Abstract 4740: Real-time impedance analysis of triple negative breast cancer cells from African Americans demonstrates oncogenic mutant p53 (mtp53) is a promoter of cellular deformability

Abstract

Abstract The TP53 gene is mutated in 80% of basal-like triple negative breast cancers (TNBC) that are more prevalent in African American (AA) populations. Despite the increased interest in studying mtp53, its oncogenic mechanism of action and direct targets remain unclear. Based on our hypothesis mtp53 can increase cell deformability essential for metastatic cell spreading. To test this hypothesis we used two approaches. (1) We used custom designed interdigitated polysilicon microelectrodes to analyze cell deformability under hyposmotic pressure in a panel of breast cancer cell lines (AA-derived cancer cell lines MDA-MB-468 (mtp53/R273H), HCC70 (mtp53R248W) and HCC1806 (p53 null) and non-AA cell lines MCF-7 (wt p53) and MDA-MB-231 (mtp53/R280K)); we also compared impedance following mtp53 knockdown in MDA-MB-468 (through shRNA-mediated inducible knockdown). (2) We optimized a protocol for analysis of deformability using the commercial xCELLigence system to achieve lower variability and stronger statistical confidence, possible through simultaneous analysis of a large set of samples (three 16-well plates) and a controlled CO2/temperature environment. The principle of the deformability measurement assay is based on using a microelectrode transmitting an electromagnetic field, and following the extent of interference created by cells seeded on top of the electrode (i.e. bio-impedance). To measure the cells capacity to deform, deionized water was applied onto the cancer cells attached to the electrode, causing them to swell and increase bio-impedance. While cancer cells exhibit the ability to deform, normal cells retain their volume and eventually crack under hyposmotic pressure. By means of the impedance-based analysis we demonstrated that among all cell lines with mtp53, wt p53 or no p53, cell lines with mtp53 HCC70 and MB-MDA-468 generated substantially higher impedance. Furthermore we found that mtp53 knockdown in the MDA-MB-468.shp53 cell line led to decreased deformability. The optimized impedance-based deformability assay using xCELLigence produced identical results. Taken together, the obtained results imply that oncogenic mtp53 possesses gain-of-function that promotes cellular deformability, potentially to a greater extent in AA-derived cell lines. The important clinical value of this study is development of a robust analytical approach that, based on mtp53 function, may potentially provide an effective detection method for metastatic breast cancer cells and a useful tool for validation of mtp53-targeted therapy. Notably, this is the first report of a new application of the xCELLigence system for analysis of cancer cell mechanical properties. Funding: BCRF and a RCMI Program grant, National Center for Research Resources (G12 RR003037) and the National Institute on Minority Health and Health Disparities (8 G12 MD007599) from the NIH. Citation Format: Nataly Shtraizent, Menglu Shi, Alla Polotskaya, Hiroshi Matsui, Jill Bargonetti. Real-time impedance analysis of triple negative breast cancer cells from African Americans demonstrates oncogenic mutant p53 (mtp53) is a promoter of cellular deformability. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4740. doi:10.1158/1538-7445.AM2014-4740