信号通路可能是 AZD9291 耐药后的一种潜在治疗策略。
