iTRAQ-Based Proteomic Profiling of Breast Cancer Cell Response to Doxorubicin and TRAIL

Breast cancer is a molecularly heterogeneous disease,
and predicting
response to chemotherapy remains a major clinical challenge. To minimize
adverse side-effects or cumulative toxicity in patients unlikely to
benefit from treatment, biomarkers indicating treatment efficacy are
critically needed. iTRAQ labeling coupled with multidimensional LC–MS/MS
of the enriched mitochondria and endoplasmic reticulum fraction, key
organelles regulating apoptosis, has led to the discovery of several
differentially abundant proteins in breast cancer cells treated with
the chemotherapeutic agent doxorubicin followed by the death receptor
ligand, TRAIL, among 571 and 801 unique proteins identified in ZR-75-1
and MDA-MB-231 breast cancer cell lines, respectively. The differentially
abundant proteins represent diverse biological processes associated
with cellular assembly and organization, molecular transport, oxidative
stress, cell motility, cell death, and cancer. Despite many differences
in molecular phenotype between the two breast cancer cell lines, a
comparison of their subproteomes following drug treatment revealed
three proteins displaying common regulation: PPIB, AHNAK, and SLC1A5.
Changes in these proteins, detected by iTRAQ, were confirmed by immunofluorescence,
visualized by confocal microscopy. These novel potential biomarkers
may have clinical utility for assessing response to cancer treatment
and may provide insight into new therapeutic targets for breast cancer.

乳腺癌是一种分子异质性疾病，预测患者对化疗的应答反应仍是一项重大临床挑战。为了在不太可能从治疗中获益的患者群体中最大限度降低不良反应或累积毒性，亟需可指示治疗疗效的生物标志物。本研究采用结合液相色谱-串联质谱（LC-MS/MS）的iTRAQ标记技术，对富集的线粒体与内质网组分——这两类调控细胞凋亡的关键细胞器——进行分析。结果在分别来自ZR-75-1和MDA-MB-231乳腺癌细胞系的571种和801种独特蛋白中，发现经化疗药物多柔比星（doxorubicin）及死亡受体配体肿瘤坏死因子相关凋亡诱导配体（TRAIL）处理的乳腺癌细胞内存在多种差异丰度蛋白。这些差异丰度蛋白涉及多种生物学过程，包括细胞组装与组织、分子转运、氧化应激、细胞运动、细胞死亡及癌症发生相关通路。尽管这两种乳腺癌细胞系的分子表型存在诸多差异，但对其经药物处理后的亚蛋白质组进行比较后发现，三种蛋白呈现出一致的表达调控模式：PPIB、AHNAK及SLC1A5。通过iTRAQ标记检测到的这些蛋白表达变化，经免疫荧光染色结合共聚焦显微镜成像得到了验证。这些具有潜在临床应用价值的新型生物标志物，不仅可用于评估癌症治疗应答反应，还可为乳腺癌新型治疗靶点的研发提供全新思路。