The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is commonly dysregulated in human cancer, making it an attractive target for novel anticancer therapeutics. We have used a mouse model of ovarian cancer generated by Kras^G12D activation and Pten deletion in the ovarian surface epithelium for the preclinical assessment of a novel PI3K/mTOR inhibitor PF-04691502. To enable higher throughput studies, we developed an orthotopic primary transplant model from these mice and evaluated therapeutic response to PF-04691502 using small-animal ultrasound and FDG-PET imaging. PF-04691502 inhibited tumor growth at 7 days by 72% ± 9. FDG-PET imaging revealed that PF-04691502 reduced glucose metabolism dramatically, suggesting FDG-PET may be exploited as an imaging biomarker of target inhibition by PF-04691502. Tissue biomarkers of PI3K/mTOR pathway activity, p-AKT (S473), and p-RPS6 (S240/244), were also dramatically inhibited following PF-04691502 treatment. However, as a single agent, PF-04691502 did not induce tumor regression and the long-term efficacy was limited, with tumor proliferation continuing in the presence of drug treatment. We hypothesized that tumor progression was because of concomitant activation of the mitogen-activated protein kinase pathway downstream of Kras^G12D expression promoting cell survival and that the therapeutic effect of PF-04691502 would be enhanced by combinatory inhibition of MEK using PD-0325901. This combination induced striking tumor regression, apoptosis associated with upregulation of Bim and downregulation of Mcl-1, and greatly improved duration of survival. These data suggest that contemporaneous MEK inhibition enhances the cytotoxicity associated with abrogation of PI3K/mTOR signaling, converting tumor growth inhibition to tumor regression in a mouse model of ovarian cancer driven by PTEN loss and mutant K-Ras. Mol Cancer Ther; 10(8); 1440–9. ©2011 AACR.