Atomic Force Microscopy of Triple Negative Breast Cancer Cells: A Predictive Value of Mechanical Phenotype

Abstract

We applied Quantitative Nanomechanical Atomic Force Microscopy (QNM-AFM) to test a response of physical phenotype of triple negative breast cancer cells to treatment with proteasome inhibitors. Genetic screens identified TNBCs as addicted to the activity of the ubiquitin-proteasome pathway, the major intracellular venue of regulated protein degradation, served by the essential protease, the proteasome. We investigated two types of proteasome inhibitors: the drug bortezomib (Velcade®) targeting active sites of the enzyme and a novel allosteric inhibitor B1 interfering with stability of the 26S proteasome. Mechanical phenotype constitutes a sensitive indicator of physiological status of cells. The most explored mechanical parameters are cell elasticity (“softness”) and surface adhesiveness (“stickiness”). It has been established that cancerous transformation makes cells softer and less sticky. These differences are attributed to remodeling of the cytoskeleton and altered expression of membrane proteins. Therefore, the physical phenotype should have a predictive value as the early indicator of the cells’ response to a drug treatment and disease stage. Indeed, it has been shown that exposure of cancer cells to anticancer drugs at least partially reverses their mechanical phenotype resembling healthy cells. Here we found that subjecting breast cancer cells to low doses of bortezomib increased their stiffness and adhesiveness about two times. Moreover, similar treatment with low doses of B1 induced almost a threefold increase of stiffness and a twofold increase in cell adhesion. Surprisingly, much higher doses of the drugs were required to inflict detectable changes in cells viability or morphology. The results point at the extraordinary sensitivity of the mechanical phenotype to detect cells’ response to anticancer drugs. We are exploring the potential predictive value of AFM-based cell surface studies in a search for effective drugs or drug combinations.