靶向p21蛋白激活激酶2逆转非小细胞肺癌细胞吉非替尼耐药  

Targeting PAK2 Reverses Gefitinib Resistance in Non-small Cell Lung Cancer Cells

作　　者：黄渊锋 李盼 杨明强 刘浩 陈丹扬 HUANG Yuan-Feng;LI Pan;YANG Ming-Qiang;LIU Hao;CHEN Dan-Yang(Department of Thoracic Surgery,Affiliated Cancer Hospital of Guangzhou Medical University,Guangzhou 510095,China;Affiliated Cancer Hospital and Cancer Research Institute,Guangzhou Medical University,Guangzhou 510095,China)

机构地区：[1]广州医科大学附属肿瘤医院胸外科,广州510095 [2]广州医科大学附属肿瘤医院肿瘤研究所,广州510095

出　　处：《中国生物化学与分子生物学报》2021年第8期1078-1084,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：广东省医学科研基金项目(No.B2020165)资助。

摘　　要：靶向突变的表皮生长因子受体(epidermal growth factor receptor,EGFR)酪氨酸激酶抑制剂(tyrosine kinase inhibitor,TKI)在非小细胞肺癌(non-small cell lung cancer,NSCLC)治疗中显示出越来越重要的作用。然而,不可避免的耐药性的出现极大限制了TKI的临床治疗效果。p21蛋白激活激酶2(p21-activated protein kinase 2,PAK2)是丝/苏氨酸激酶家族的重要成员,在肿瘤发生与发展中发挥重要的作用。本研究拟探讨靶向抑制PAK2逆转非小细胞肺癌吉非替尼耐药的作用及可能机制。首先Western印迹检测发现,相比非小细胞肺癌吉非替尼敏感细胞HCC827,耐药细胞HCC827/GR中PAK2的蛋白质磷酸化水平显著上调(P<0.01),而PAK2总蛋白质水平未见明显变化。采用PAK2抑制剂FRAX597和G5555诱导HCC827/GR细胞,MTS及克隆形成结果显示,抑制剂FRAX597和G5555均能显著增加HCC827/GR细胞对吉非替尼的敏感性(P<0.01)。流式细胞术分析发现,抑制剂FRAX597处理诱导HCC827/GR细胞发生G2/M期阻滞,并增加吉非替尼诱导的HCC827/GR细胞的凋亡以及切割胱天蛋白酶3(cleaved caspase 3)表达(P<0.01)。最后,Western印迹结果显示,抑制剂FRAX597处理能够抑制HCC827/GR细胞BCL-2和CDK4蛋白质表达(P<0.05),上调BAX、p21和p27蛋白质表达(P<0.05)。综上所述,PAK2活化与非小细胞肺癌吉非替尼耐药性密切相关;靶向抑制PAK2活化能够诱导细胞周期阻滞及凋亡,从而提高非小细胞肺癌细胞对吉非替尼的敏感性。EGFR tyrosine kinase inhibitor(EGFR-TKI)-targeted therapy has been playing an important role in the treatment of non-small cell lung cancer(NSCLC).However,unavoidable therapeutic resistance significantly limits the clinical efficacy of TKI.As an important member of the PAK family of serine/threonine kinases,p21-activated protein kinase 2(PAK2)plays a critical role in tumorigenesis and tumor development.The aim of this work is to investigate the effect and molecular mechanism of inhibition of PAK2 on the reversal of gefitinib resistance in NSCLC.Firstly,Western blotting was used to detect the expression and phosphorylation level of PAK2 in gefitinib-resistant HCC827/GR cells.The results showed that the phosphorylation level of PAK2 was significantly increased in HCC827/GR cells compared with HCC827 cells(P<0.01),while the total protein level of PAK2 did not change.Then,HCC827/GR cells were treated with PAK2 inhibitors FRAX597 or G5555.The MTS assay and clone formation assay results showed that FRAX597 or G5555 significantly increased the sensitivity of HCC827/GR cells to gefitinib(P<0.01).Flow cytometry analysis showed that treatment of FRAX597 could induce cell cycle arrest in G2/M phase of HCC827/GR cells,and increased gefitinib-induced apoptosis and cleaved Caspase-3 expression(P<0.01).Western blotting results showed that treatment of FRAX597 significantly inhibited the expression of BCL-2 and CDK4(P<0.05),but increased the expression of BAX,p21 and p27(P<0.05).Taken together,these results indicated that activation of PAK2 is closely related to NSCLC with gefitinib resistance.Inhibition of PAK2 could induce cell cycle arrest and cell apoptosis,which increased the sensitivity of NSCLC cells to gefitinib.

关 键 词：非小细胞肺癌 吉非替尼耐药 p21蛋白激活激酶2 PAK2抑制剂FRAX597 

分 类 号：R734.2[医药卫生—肿瘤]