Expression of mutant P85α promoted transformation of GBM-relevant cells both <i>in vitro</i> and <i>in vivo</i>.

<p>(A) Hs683 cells co-expressing wildtype or mutant P85α and wildtype P110α were seeded in soft agar in triplicate. The mean colony numbers (±SD) from a representative experiment are shown. All three mutant P85α constructs significantly (<i>p</i><0.001) promoted colony formation relative to cells expressing wildtype P85α or untransduced cells. (B) E6/E7/hTERT-immortalized normal human astrocytes co-expressing wildtype or mutant P85α and wildtype P110α were injected subcutaneously in nude mice (n = 8 injections for DKRMSNS560del subline, and n = 10 injections for all other sublines). Tumor volume measurements taken 61 days post-transplantation show that P85α mutant tumors were larger. (C) The total penetrance for each subline upon termination of the experiment 300 days post-injection is shown. (D) Whole cell lysates were generated from two tumors of each subline. Western blotting confirmed that tumors retained expression of mutant P85α, and that tumors expressing mutant P85α demonstrated higher levels of pAKT. Numerical values below the pAKT panel of the immunoblot represent quantification of the relative protein level by densitometry (normalized to AKT).