(306a) Receptor Tyrosine Kinase Inhibitor Efficacy in Carcinoma Is Stiffness Dependent

There is a critical need for model systems to support the efforts of drug development. In particular, several groups are creating tissue mimics that capture key features of the in vivo cell and tissue microenvironment to test true drug efficacy and latent toxicity in physiologically relevant microenvironments.  Our lab has developed a high-throughput system to quantify carcinoma cell response to receptor tyrosine kinase (RTK) inhibitors during tumor growth and stroma stiffening. 

This platform consists of a new class of hydrogels with an extremely wide range of mechanical properties that combine poly(ethylene glycol) (PEG), and  phosphorylcholine (PC) zwitterions. The Young’s modulus of these “PEG-PC” hydrogels can be tuned over four orders of magnitude, much greater than conventional PEG-diacrylate or PEG–dimethacrylate gels. We are synthesizing these PEG-PC hydrogels within a silane-treated glass 96-well plate, wherein we can control the modulus of the hydrogels from 900Pa to 10MPa, and couple full-length ECM proteins to the gel surface. These ECM proteins are made in combinations to capture the integrin-binding present pre-tumorigenesis (collagen IV, fibronectin, and laminin), and late in tumor progression (collagen I, III, and fibronectin). We have used this system to quantify carcinoma cell (liver: HEP3B, and breast: MDA-MB-231, SKBR-3, and BT549) response to sorafenib, an FDA-approved RTK inhibitor, as a function of tissue stiffness and integrin binding.

We have observed that tissue stiffening increases the resistance of carcinoma cells to sorafenib by nearly three-fold, and there is a synergistic effect on collagen-rich, stiff environments, compared with soft, pre-tumorigenic integrin-binding proteins.  This stiffness-conferred chemo-resistance is dampened when sorafenib is co-administered with ROCK inhibitor, β1 integrin blocking antibodies, or an epidermal growth factor receptor (EGFR) inhibitor.  Interestingly, these combinatorial treatments were more effective on triple negative breast cancer cells than Herceptin receptor overexpressing cells.  Toward more rational identification of combinatorial treatments, we used a bead-based ELISA (MAGPix, Luminex) to identify JNK as a promising phospho-protein target, which provided survival signals during substrate stiffening.  Dosing carcinoma cells with JNK inhibitor and sorafenib was the one combination, which eliminated stiffness sensitive resistance.  Most excitingly, we have identified a sub-population of carcinoma cells, within these cell lines, which express qualities of cancer stem cells, and are enriched during substrate stiffening.  We are currently investigating RTK inhibitor combinatorial treatments to eliminate this dangerous, drug resistant subpopulation of cells. 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.