Cell–Gel Mechanical Interactions as an Approach to Rapidly and Quantitatively Reveal Invasive Subpopulations of Metastatic Cancer Cells

Abstract

We present a novel mechanobiology-based invasiveness assay to rapidly and quantitatively evaluate the mechanical invasiveness of metastatic cancer cells and identify invasive subpopulations, without need for chemoattractants and independent of serum content. A commonly accepted assay to determine metastatic potential in vitro is the Boyden chamber assay, where the percentage of serum-starved cells that can long-term transmigrate/invade through subcell size membrane pores is quantified; those experiments typically take 2–3 days and require serum-starvation. To squeeze through the small pores, the invasive cells must be pliable, yet they are also able to force their way through flexible microenvironments. We have previously shown that metastatic breast cancer cells will deform and indent soft, impenetrable, elastic gels within 2 h of seeding, without requiring serum starvation. Specifically, in cell lines with higher metastatic potential, a larger percentage of cells will indent gels and typically also to deeper depths. Thus, we are able to rapidly reveal mechanically invasive subpopulations, which are likely those that lead to high metastatic potential. By comparing the Boyden chamber and gel mechanobiology assays, we show that the gel-indenting cell subpopulations are part of the group that successfully transmigrates through the Boyden chamber membrane (8 μm pores). Thus, we are able to rapidly (within 2 h of seeding and using the standard cell media), provide a quantitative measure of the mechanical invasiveness of cancer cells, which is correlated to the metastatic potential but is an independent parameter; we evaluate numbers of indenting cells and their indentation depth. Moreover, the mechanical invasiveness assay allows focus on specific (invasive or noninvasive) cells within the sample to identify specific surface markers, determine invasive mechanisms, and evaluate effects of applied drugs and treatments on the different subpopulations.