Extra file two. Figure S2 demonstrates that LY294002 correctly inhibits PI3K by evidence of reduced phosphorylated AKT protein ranges within the 4 PTEN mutant melanoma cell lines that normally express large amounts of pAKT, Additionally, Additional file three. Figures S3 and Added file four. Figure S4 present the concentration result curves for single agent LY294002 and E6201 respectively, wherever each medication had been added 24 hours following plating. The six melanoma cell lines examined displayed related trends in E6201 sensitivity when compared with our preceding experiments, with MM622, MM540, UACC903, and WM35 being essentially the most sensitive and UACC558 and UACC647 remaining much less delicate, Surprisingly, all cell lines showed equivalent sensitivity to LY294002, with IC50 ranging from eleven uM to 17 uM.
This was sudden, as one would predict MM540 and WM35 cells kinase inhibitor Paclitaxel for being somewhat resistant to PI3K inhibition offered the lack of detectable levels of pAkt Camptothecine indicating no constitutive PI3K activation in these cell lines. A past study by Smalley and other people, on the other hand, reported a equivalent sensitivity of WM35 cells to LY294002. The concentration response curves for E6201 and LY294002 combinations, normalized to a dimethyl sulf oxide handle are offered in Added file 4. Figure S4. As variations in synergy may well exist at vary ent drug result amounts, we graphed person combin ation index values for LY294002 with raising concentrations of E6201 for each cell line, As shown in Figure 5A, evaluating the individual com bination index for all combinations tested uncovered that E6201 and LY294002 exhibit synergistic exercise in all six melanoma cell lines, irrespective of E6201 sensitivity or PTEN or pAkt status.
Interestingly, various patterns of synergy have been observed amid the groups of cell lines tested. Whilst almost all of the cell lines showed an in creasing combination index at greater concentrations of E6201, UACC647 and UACC558 cells showed a decreasing combination index or enhanced synergy with expanding concentrations of E6201. Notably, this pattern observed for UACC647 and UACC558 cells occurs within the context of high pAkt and relative resistance to E6201, supporting the hypoth esis that administration of a PI3K inhibitor can sensitize E6201 resistant cells with large pAkt amounts to E6201. In summary, the blend of E6201 and LY294002 resulted in synergistic action in all 6 melanoma cell lines examined, as defined by a blend index 1.