Department of Oncology, Guangzhou General Hospital of Guangzhou Military Command, Guangzhou, Guangdong, China.
PLoS One. 2014 Dec 4;9(12):e114074. doi: 10.1371/journal.pone.0114074. eCollection 2014.
Recent clinical trials showed that the sequential combination of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) and chemotherapy could prolong the PFS and/or OS of advanced non-small cell lung cancer (NSCLC) patients with EGFR mutation. The aim of present study was to assess the optimal combination sequence and to explore its possible mechanism.
PC-9 cells and A549 cells, the lung adenocarcinoma cells with mutant and wide-type EGFR respectively, were treated with docetaxel/gefitinib alone or in different combination schedules. The EGFR and K-ras gene status was determined by qPCR-HRM technique. Cell proliferation was detected by MTT assay. The expression and phosphorylation of EGFR, ERK, Akt and IGF-1R were detected by western blot. Cell cycle distribution was observed by flow cytometry.
Only sequential administration of docetaxel followed by gefitinib (D → G) induced significant synergistic effect in both cell lines (Combination Index<0.9). The reverse sequence (G → D) resulted in an antagonistic interaction in both cell lines (CI>1.1), whereas the concurrent administration (D+G) showed additive (0.9<CI<1.1)-synergistic effect in PC-9 cells and antagonistic-additive effect in A549 cells. Mechanism studies showed that docetaxel-induced phosphorylation of EGFR and ERK was repressed by subsequently used gefitinib, but not by concurrent exposure of gefitinib. The gefitinib-repressed phosphorylation of EGFR and ERK was reversed neither by concurrent nor by subsequent administration of docetaxel. D+G reinforced their inhibition on the phosphorylation of IGF-1R in PC-9 cells.
The cytotoxic drugs followed by EGFR-TKIs may be the optimal combination for antiproliferative effects in EGFR-mutant NSCLC cells, and the phosphorylation of EGFR and ERK might contribute to this effect.
最近的临床试验表明，表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKIs）与化疗的序贯联合可延长具有 EGFR 突变的晚期非小细胞肺癌（NSCLC）患者的无进展生存期（PFS）和/或总生存期（OS）。本研究旨在评估最佳联合顺序，并探讨其可能的机制。
分别用多西紫杉醇/吉非替尼单药或不同联合方案处理具有突变型和野生型 EGFR 的肺腺癌 PC-9 细胞和 A549 细胞。采用 qPCR-HRM 技术检测 EGFR 和 K-ras 基因状态。用 MTT 法检测细胞增殖。用 Western blot 法检测 EGFR、ERK、Akt 和 IGF-1R 的表达和磷酸化。用流式细胞术观察细胞周期分布。
只有多西紫杉醇序贯吉非替尼（D→G）在两种细胞系中均诱导出显著的协同作用（组合指数<0.9）。相反，吉非替尼序贯多西紫杉醇（G→D）在两种细胞系中均产生拮抗作用（CI>1.1），而同时给药（D+G）在 PC-9 细胞中表现出相加-协同作用，在 A549 细胞中表现出拮抗-相加作用。机制研究表明，多西紫杉醇诱导的 EGFR 和 ERK 磷酸化被随后使用的吉非替尼抑制，但不是通过同时暴露于吉非替尼。吉非替尼抑制的 EGFR 和 ERK 磷酸化既不能通过同时给药也不能通过随后给药来逆转。D+G 增强了它们对 PC-9 细胞中 IGF-1R 磷酸化的抑制作用。
对于 EGFR 突变型 NSCLC 细胞的增殖抑制作用，细胞毒性药物继之以 EGFR-TKIs 可能是最佳联合方案，EGFR 和 ERK 的磷酸化可能有助于这种作用。
