Abstract 2494: Adhesive signature technology for tumor initiating cell purification in cancer research

Abstract

Abstract In spite of major therapeutic advances, cancer relapse and low rates of patient response persist. This failure is due in part to a small subpopulation of tumor initiating cells (TICs) with stem cell like properties that are responsible for the growth of the tumor and the progression of metastasis. Currently, no efficient and reliable methods to isolate TICs for study exist. Our lab has developed a methodology to isolate cells based on their unique adhesion binding strength to a matrix. The novel technology (micro-Stem cell High- Efficiency Adhesion based Recovery [μSHEAR]) consists of a microfluidic device that applies varying degrees of detachment shear forces on cells. Using this device, human pluripotent stem cells and their progeny have been isolated with high reproducibility, yield (>97%), purity (95-99%), and survival (>95%) rates (Singh et al, Nature Methods 2013). The process is fast (10 min), label free, and scalable. The objective of this research is to isolate the rare TICs from the general cancer cell population by exploiting differences in adhesion strength. To study these differences, we measured the adhesion strength (adhesive signature) of a panel of breast cancer cell lines to fibronectin using a hydrodynamic assay. Based on this screen, we selected the MDA-MB- 231, MDA-MB-453, and MCF7 cell lines for purification of TICs using the uSHEAR technology. Briefly, microfluidic channels were sterilized and coated with fibronectin. MDA-MB- 231, MDA-MB-453 or MCF7 breast cancer cells were enzymatically disassociated, pipetted into the inlet reservoir, and cultured in the device for 24h before detachment experiments. Cells were exposed to different shear forces to selectively detach cell sub-populations. Recovered cell/colonies were counted, seeded into a mammosphere formation assay (MFA), and analyzed after 10 days The ability to form mammospheres is characteristic of TICs. After uSHEAR mediated separation of breast cancer cells into three fractions, the strongest adhering fraction consistently produced larger and more mammospheres. Furthermore, as the selection adhesive force for the adhered fraction was increased, greater increases in mammosphere number and size were observed. After 10 days, the mammospheres were disassociated and the number of cells quantified. A 5-15 fold increase in the final number of cells was observed in the strongly adherent fractions of cells, compared to 0.8-1.7 fold increase in the unsorted controls. Our results show that cancer cells with a higher mammosphere formation potential, a hallmark of characteristic TICs, have a higher adhesion strength. Furthermore, these cells can be enriched via adhesion based separation giving rise to more and larger mammospheres than their less adherent counterparts. Our results suggest that TICs could be separated based on adhesive forces. Future studies are aimed at characterizing the isolated cells as TICs and testing the isolation strategy in primary tumors. Citation Format: Efrain A. Cermeno, Austin P. Veith, Susan N. Thomas, Andres J. Garcia. Adhesive signature technology for tumor initiating cell purification in cancer research. [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 2494.