Fig. 5

Comparative of NUAK1 inhibition versus mTOR inhibition on Akt signaling. A IB of combined inhibition of NUAK1 and mTOR on Akt signaling. MDA-MB-231 cells were serum-starved overnight followed by 1-h of pretreatment with DMSO, HTH-01-015 (10 µM) or HTH-01-015 (10 µM) plus Torin1 (100 nM) before stimulation with EGF. B IB of Akt Ser-473 phosphorylation under NUAK1 inhibition in MDA-MB-231 shCtrl and MDA-MB-231 shRictor cells. Stable cells were serum-starved overnight followed by 1-h of pretreatment with DMSO or HTH-01-015 (10 µM) before stimulation with EGF for 60 min. C IB of NUAK1 and mTOR effect on Akt signaling. MDA-MB-231 cells were serum-starved overnight followed by 1-h of pretreatment with DMSO, HTH-01-015 (10 µM) or Torin1 (100 nM) before stimulation with EGF for 0 and 20 min. D Quantification of Akt signaling pathway from C. Each bar represents the mean ± SD, n = 3. Data from 3 independent experiments at 20 min of EGF stimulation were analyzed by student t test. E IB of Akt/TSC2 signaling under NUAK1 inhibition in MDA-MB-231 shCtrl and MDA-MB-231 shRictor cells. Stable cells were serum-starved overnight followed by 1-h pretreatment with DMSO or HTH-01-015 (10 µM) before stimulation with EGF for 20 min. GAPDH and/or α-tubulin were used as loading controls