Metastasis-related Plasma Membrane Proteins of Human Breast Cancer Cells Identified by Comparative Quantitative Mass Spectrometry

Abstract

The spread of cancer cells from a primary tumor to form metastasis at distant sites is a complex multistep process. The cancer cell proteins and plasma membrane proteins in particular involved in this process are poorly defined, and a study of the very early events of the metastatic process using clinical samples or in vitro assays is not feasible. We have used a unique model system consisting of two isogenic human breast cancer cell lines that are equally tumorigenic in mice; but although one gives rise to metastasis, the other disseminates single cells that remain dormant at distant organs. Membrane purification and comparative quantitative LC-MS/MS proteomics identified 13 membrane proteins that were expressed at higher levels and three that were underexpressed in the metastatic compared with the non-metastatic cell line from a total of 1919 identified protein entries. Among the proteins were ecto-5-nucleotidase (CD73), NDRG1, integrin ␤1, CD44, CD74, and major histocompatibility complex class II proteins. The altered expression levels of proteins identified by LC-MS/MS were validated using flow cytometry, Western blotting, and immunocyto-and immunohistochemistry. Analysis of clinical breast cancer biopsies demonstrated a significant correlation between high ecto-5-nucleotidase and integrin ␤1 expression and poor outcome, measured as tumor spread or distant recurrence within a 10-year follow-up. Further the tissue analysis suggested that NDRG1, HLA-DR␣, HLA-DR␤, and CD74 were associated with the ER

Blockade of A 2A receptors potently suppresses the metastasis of CD73 + tumors

“…Our data have significant therapeutic potential in the treatment of tumor metastasis given that (i) CD73 expression was shown to be a predictor of metastasis in breast cancer patients (13) antagonist CVT-6883 has undergone a phase I trial (25). All three compounds were safe and well-tolerated, indicating that a pathway to the clinic for drugs of this class maybe relatively straightforward.…”

“…All three compounds were safe and well-tolerated, indicating that a pathway to the clinic for drugs of this class maybe relatively straightforward. Because a link has been shown between CD73 expression and metastasis in breast cancer patients (13), further investigation into the therapeutic potential of A 2A /A 2B blockade is warranted.…”

“…Understanding the mechanism by which CD73 increases tumor metastasis is vital to developing new strategies that target metastasis because there are no anti-CD73 mAbs currently in clinical development. The clinical relevance of this pathway is underlined by the correlation between CD73 expression and an increased risk of metastasis in breast cancer (13,14). Thus, the CD73 pathway has been proposed as a novel target for immunotherapy (15).…”

Biomarkers for EMT and MET in breast cancer: An update

“…It plays an important role in the metastasis of breast cancer (30). Some studies have shown an upregulation of CD44 in a metastatic cell line as compared with a non-metastatic cell line (31). There is a shift in CD44 expression from the variant isoform (CD44v) to the standard isoform (CD44s) during EMT.…”

Cancer: A proteomic disease

“…These results indicate that multiple subcellular locations exist for proteins in processes related to breast cancer. Leth-Larsen et al [10,11] identified 13 membrane proteins that were over-expressed and three that were under-expressed in a metastatic versus non-metastatic cell line from a total of 1919 protein entries. High ecto-5′-nucleotidase and integrin β1 expression correlated with tumor spread or distant recurrence, and poor outcome within a 10-year follow-up [10].…”