Co-operative induction of apoptosis by ABT-199 with pladienolide B or torin1.

<p><b>(A)</b> MCL-1 long form (L) and short form (S) transcripts (i) and protein (ii) were quantified in untreated cells and cells treated with 10 nM pladienolide B (PLB) for 4 hours. <b>(B)</b> 4E-BP1 phosphorylation (i) and MCL-1 protein (ii) were quantified in untreated cells and cells treated with 1μM torin1 for 4 hours. (<b>C, D)</b> Cells were incubated with 10 nM ABT-199 (199, turquoise), 10 nM pladienolide B (PLB, red), 1 μM torin1 (green) or the indicated combinations (bright blue bar, height = effect with both agents in combination–sum of effects with agents individually). <b>(C)</b> After 4 hours cells were fixed and processed for Cytochrome C release. <b>(D)</b> After 4 hours DiOC6 was added for a further 75 minutes and 7-amino actinomycin D for the last 30 minutes. (Mean+/- SD for n = 3). Fold excess additivism (FEA) is shown on the figures and was calculated as a ratio of observed to expected values after corrections according to the Bliss algorithm (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0190682#sec002" target="_blank">methods</a>). Asterisks indicate observed values significantly higher than expected values (P<0.05).</p>