Figure 7

Organotypic collagen invasion of SCC12 cells is inhibited by BDP5290. (A) SCC12 human squamous cell carcinoma cells were treated with indicated concentrations of BDP5290 or Y27632 for 30 minutes prior to cell lysis and quantitative western blotting with indicated antibodies. (B) Inhibition of MLC phosphorylation by BDP5290 and Y27632. All results are shown as mean ± standard error of n = 3 independent replicates. (C) Alamar Blue metabolism of SCC12 cells after 24 hours in the presence of DMSO vehicle, 2 μM BDP5290 or 2 μM Y27632. Readings were compared to the Alamar Blue metabolism in DMSO treated cells that were used as the viability standard of 100% for each replicate experiment. (D) Percentage cell confluence of 35,000 cells (set to 100% at t = 0) plated in replicate wells of 96 well plates in the presence of DMSO vehicle, 2 μM BDP5290 or 2 μM Y27632 was determined after 21 hours by imaging with an Incucyte Zoom. (E) Example images of SCC12 cells invading collagen, that had been conditioned by cancer associated fibroblasts for five days prior to their removal, in the presence of DMSO vehicle 2 μM BDP5290 or 2 μM Y27632 for 10 days. (F) Number of invading cells was determined in 5 random fields for each condition for each separate replicate experiment. All results are shown as mean ± standard error of n = 3 independent replicates.