Department of Biomedical Imaging and Image-guided Therapy , Medical University of Vienna , Währinger Gürtel 18-20 , 1090 Vienna , Austria.
Comprehensive Cancer Center of the Medical University of Vienna , Spitalgasse 23 , 1090 Vienna , Austria.
Anal Chem. 2018 Nov 20;90(22):13178-13182. doi: 10.1021/acs.analchem.8b03756. Epub 2018 Nov 1.
Cancer cells communicate with the whole organism via extracellular vesicles (EVs), which propagate molecular information in support of the malignant phenotype. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was employed for protein profiling of EVs derived from CCL-228 as the primary colon tumor, the lymph node metastasis CCL-227, and subclones resistant to 5, 25, and 125 μM 5-fluorouracil (FU). EVs were harvested from cell culture supernatant by ultracentrifugation to serve as a model for circulating cancer cell-derived biomarker carriers from body fluids (i.e., liquid biopsy). Protein mass spectra were recorded using standard MALDI matrixes (e.g., CHCA, sinapinic acid) in the range m/ z 2000-20000 on different MALDI-TOF-MS systems and subjected to multivariate data analysis . By using hierarchical clustering, PCA and PLS-DA, discriminatory protein patterns of the EVs from the different cell populations were obtained. Peaks in the range m/ z 2000-6500 and m/ z 5500-15500 were found to be unique to EVs and the cells, respectively. This clearly demonstrates the differential expression of proteins in EVs as the result of an increasing chemoresistance of their parent cells. The sensitivity of the MALDI-MS based assay was in the low μg/mL (≈1.2-5 × 10 particles/mL) range. Consequently, our MALDI-MS protein profiling approach shows the potential to serve as novel tool for minimally invasive cancer diagnostics and chemotherapy monitoring in the future, e.g., for early detection of therapy resistance without biopsy.
癌细胞通过细胞外囊泡(EVs)与整个机体进行通讯,这些囊泡传播分子信息以支持恶性表型。基质辅助激光解吸/电离飞行时间质谱(MALDI-TOF-MS)被用于分析源自 CCL-228(原发性结肠肿瘤)、CCL-227(淋巴结转移)以及对 5、25 和 125 μM 5-氟尿嘧啶(FU)耐药的亚克隆的 EVs 的蛋白质谱。通过超速离心从细胞培养上清液中收获 EVs,作为来自体液(即液体活检)的循环癌细胞衍生生物标志物载体的模型。使用标准 MALDI 基质(例如 CHCA、sinapinic 酸)在不同的 MALDI-TOF-MS 系统上记录蛋白质质谱,范围为 m/z 2000-20000,并进行多元数据分析。通过使用层次聚类、PCA 和 PLS-DA,获得了来自不同细胞群体的 EVs 的有区别的蛋白质图谱。在 m/z 2000-6500 和 m/z 5500-15500 范围内的峰分别被发现是 EVs 和细胞所特有的。这清楚地表明,EVs 中蛋白质的差异表达是其亲本细胞化学抗性增加的结果。MALDI-MS 测定的灵敏度在低 μg/mL(≈1.2-5×10 个颗粒/mL)范围内。因此,我们的 MALDI-MS 蛋白质谱分析方法显示出作为未来微创癌症诊断和化疗监测的新型工具的潜力,例如,在没有活检的情况下早期检测治疗耐药性。