Abstract 249: Breast cancer cell vesiculation is driven by calpain: implications in cancer therapy

Abstract

Abstract Microparticles (MPs) are small membrane vesicles (0.1 to 1 μm in diameter) released from the plasma membranes of most cell types, (malignant and non-malignant) upon cellular activation or during apoptosis. We discovered that MPs provide a “non-genetic” mechanism for the acquisition of multidrug resistance and increased metastatic capacity in tumour cell populations, whereby they serve as vectors in the intercellular transfer of functional cancer proteins and nucleic acids. By this mechanism, MPs effectively confer the transfer, dissemination and dominance of within cancer cell populations within a matter of hours. The aim of this study was to define the pathways governing membrane vesicle biogenesis in malignant and non-malignant cells with the aim of identifying cancer specific mediators, which could serve as novel therapeutic targets in preventing microvesicle induced deleterious traits in cancer. We preformed a comparative analysis using the non-malignant human brain endothelial cell line (HBEC-D3) cells and drug sensitive and resistant human breast adenocarcinoma cells (MCF-7) and (MCF-7/DX) respectively. Cell surface topography and vesiculation pits were visualized at rest, and in the +/- calcium ionophore, A23187 and the Calpain inhibitor ALLM using contact mode Atomic Force Microscopy (Nanowizard, JPK Instruments, Germany) on fixed cells. We observed that at rest, cells from both malignant cell lines were high vesiculators relative to the HBEC-D3 cells. In the presence of ALLM we observed a direct inhibition of vesiculation in the malignant cells whilst vesiculation was enhanced in the non-malignant cells. Upon the release of calcium from intracellular stores using A23817, we observe an increase in vesiculation across all cell types, however to a greater extent in the HBEC-D3 cells. Interestingly, only upon the release of intracellular calcium in HBEC-D3 cells do we see an inhibition of vesiculation by ALLM. These results clearly demonstrate an alternative pathway governing vesiculation at rest for the non-malignant HBEC-D3 cells. Whereas contrary to this, vesiculation at rest in malignant cells appears to be calpain dependent. Future work aims to validate these findings and examine the presence of distinct pathways of vesiculation in malignant cells relative to non-malignant cells. These results have important implications in defining novel strategies to selectively targeting malignant cells and for the circumvention of deleterious traits acquired through intercellular exchange of extracellular vesicles. Citation Format: Jack Taylor, Ritu Jaiswal, Mary Bebawy. Breast cancer cell vesiculation is driven by calpain: implications in cancer therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 249.