Scaffold stiffness influences breast cancer cell invasion via EGFR-linked Mena upregulation and matrix remodeling

Matrix Biol. 2020 Jan;85-86:80-93. doi: 10.1016/j.matbio.2019.07.006. Epub 2019 Jul 16.

Abstract

Clinically, increased breast tumor stiffness is associated with metastasis and poorer outcomes. Yet, in vitro studies of tumor cells in 3D scaffolds have found decreased invasion in stiffer environments. To resolve this apparent contradiction, MDA-MB-231 breast tumor spheroids were embedded in 'low' (2 kPa) and 'high' (12 kPa) stiffness 3D hydrogels comprised of methacrylated gelatin/collagen I, a material that allows for physiologically-relevant changes in stiffness while matrix density is held constant. Cells in high stiffness materials exhibited delayed invasion, but more abundant actin-enriched protrusions, compared to those in low stiffness. We find that cells in high stiffness had increased expression of Mena, an invadopodia protein associated with metastasis in breast cancer, as a result of EGFR and PLCγ1 activation. As invadopodia promote invasion through matrix remodeling, we examined matrix organization and determined that spheroids in high stiffness displayed a large fibronectin halo. Interestingly, this halo did not result from increased fibronectin production, but rather from Mena/α5 integrin dependent organization. In high stiffness environments, FN1 knockout inhibited invasion while addition of exogenous cellular fibronectin lessened the invasion delay. Analysis of fibronectin isoforms demonstrated that EDA-fibronectin promoted invasion and that clinical invasive breast cancer specimens displayed elevated EDA-fibronectin. Combined, our data support a mechanism by which breast cancer cells respond to stiffness and render the environment conducive to invasion. More broadly, these findings provide important insight on the roles of matrix stiffness, composition, and organization in promoting tumor invasion.

Keywords: Biomaterials; Extracellular matrix; Tumor microenvironment.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Breast Neoplasms / genetics
  • Breast Neoplasms / metabolism
  • Breast Neoplasms / pathology*
  • Cell Culture Techniques
  • Cell Line, Tumor
  • Cell Movement
  • ErbB Receptors / metabolism
  • Extracellular Matrix / metabolism
  • Extracellular Matrix / pathology*
  • Female
  • Fibronectins / genetics
  • Fibronectins / metabolism
  • Gene Knockdown Techniques
  • Humans
  • Hydrogels
  • Microfilament Proteins / metabolism*
  • Neoplasm Invasiveness
  • Phospholipase C gamma / genetics
  • Spheroids, Cellular / cytology
  • Spheroids, Cellular / metabolism
  • Spheroids, Cellular / pathology
  • Transcriptional Activation

Substances

  • Enah protein, human
  • FN1 protein, human
  • Fibronectins
  • Hydrogels
  • Microfilament Proteins
  • extra domain A fibronectin, human
  • EGFR protein, human
  • ErbB Receptors
  • PLCG1 protein, human
  • Phospholipase C gamma