281019 Extracellular Matrix Stiffness and Drug Resistance in Carcinoma
Extracellular Matrix Stiffness and Drug Resistance in Carcinoma
Thuy V. Nguyen and Shelly R. Peyton
University of Massachusetts, Amherst, MA, USA
The in vivo tumor niche is comprised of both cells and extracellular matrix (ECM), which provides important physical and chemical cues, that regulate tumor cell growth, motility, and perhaps, the ability of cells to respond to drugs. We hypothesize that receptor tyrosine kinase (RTK) inhibitors, in particular, may show misleading efficacy when tested on cells cultured on tissue culture plastic, due to the unnatural microenvironment. In response, we are developing a novel high-throughput biomaterial system to study how carcinoma cells respond to RTK inhibitors in the presence of a physiologically relevant matrix cues.
During disease progression in tumor microenvironment, the ECM is remodeled by fibroblasts. Collagen IV and laminin are replaced by fibrillar collagen I, III, and fibronectin. This ECM turnover increases the total protein content as well as the local tissue modulus (from less than 1kPa to 20-100kPa or more). In order to capture these physical and chemical tissue changes, we are developing a novel, high-throughput, PEG-PC hydrogel biomaterial system. Briefly, poly(ethylene glycol) dimethacrylate (PEG) and the zwitterion 2-Methacryloyloxyethyl phosphorylcholine (PC) form an optically clear hydrogel via radical polymerization in a silane-treated glass 96-well plate. We can control the modulus of the hydrogels anywhere from one to hundreds of kPa by tuning the PEG crosslinker content. The PC zwitterion is extremely hydrophilic, which allows our PEG-PC gels to achieve lower moduli (we can form gels with only 0.5 wt% PEG crosslinker), and block non-specific protein interactions better than PEG-only gels. We incorporate small amounts of acrylate-PEG-succinimidyl valerate, which is an amine-reactive group, to couple full-length ECM proteins to the gel surface. We are developing this tunable hydrogel system to study how tissue modulus and integrin-binding ligands control the ability of carcinoma cells to respond to RTK inhibitors. As a mimic of precancerous tissue, we have a 5kPa gel with collagen IV, fibronectin, and laminin, and as a cancerous tissue, we have 20 and 55kPa gels with collagen I, III, and fibronectin. We have used this system to test carcinoma cell (liver: HEP3B, and breast: MDA-MB-231, MCF7, SKBR-3, and BT549) response to sorafenib, an FDA-approved RTK inhibitor. We are quantifying the cell response (apoptosis) with a proliferation assay and a real-time caspase 3/7 reporter.
Thus far, we have observed that both breast and liver carcinoma cells exhibit chemo-resistance in cancerous microenvironments. In fact, at least 30% additional sorafenib is required to inhibit proliferation in cells on cancerous tissues relative to healthy tissues. This resistance is coincided by visible morphological changes as well, wherein the cancerous microenvironment promotes less cell-cell contact and a spindle shape. We are currently investigating combinatorial treatments by targeting ERK/Akt signaling pathways to overcome matrix-conferred resistance. The drug resistance we have observed here may be one reason that drugs that show efficacy on tissue culture plates eventually fail in clinical trials. We propose that this high-throughput biomaterial system may serve as a system that pharmaceutical companies can use to rule out false positives and potentially save billions of dollars in the drug development pipeline.
Thuy V. Nguyen
Department of Chemical Engineering
University of Massachusetts
686 North Pleasant Street
159 Goessmann Lab
Amherst, MA 01003
Phone number: 404-933-4077