C95
Development of 3D tumour cell and HUVEC assays for phenotypic assessment of small molecular weight and biologic therapeutics
Sarah Ross, Cath Eberlein, Margaret Veldman-Jones, Weir Hazel, Natalie Byrne, Simon Barry
Cancer and Infection Research Area, AstraZeneca,, Alderley Park, Macclesfield, Cheshire, SK10 4TG, UK
Three dimensional in vitro tumour cell and endothelial cell culture models may more closely resemble the in vivo situation than cells grown in a monolayer. Studies have shown differences in cell morphology, growth, gene expression, cell-cell and cell-environment interactions, which would indicate that 3D assays could be advantageous in selecting clinically relevant cancer therapies.
We have developed a 96-well format, 3D colony cell assay and have explored methods of quantifying inhibition of colony growth by anti-tumour therapies. The 3D growth of a panel of tumour cell lines, representing the major solid tumour classes, has been assessed comparing soft agar and matrigel as 3D matrices. Better colony growth for more cell types was observed in matrigel. Inhibition of 3D growth by cytotoxic agents and inhibitors of tumour promoting signalling pathways has been examined. MTT staining of colonies and colony analysis using GelCount (Oxford optronix) were compared as methods of quantifying colony growth and sensitivity to anti-tumour agents. The Gelcount platform gave similar results to staining with MTT, but with the ability to identify more subtle effects of agents on colony growth and measurement of additional parameters including colony diameter.
In addition a 3D in vitro bead angiogenesis co-culture assay has been developed with Human Umbilical Vein Endothelial cells (HUVEC) and fibroblasts. 3D growth of HUVECs coated onto cytodex 3 beads has been analysed in response to growth factors and inhibitors of angiogenesis. A method for quantitative analysis of 3D tube growth has been developed using fluorescence imaging and a Cellomics ArrayScan platform.
