Figure 5. Differential effects of drug combination on PI3K/AKT signaling and activation of ER-stress transducers and death sensors.

The effects of DTX (10 nM), NFR (5 µM) and CUR (5 µM), alone and in combination, on AKT-phosphorylation (a, b), ER-stress transducers (c–f), and ER-stress death sensors (g–l) were investigated by western blotting using lysates from both C4-2B and RWPE-1 cells. In panels (a) and (b), both total AKT (t-AKT) and IGF-1 induced AKT activation (p-AKT) levels are shown after 6 hrs post drug exposures. In panels (c–f), the expression of BiP and p-eIF2α, at 6 hrs of drug exposure is shown. The expression of CHOP and ATF4 following 3 hr drug treatment are shown in both C4-2B (g & i) and RWPE-1 (h & j) cells. Expression of TRIB3 following 6 hr drug treatment, alone or in combination, are shown in both C4-2B (k) and RWPE-1 (l) cells. Band intensities were quantified by the Image-J software. For both p-AKT and p-eIF2α quantifications, band intensities were first normalized to β-actin levels and then with either t-AKT or t-eIF2α levels, respectively. For all other proteins, normalization with β-actin levels was carried out. Treatment specific changes (lanes 2–8) are expressed as fold changes compared to untreated controls (lane-1).