Human solid tumours are comprised of epithelial cells interacting with a surrounding stroma or microenvironment. The stroma consists of several cell types including immune cells, endothelial cells and fibroblasts as well as macromolecules constituting the extracellular matrix. It has become clear from both pre-clinical and clinical studies that the tumour stroma plays a critical role in tumour progression and response to therapeutics. TGF-b is one of a number of pivotal regulators of the tumour microenvironment. Among its roles, TGF-b drives the cross-talk between tumour cells and fibroblasts resulting in a cancer associated fibroblast phenotype. This activated CAF state is linked to increased progression and survival of tumour cells and response of tumours to therapeutics.

Modeling this cross-talk is challenging however we have developed in vitro assays using tumour cells from a range of tissue types, for example NSCLC, breast, pancreatic and prostate, co-cultured with normal human fibroblasts and assessed cell/cell and cell/extracellular matrix interactions using a range of end-points including -smooth muscle actin and fibronectin. We have further characterized the assays, for NSCLC cell lines, using pharmacologically active agents to modulate canonical and non-canonical TGF-beta signaling pathways and assessed the effects on tumour cell growth, fibroblast activation and secretion of key growth factors, cytokines and proteases. We have also used RNA expression profiling of a selection of genes to investigate signaling pathways activated in tumour and fibroblast co-cultures compared to when the cells are grown as monocultures.

We will describe the development of methods to establish and quantify the different end-points explored in tumour and fibroblast co-culture assays and show data using inhibitors to validate these methods with NSCLC cell lines. We will show data from further comparison of monocultures and co-cultures by analysis of RNA and protein in cell lysates and conditioned media samples.

The development of more complex in vitro assays with more than a single cell type allows greater insight into the various tumour-stromal interactions that take place in tumours in vivo. This allows early understanding and testing of agents which may provide therapeutic benefit in limiting tumour-stromal interactions and signalling in patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B127.