Abstract LB-57: Multiplexed detection of phospho-ERK as a marker of H-Ras signaling, EGFR and cancer stem cell marker expression in breast cancer cells recovered from human blood

Breast cancer is the most frequent malignancy among women, causing 373,000 deaths per year worldwide. Recent studies indicate that breast cancer is initiated by breast cancer stem cells (BCSCs) which express CD44+/high CD24−/low surface markers. Mammospheres are non-adherent spherical cell clusters grown in selective culture conditions. Our experimental results using mammosphere cultures of different metastatic, human breast cancer cells such as MDA-MB-231 and MDA-MB-468, showed drastic enrichment of breast cancer stem-like cells with the phenotype of CD44+/high CD24−/low in flow cytometry analysis, with specific antibody conjugates. Using the FDA-approved Veridex CellSearch System, we successfully recovered Epcam+/cytokeratin+/CD45− breast cancer cells from 7.5 ml of human blood to which a known number of MDA-MB-468 or H-Ras transformed MCF10A cells had been added. When these recovered breast cancer cells were subjected to Q-Dot conjugated p-ERK antibody staining and quantification, signal intensity clearly demonstrated higher levels of nuclear p-ERK in MCF10A-Ras cells whereas immunostaining for surface EGFR expression was much stronger in MDA-MB-468 cells. We are modeling mammosphere-derived breast cancer cells enriched for stem cell markers by orthotopic implantation into mammary fat pads of nude mice, sampling blood from tumor-bearing animals to detect the presence of Circulating Tumor Cells (CTCs) using the Veridex CellSearch System and further for breast cancer specific multiplexed marker analysis. These stem cell marker enriched xenografted human breast tumors are facilitating our understanding of cellular signaling pathways active in CTCs, and also in understanding of the role of cancer stem cells in human breast cancer metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-57. doi:10.1158/1538-7445.AM2011-LB-57