Fig 5. GSK461364 plus docetaxel reduce tumorsphere formation of SUM149 and SUM159.

(A, histograms and B, pictures) For the primary tumorsphere assay, we analyzed the tumorsphere formation in presence or absence of docetaxel, GSK461364, and the combination at the IC₅₀ concentrations identified in the proliferation assay; SUM149 and SUM159 cells were plated at a density of 20,000 cells per well in ultra-low attachment 6 well plates and after 24 hours treated with DMSO (control), docetaxel IC₅₀, GSK461364 IC₅₀, or both in combination. Formed spheres were counted using the Celigo Imaging Cytometer (Nexcelom, USA). GSK461364 plus docetaxel decreased primary tumorsphere formation of SUM149 and SUM159 (Interaction Test, p = 0.01* and p < 0.001⁺, respectively). (C, histograms and D, pictures) Dissociated tumorspheres (N = 500) were plated in ultra-low attachment 96 well plates (Costar^®, USA) in serum-free medium for an additional seven days in the absence of drug. Secondary passage tumorspheres were then counted as before. GSK461364 plus docetaxel decreased primary tumorsphere formation of SUM149 and SUM159 (Interaction Test, p < 0.001⁺ and p < 0.001⁺, respectively). Poisson regression model with drug interaction was used to assess statistical differences.