Fig. 5.

Combination studies of BEZ235 and Dox demonstrate synergy in LMS cell lines in vitro. a Dose schedule used in combination treatment. Three treatment schedules were investigated: schedule 1—concurrent treatment for 72 h, schedule 2—single agent therapy with inhibitor for the first 24 h followed by concurrent Dox treatment for the next 48 h and schedule 3—single agent Dox treatment for the first 24 h followed by concurrent treatment with the inhibitor for 48 h. b Combination index (CI) graphs resulting from the treatment of SKLMS1 cells with BEZ235 or BKM120 and Dox following 3 dosing schedules to determine optimal treatment regime. Viability was determined using ATPlite and analysed using CalcuSyn software. Treatment with BEZ235 (15–240 nM) and Dox (125–2000 nM) showed synergy in all 3 schedules (CI < 0.9), while combination BKM120 and Dox treatment was not synergistic in any of the treatment schedules (n = 3). For detailed CI ranges see Additional file 1: Table S2. c Immunoblot demonstrates decrease in p-AKT^S473, and p-4EBP1^T37/46 levels in total lysates from STS39 cells treated with BEZ235 for 72 h at the indicated concentrations and in combination with Dox. Total AKT, 4EBP1 and tubulin levels are shown as the loading controls