Quantitative proteomic analysis identifies TUBB2A as a novel protein biomarker for prediction of distant metastatic breast cancer

Metastasis of breast cancer to other distant organs is fatal to patients. However, few studies have revealed biomarkers associated with distant metastatic breast cancer. Furthermore, the inability of current biomarkers such as HER2, ER and PR, in accurately differentiating between distant metastatic breast cancers from non-distant metastatic ones necessitates the development of novel biomarkers. An integrated proteomics approach that combines filter-aided sample preparation, tandem mass tag labeling (TMT), high pH fractionation, and high resolution MS was applied to acquire in-depth proteome data of distant metastatic breast cancer FFPE tissue. Bioinformatics analyses for gene ontology and signaling pathways using differentially expressed proteins (DEPs) were performed to investigate molecular characteristics of distant metastatic breast cancer. In addition, real-time polymerase chain reaction (RT-PCR) and invasion/migration assays were performed to validate the differential regulation and functional capability of biomarker candidates. A total of 9,459 and 8,760 proteins were identified from the pooled sample set and the individual sample set, respectively. Through our stringent criteria, TUBB2A was selected as a novel biomarker. The metastatic functions of the candidate were subsequently validated. Bioinformatics analysis using DEPs were able to characterize the overall molecular features of distant metastasis as well as investigate the differences across breast cancer subtypes. Our study is the first to explore the distant metastatic breast cancer proteome using FFPE tissue. The depth of our dataset enabled the discovery of novel biomarker and the investigation of proteomic characteristics of distant metastatic breast cancer. The distinct molecular features of breast cancer subtypes were also observed. Our proteomic data has important utility as a valuable resource for the research on distant metastatic breast cancer.