Oral Presentation Symposium on Proteases and the Tumouri Microenvironment 2017

Bioengineered microtissues enable analysis of the human prostatic tumour microenvironment in vitro (#5)

Brooke A Pereira 1 , Natalie L Lister 1 , Birunthi Niranjan 1 , Mitchell G Lawrence 1 , Dietmar W Hutmacher 2 3 , Stuart J Ellem 1 , Elena M De-Juan-Pardo 2 3 , Gail P Risbridger 1
  1. Department of Anatomy and Developmental Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia
  2. Centre In Regenerative Medicine, Institute of Health & Biomedical Innovation, Queensland University of Technology, Kelvin Grove, QLD, Australia
  3. ARC Centre In Additive Biomanufacturing, Queensland University of Technology, Kelvin Grove, QLD, Australia

The tumour microenvironment (TME) plays a fundamental role in prostate tumourigenesis. Classical tissue recombination experiments show tumour stroma directs tumour formation in benign epithelia, yet despite this, stromal components are frequently absent from human prostate cancer (PCa) models, including cancer associated fibroblasts (CAFs) and the extracellular matrix (ECM) they deposit. Here we describe a three-dimensional (3D) human prostate TME microtissue model which has been utilised to interrogate the contribution of CAFs and ECM, and how they promote tumourigenicity in the prostate.

Scaffolds were formatted from poly(ε-caprolactone) (PCL) via melt electrospinning writing (MEW) in a 0/90⁰ pattern, forming flexible 15x15mm2 meshes with a pore size of ~150µm. Scaffold surface were treated with plasma to improve cell attachment. Validated patient-derived primary CAFs or non-malignant prostatic fibroblasts (NPFs) were incorporated into the scaffolds, forming a 3D microtissue. Native ECM deposition deposited by the fibroblasts was evident. Once confluent, tagged benign epithelial cells (BPH-1 or RWPE-1) were co-cultured on the microtissue constructs. Subsequently, microtissues were fixed and tumourigenicity was assessed by analysing the 3D morphological transformation of the epithelial cells using confocal microscopy. Additionally, migration assays were performed to further quantify invasive potential.

The data show that CAF and NPF proliferate and deposit morphologically distinct ECM to form a 3D stromal network within the scaffolds. CAF, but not NPF, microtissues induce an invasive morphology in the benign epithelium. Furthermore, as the epithelial cells malignantly transform they develop invasive invadopodia, which are only detected in the microtissue model.

Overall, our results highlight the importance of including TME components in in vitro cancer models. CAFs and ECM significantly contribute to prostatic epithelial tumourigenesis. These results have prompted further transcriptomic analyses (RNASeq) of the epithelial cells co-cultured with CAF/NPF to fully elucidate the signalling pathways in the prostatic TME.