Analysis of RAS mutation and Akt activation in primary MM samples. (A) RAS mutation status and sensitivity to Akt blockade in freshly isolated primary MM cells (n = 25). Viability measurements of MM cells cocultured with bone marrow stromal cells were performed after 5-day treatment with 10μM Akti-1,2. Sensitivity to Akt inhibition shows no obvious correlation with the presence or absence of RAS mutation. Medians are indicated. (B) Correlation between Akti-1,2 (10μM) sensitivity of freshly isolated, cocultured primary MM cells and the presence or absence of phospho-Akt in immunohistochemical stains of matched bone marrow biopsies (n = 19). Phospho-Akt positivity in the biopsies was significantly (P < .05) correlated with sensitivity to Akt inhibition of the respective MM cells. (C) RAS mutation in primary MM cells and phospho-Akt status in corresponding bone marrow biopsies (n = 59). RAS mutations were present in 16 of 34 phospho-Akt-positive and in 8 of 25 phospho-Akt-negative samples. (D) Two examples of MM patient biopsies with immunofluorescent double staining for plasma-cell marker CD138 (green) and either phospho-Akt Ser473 or phospho-Akt Thr308 (both in red). A phospho-Akt-negative (patient 6) and a phospho-Akt-positive (patient 20) sample are shown. Bar represents 75 μm.