Fig. 5.

Melanoma cells co-cultured with TAB cells or recombinant IGF-1 are resistant to signaling inhibitors. a Melanoma cells (451Lu) co-cultured with TAB cells for 4–5 days are resistant to subsequent BRAFi (PLX4720) and MEKi (PD0325901) treatment (red lines) as compared with tumor cells cultured in the presence of NB cells (blue lines) or media control only (black lines). Co-cultured tumor cells were treated with drugs in triplicates for 72 h and viability was determined using the AlamarBlue assay. Results are expressed as relative viability of melanoma cells and drug responses are compared with appropriate controls. Data represents mean + SE of triplicate samples and p values (t-test) as indicated are for drug doses 0.1 to 10 nM of BRAFi and 0.01 nM to 1 nM of MEKi. b Melanoma cells treated with recombinant IGF-1 show resistance to BRAFiand MEKi. Melanoma cells (451Lu) were treated every day for a total of 5 days with recombinant IGF-1 (25 ng/ml; red lines), then harvested and their dose responses to BRAFiand MEKi determined as above. Results are compared with untreated melanoma cells as controls (blue lines). c, d Melanoma cells (451Lu (c) and WM989 (d)) co-cultured with TAB cells for 4 days in presence of anti-IGF-1 neutralizing antibody (blue lines) or media control (black lines) are sensitive to subsequent BRAFi and MEKi treatment as compared with cultures incubated with control antibody (red lines)