Objective: To explore the intrinsic connection between activation of classical nuclear factor-κB (NF-κB) pathway and gefitinib resistance in human lung adenocarcinoma H1650 cells.
Methods: Human lung adenocarcinoma H1650 cells were exposed to gefitinib continuously for 60 days to obtain resistant H1650 cells. The expressions of P-IκBα, P-p50 and P-p65 in the cytoplasm or nuclei were detected using Western blotting in human lung adenocarcinoma HCC827 cells, parental H1650 cells and gefitinib-resistant H1650 cells. The effects of gefitinib alone or in combination with PDTC on the survival rate and expressions of NF-κB P-p50 and P-p65 were compared among the 3 cell lines.
Results: Gefitinib-resistant H1650 cells showed increased cytoplasmic and nuclear P-IκBα expressions. The expressions of P-p50 and P-p65 differed significantly among the 3 cell line, decreasing in the order of resistant H1650 cells, parental H1650 cells, and gefitinib sensitive HCC827 cell lines (P<0.05 or 0.01). Treatment with gefitinib alone resulted in a significantly lower cell inhibition rate in resistant H1650 cells than in the parental H1650 cells (P<0.05) and HCC827 cells (P<0.01). The resistant H1650 cells had a significantly higher expression of P-p50 and P-p65 than other two cell lines (P<0.05). In both the resistant and parental H1650 cells, gefitinib significantly lowered P-p50 and P-p65 expressions (P<0.05 or 0.01), and the combined treatment with gefitinib and PDTC significantly decreased the cell survival rate and further lowered the cytoplasmic and nuclear expressions of P-p50 and P-p65 (P<0.01 or 0.01).
Conclusion: The activation of classical NF-κB pathway is a key factor contributing to transformation of the parental H1650 cells into gefitinib-resistant cells. Gefitinib combined with PDTC can inhibit P-IκBα production and NF-κB P-p50 and P-p65 activation to suppress the survival of residual H1650 cells and the generation of gefitinib-resistant cells.
目的: 探究NF-κB经典通路激活与人肺腺癌H1650细胞吉非替尼耐药的内在联系。
方法: 选择肺腺癌HCC827和H1650亲代细胞株, 吉非替尼连续处理60 d, 检测细胞株生长情况和各细胞株595nm吸光度, 确定各细胞株细胞存活率和H1650亲代细胞被诱导成H1650耐药细胞株。Western blot检测各细胞株P-IκBα、NF-κB P-p50和P-p65表达量, Image J灰度仪计算出胞浆和胞核P-p50和P-p65与内参蛋白灰度比值, 比较吉非替尼和吉非替尼加吡咯烷二硫代氨基甲酸盐(PDTC)联合用药状态下, 各细胞株细胞存活率和P-IκB、P-p50和P-p65的表达。
结果: 在无吉非替尼和吉非替尼加PDTC药物干预下, H1650耐药细胞PIκBα表达增加, 胞浆和胞核P-p50和P-p65表达, H1650耐药细胞株>H1650亲本细胞株>吉非替尼敏感HCC827细胞株(P < 0.05, P < 0.01);单独使用吉非替尼干预, H1650耐药细胞株细胞抑制率较H1650亲本细胞株和HCC827细胞株降低(P < 0.05, P < 0.01), H1650耐药细胞株P-p50和P-p65表达较其他两类细胞株升高(P < 0.05);与无吉非替尼干预比较, H1650亲代和耐药细胞P-p50和P-p65均有显著降低(P < 0.05, P < 0.01);吉非替尼联合PDTC干预, 显示H1650亲代和耐药细胞存活率、胞浆和胞核Pp50和P-p65表达均较吉非替尼组降低(P < 0.01, P < 0.01)。
结论: NF-κB经典传导通路激活是H1650亲代细胞残存和转化为吉非替尼耐药细胞的主要因素之一, 吉非替尼联合PDTC用药可抑制P-IκBα生成和P-p50、P-p65的激活, 从而降低H1650残存细胞生存率和耐药细胞的产生。