Wang Yi, Cao Jiang, Zeng Su
Cancer Institute, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, China.
Shi Yan Sheng Wu Xue Bao. 2003 Oct;36(5):342-6.
In an attempt to study the whole protein expression alterations of tumer cells after becoming multidrug-resistant, which may provide useful information on new drug target identification, an adriamycin-resistant variant of the human leukemia cell line K562 (K562/ADR) was developed in vitro by continuous exposure to adrimycin. MTT assay was used to determine IC50 of K562/ADR cells to adriamycin (ADR), cisplatin (DDP), 5-fluorouracil (5-FU) and vincristin (VCR). The total proteins of K562 and K562/ADR were separated by two-dimensional gel electrophoresis and visualized by silver staining. Proteins with significant expression alterations were selected and their peptide mass fingerprints (PMFs) were obtained by matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS). The PMFs were used to search NCBInr database by AutoMS-Fit software. The results showed that K562/ADR cell demonstrated cross-resistance to other antineoplastic drugs. The IC50 of K562/ADR cells to ADR, DDP, 5-FU, VDR were much higher than those of K562. The proteins differentially expressed in the two cell lines were identified as cell cycle-related proteins, zinc finger protein 165, etc. These proteins are involved in cell cycling and transcription regulation, whose expression alterations may contribute to the multidrug resistant phenotype of K562/ADR cells.
为了研究肿瘤细胞产生多药耐药后全蛋白表达的变化，这可能为新药靶点的鉴定提供有用信息，通过持续暴露于阿霉素在体外建立了人白血病细胞系K562的阿霉素耐药变体（K562/ADR）。采用MTT法测定K562/ADR细胞对阿霉素（ADR）、顺铂（DDP）、5-氟尿嘧啶（5-FU）和长春新碱（VCR）的半数抑制浓度（IC50）。通过二维凝胶电泳分离K562和K562/ADR的总蛋白，并用银染法进行可视化。选择表达有显著变化的蛋白质，通过基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF-MS）获得其肽质量指纹图谱（PMF）。利用AutoMS-Fit软件将PMF在NCBInr数据库中进行检索。结果显示，K562/ADR细胞对其他抗肿瘤药物表现出交叉耐药性。K562/ADR细胞对ADR、DDP、5-FU、VDR的IC50远高于K562细胞。在这两种细胞系中差异表达的蛋白质被鉴定为细胞周期相关蛋白、锌指蛋白165等。这些蛋白质参与细胞周期和转录调控，其表达变化可能导致K562/ADR细胞的多药耐药表型。
