Dox E17K WT Basal mediumGrowth medium p-AKT S473 FLAG PI3K  p-AKT T308 p-mTOR S2448 p-mTOR S2481 mTOR Actin Davies BR et al. Supplementary Figure 1 MCF10-AKT1 E17K and MCF10A-AKT1 WT cells were cultured in basal or growth medium in the presence or absence of dox, and proteins were detected by Western blotting.

2 -Dox+Dox Davies BR et al. Supplementary Figure 2 AKT1 E17K AKT1 WT HER2 V659E RFP AKT1 E17K AKT1 WT HER2 V659E RFP Triplicate wells are shown from colony formation assay in soft agar.

3 Davies BR et al. Supplementary Figure 3 AKT1 E17K -DoxAKT1 E17K +Dox AKT1 WT -DoxAKT1 WT +Dox RFP +DoxRFP -Dox Mean colony diameter (  M) -Dox+Dox AKT1 E17K AKT1 WT 9597 RFP9196 Mean number of cells per colony -Dox+Dox AKT1 E17K AKT1 WT 6164 RFP6763 Number of colonies of particular diameters, mean colony diameter and mean number of cells per colony are shown from the soft agar assay.

4 Genomic DNA HBCx2 DNA Davies BR et al. Supplementary Figure 4 Detection of AKT1 E17K mutation in the HBCx2 explant model

GI50 = 2.4  MGI50 = 1.6  M Davies BR et al. Supplementary Figure 5 Effect of AKT inhibitors on signaling proteins and growth of MGH-U3 cells