稳定表达FIP1L1-PDGFRA及其突变体细胞株的构建和耐药性评价  

Construction and drug resistance evaluation of cell strains which stably express FIP1L1-PDGFRA protein and its mutants

作　　者：刘婧雯 宋昊昕 朱蕾[1] LIU Jingwen;SONG Haoxin;ZHU Lei(Department of Pharmacology,Institute of Basic Medical Sciences CAMS,School of Basic Medicine PUMC Medical Epigenetics Research Center CAMS,Beijing 100005,China)

机构地区：[1]中国医学科学院基础医学研究所、北京协和医学院基础学院、中国医学科学院医学表观遗传研究中心药理学系,北京100005

出　　处：《基础医学与临床》2024年第1期63-68,共6页Basic and Clinical Medicine

基　　金：中国医学科学院医学与健康科技创新工程项目(2021-I2M-1-005);中央高水平医院临床科研业务费(2022-PUMCH-C-025)。

摘　　要：目的构建能稳定表达FIP1L1-PDGFRA融合基因(F/P)及其T674I和D842V突变体的小鼠原B细胞株BaF3-F/P、BaF3-F/P-T674I和BaF3-F/P-D842V,并通过考察它们对酪氨酸激酶抑制剂(TKIs)的响应以评价其活性。方法用慢病毒感染技术将目的基因导入BaF3细胞;RT-qPCR检测mRNA表达;CCK-8法检测TKIs对稳转细胞株增殖的抑制作用。结果构建的BaF3-F/P、BaF3-F/P-T674I和BaF3-F/P-D842V细胞株均能转录FIP1L1和PDGFRA mRNA,并表现出不依赖IL-3增殖的恶性表型特征以及对相应TKIs的敏感性。结论成功构建了能稳定表达F/P及其T674I、D842V突变体的小鼠原B细胞株,为开发以此为靶点的化合物提供良好的细胞模型。Objective To construct mouse BaF3-FIP1L1-PDGFRA(F/P),BaF3-F/P-T674I and BAF3-F/P-D842V pre-B cell strains which stably express F/P fusion protein and its T674I and D842V mutants in order to evaluate their activity by checking their responses to tyrosine kinase inhibitors(TKIs).Methods Lentivirus infected technique was used to transfect the target gene into BaF3 cells.RT-qPCR was used to detect mRNA expression,and CCK-8 method was used to detect the inhibitory effect of TKIs on the proliferation of stable cell strains.Results The constructed BaF3-F/P,BaF3-F/P-T674I and BAF3-F/P-D842V cell strains all transcripted FIP1L1 and PDGFRA mRNA.They exhibited malignant phenotypic characteristics of proliferation independent of IL-3 and sensitivity to corresponding TKIs.Conclusions The pre-B-cell strains stably expressing F/P and its T674I and D842V mutants are successfully constructed,which provide a good cell model for the development of compounds targeting at those molecules.

关 键 词：FIP1L1-PDGFRA融合基因 基因突变 慢病毒载体 BaF3细胞 慢性嗜酸粒细胞白血病 

分 类 号：R967[医药卫生—药理学]