Treatment options for triple-negative breast cancer (TNBC) are limited. Current 2D cancer models fail to accurately model the tumour microenvironment of breast cancer. Alterations to extracellular matrix (ECM) composition have been shown to play a key role in the epithelial-mesenchymal transition (EMT) process involved in breast cancer progression (1). This highlights the need for the development of a representative in vitro 3D model, in which to study TNBC behaviour and identify new treatment targets. This project aims to develop a collagen-based scaffold model composed of matrix components of breast tissue, including glycosaminoglycans, hyaluronic acid (HyA) and chondroitin sulphate (CS), both elevated in tumours, to investigate their role in the EMT process, within TNBC.
Collagen-based scaffolds comprised of varying concentrations of HyA, or CS were fabricated using previously optimised protocols (2). Scaffold characterisation was performed using SEM, porosity measurements, and mechanical testing. AlamarBlue and DNA assays were performed to assess the metabolic activity and growth of TNBC cell lines, MDA-MB-231 and MDA-MB-436, in comparison to normal breast epithelial cell line, MCF10a. The migratory ability of TNBC cells in 3D was assessed with H&E staining. A cytokine array and qPCR were performed to assess the effect of varying HyA and CS concentrations on MCF10a and MDA-MB-231 behaviour and to determine expression levels of markers associated with the EMT process.
All scaffolds were highly porous and had a uniform pore distribution, with an average stiffness of 1kPa and are therefore within the stiffness range of cancerous breast tissue (1kPa-4kPa). Each scaffold type exhibited huge biocompatibility for each cell line. TNBC cells were more metabolically active on CHyA scaffolds than CCS scaffolds and TNBC cells proliferated at a faster rate than MCF10a cells. Each scaffold type supported the migration of TNBC cells. Change in HyA or CS concentration did not affect cell proliferation but altered the expression of pro-inflammatory cytokines. Interestingly, an increase in HyA increased pro-inflammatory cytokine expression and an increase in CS decreased pro-inflammatory cytokine expression in MDA-MB-231 cells. The effect of glycosaminoglycan type and concentration on cytokine expression requires further investigation. Alterations in the expression of EMT associated markers differed with an increase in HyA and an increase in CS concentration, findings which require further investigation.
Collagen-based scaffolds composed of varying ECM components have been developed. Altering the mechanical stiffness of the collagen-based scaffolds within a range that represents cancerous breast tissue is achievable. Each scaffold is highly porous and supports cell viability and proliferation. Varying concentrations of HyA and CS alters the cytokine expression profile of MCF10a and MDA-MB-231 cells, highlighting the effect of changes to the ECM composition on cancer progression. This finding will be further investigated in future studies. In summary, the collagen-based scaffolds have the potential to mimic the ECM of breast tissue and have the capacity to be used as 3D models for breast cancer research.
Acknowledgements: Funded by Health Research Board
(1) Scott LE, Weinberg SH, Lemmon CA. Front Cell Dev Biol, (2019)
(2) Haugh MG et al., Tissue Eng - Part A,1201–8 (2011)