非小细胞肺癌ALK-TKIs耐药株的构建及耐药后序贯治疗的效果探究  

Establishment of ALK-TKIs resistant cells in non-small cell lung cancer and the efficacy of sequential therapy following ALK-TKIs resistance

作　　者：王鑫烨 李浩 李梅[1] 刘子威[1,2] 吕丽婷 WANG Xinye;LI Hao;LI Mei;LIU Ziwei;LYU Liting(Department of Tumor Chemotherapy,Affiliated Hospital of Nantong University,Jiangsu 226001;Medical School of Nantong University)

机构地区：[1]南通大学附属医院肿瘤化疗科,江苏226001 [2]南通大学医学院

出　　处：《南通大学学报(医学版)》2025年第1期7-13,共7页Journal of Nantong University(Medical sciences)

基　　金：国家自然科学基金青年基金资助项目(81602010);南通市卫生健康委员会科研课题(MS2024017)。

摘　　要：目的:探索间变性淋巴瘤激酶-酪氨酸激酶抑制剂(anaplastic lymphoma kinase-tyrosine kinase inhibitors,ALK-TKIs)耐药后序贯其他ALK-TKIs治疗的效果。方法:首先,收集12例序贯使用两种ALK-TKIs的非小细胞肺癌(non-small cell lung cancer,NSCLC)患者信息,分析克唑替尼、阿来替尼及洛拉替尼在真实世界中序贯使用的疗效。其次,构建H3122-克唑替尼耐药株(H3122-CR)、阿来替尼耐药株(H3122-AR)及洛拉替尼耐药株(H3122-LR),通过MTT及细胞克隆形成实验证明耐药株构建成功。最后,为探索ALK-TKIs耐药后序贯后代TKIs或一代、二代TKIs再挑战是否发挥抗肿瘤作用,通过MTT及克隆形成实验,在H3122-CR耐药株中使用阿来替尼和洛拉替尼,在H3122-AR耐药株中使用克唑替尼及洛拉替尼,或在H3122-LR耐药株中使用克唑替尼及阿来替尼,分别检测各代ALK-TKIs抗肿瘤细胞活性的作用。结果:真实世界中,一代ALK-TKI耐药后序贯使用二代或三代ALK-TKIs,或二代ALK-TKIs耐药后序贯使用三代ALK-TKI均可有一定临床获益。MTT实验表明,H3122细胞的克唑替尼IC_(50)值为341.7 nmol/L,H3122-CR耐药株的克唑替尼IC_(50)值为2.766μmol/L;H3122细胞的阿来替尼IC_(50)值为131.5 nmol/L,H3122-AR耐药株的阿来替尼IC_(50)值为1.07μmol/L;H3122细胞的洛拉替尼IC_(50)值为42.85 nmol/L,H3122-LR耐药株的洛拉替尼IC_(50)值为3.294μmol/L。细胞克隆形成实验提示,200 nmol/L克唑替尼、阿来替尼、洛拉替尼均不足以抑制H3122-CR、H3122-AR、H3122-LR耐药株的克隆形成能力,但完全抑制了亲本株的克隆形成能力,提示耐药株构建成功。通过MTT实验及克隆形成实验,在H3122-CR耐药株中,阿来替尼及洛拉替尼均可抑制肿瘤细胞的活性;在H3122-AR耐药株中,克唑替尼抗肿瘤细胞活性甚微,而序贯洛拉替尼可以发挥抗肿瘤作用。在H3122-LR耐药株中,克唑替尼及阿来替尼有些许抗肿瘤作用。结论:克唑替尼耐药后,序贯阿来替Objective:To explore the efficacy of sequential therapy with other ALK-TKIs.Methods:Firstly,twelve non-small cell lung cancer(NSCLC)patients who used two ALK-TKIs were enrolled in the study,to analyze the real-world efficacy of sequential use of crizotinib,alectinib,or lorlatinib.Then,H3122-crizotinib-resistant(H3122-CR),alectinib-resistant(H3122-AR),and lorlatinib-resistant(H3122-LR)cells were established.And MTT and cell clone formation experiments were performed to verify the TKIs resistance.To further investigate whether sequential administration of TKIs exerts antitumor effects after ALK-TKIs resistance,MTT and clone formation experiments were conducted.Specifically,H3122-CR cells were treated with alectinib and lorlatinib,H3122-AR cells were treated with crizotinib and lorlatinib,H3122-LR cells were treated with crizotinib and alectinib,to evaluate the antitumor activity of various generations of ALK-TKIs.Results:In the real-world study,sequential use of second-or third-generation ALK-TKIs with resistance to first-generation ALK-TKI,or third-generation ALK-TKI with resistance to second-generation ALK-TKIs,demonstrated clinical benefit.MTT experiments showed that the IC_(50)value of crizotinib in H3122 cells was 341.7 nmol/L,which was 2.766μmol/L in H3122-CR cells.The IC_(50)value of alectinib in H3122 cells was 131.5 nmol/L,and 1.07μmol/L in H3122-AR cells.The IC_(50)value of lorlatinib in H3122 cells was 42.85 nmol/L,and 3.294μmol/L in H3122-LR cells.Cell clone formation experiments indicated that 200 nmol/L crizotinib,alectinib,or lorlatinib were insufficient to inhibit the clone formation ability of H3122-CR,H3122-AR,and H3122-LR resistant cells,but completely inhibited the clone formation ability of the parental cells,confirming successful construction of the resistant cells.MTT and clone formation experiments demonstrated that both alectinib and lorlatinib could inhibit the activity of H3122-CR cells.In H3122-AR cells,sequential use of crizotinib was ineffective,but sequential use of lorlatinib exhi

关 键 词：非小细胞肺癌 间变性淋巴瘤激酶 克唑替尼 阿来替尼 洛拉替尼 靶向治疗耐药 

分 类 号：R734.2[医药卫生—肿瘤]