Designing and utilizing synthetic extracellular matrices to probe breast cancer cell activation in response to microenvironment cues
June 5-9, 2018
Interactions between breast cancer cells and their microenvironments are essential in tumor growth, metastasis, and recurrence. The tumor stroma undergoes constant structural changes, including degradation, redeposition, and crosslinking of collagens with gradients in matrix stiffness and composition that drive invasion and metastasis. At metastatic sites, similar remodeling events that occur with injury and aging are hypothesized to promote reactivation of dormant tumor cells in recurrence. Approaches are needed for testing hypotheses about pivotal cell-matrix interactions in the progression of breast cancer for identifying key regulators and improving treatment strategies.
In this work, we have designed well-defined materials to mimic key aspects of tumor microenvironments toward studying such complex phenomena in vitro. Specifically, we have created synthetic extracellular matrices with well-defined biophysical and biochemical properties that enable three-dimensional (3D) culture of breast cancer cells and niche cells over weeks. A biologically inert polymer, multi-arm poly(ethylene glycol) functionalized with thiols (Mn ~ 20 kDa), was reacted with integrin-binding and cell-degradable peptides decorated with one and two allyloxycarbonyl protecting groups, respectively, by rapid light-triggered thiol-ene polymerization for independent control of matrix density and composition. The elasticity, or ‘stiffness’, of these matrices has been tuned to mimic a variety of soft tissues (Young’s modulus E~0.5-20 kPa), from healthy and cancerous mammary tissues to metastatic site bone marrow and lung tissues. Further, the biochemical content has been tuned with receptor-binding peptides derived from laminin (IKVAV, laminin receptor), collagen ((POG)3POGFOGER(POG)4, α2β1 and α1β1), and fibronectin/vitronectin (RGDS, αVβ3 and α5β1 amongst others) and a crosslinking peptide derived from collagen (GPQG↓IWGQ, degraded MMP-1, -2, and -9 amongst others).
We hypothesized that a microenvironment rich in collagen and fibronectin/vitronectin, mimicking aspects of remodeled tissues, would activate breast cancer cells relative to a laminin-rich epithelium-like microenvironment, building upon seminal studies in naturally-derived matrices and in vivo. To test this, we cultured breast cancer cells of different metastatic potential (estrogen receptor positive [ER+, T47Ds] and triple negative [ER-, MDA-MB-231s]) within different matrix densities and compositions. Both cell types exhibited high viability (> 90%), and cell activation in response to different matrix compositions was assayed by examining proliferation (metabolic activity, Ki-67, cell/cluster number and volume) and phenotype (morphology; E-cadherin, vimentin). Increased matrix density decreased elongation of ER- cells and proliferation of both cell types. Increased collagen content increased the proliferation of the ER+ cells and proliferation and elongation of ER- with mass and stellate morphologies, respectively, like observed in naturally-derived matrices. These studies demonstrate a new tool for controlled 3D culture of breast cancer cells relevant for both fundamental and applied research, with on-going investigations incorporating niche cells and triggered matrix changes.
April M. Kloxin, Lisa Sawicki, Elisa Ovadia, and Lina Pradhan, "Designing and utilizing synthetic extracellular matrices to probe breast cancer cell activation in response to microenvironment cues" in "Nanotechnology in Medicine II: Bridging Translational in vitro and in vivo Interfaces", Millicent Sullivan, PhD, University of Delaware, USA Josué Sznitman, Dr. Sc., Technion-Israel Institute of Technology, Israel Lola Eniola-Adefeso, PhD, University of Michigan, USA Srivatsan Kidambi, PhD, University of Nebraska - Lincoln, USA Eds, ECI Symposium Series, (2018). http://dc.engconfintl.org/nanotech_med_ii/17