High grade astrocytomas (HGA) remain fatal without effective treatment. Using an inducible Cre-driven adult astrocyte-specific system, we explored the relative roles of key pathways perturbed in human glioblastomas (grade IV; GBM) in initiation and progression of HGA. The likely event combinations (engineered and spontaneous) yielding disease indicate grade-specific roles for each aberration and suggest specific progression mechanisms from grade II [induced only by pRb-tumor suppression (TS) inactivation] to III (addition of KrasG12D activation, with spontaneous inactivation of p53 by mutation or mislocalization), to Grade IV [further addition of PTEN inactivation (spontaneous or engineered)] without IDH1 mutation. In the transition from grade II to III disease and subsequent to KrasG12D activation, Trp53 missense mutations congruent with human GBM mutations. This study underscores the importance of stochastic events with evident tumor heterogeneity in order to recapitulate disease properties. Importantly, murine GBM transcriptomes showed concordance with the highly aggressive human mesenchymal GBM subclass.To determine whether EGFR played a role on tumor progression in our astrocytoma mouse model with inactivation of Rb-TS and activation of KrasG12D (TR model), we performed immunohistochemistry (IHC) study and FISH analysis on TR tumors, and found that EGFR was amplified/overexpressed. Inhibition of EGFR by EGFR inhibitor Erlotinib showed resistance in vivo and in vitro on TR tumors, which is consistent with the human clinical data. However, these tumors showed sensitivity to multiple RTK inhibitors in vitro, indicating that other RTKs may compensate for single EGFR inhibition. We are currently working on molecular pathway analysis to dissect the compensatory mechanism of EGFR inhibition. Song Y, Zhang Q, Bash R, Kutlu B, Difilippantonio S, Yin C, Gilbert D, Wang C, Yang C, Bullitt E, Kafri T, McCarthy K, Louis D, Hood L, Miller CR, Van Dyke T. An evolutionary path to glioblastoma: Insight into etiology from engineered mice. (Under review at Cancer Discovery)