Fas and PI3K/Akt signaling pathways pivotally impact on cancer cell death and survival respectively and are considered as promising targets for innovative anticancer therapies. To better characterize the combination effect of PI3K/Akt inhibitors and Fas agonists and understand the profile of the interaction between PI3K/Akt and Fas signaling, we qualitatively and quantitatively evaluated the combination effect of PI3K/Akt inhibitors LY294002, Akt inhibitor VIII and FasL. At the concentration that can block cell cycle progression and DNA synthesis but not elicit apoptosis, these inhibitors potentiate FasL to induce apoptosis. At higher concentrations, when the PI3K/Akt inhibitors induce apoptosis, they synergize FasL to induce apoptosis. In addition, PI3K/Akt inhibition significantly facilitates the Fas-mediated apoptotic signaling. Understanding the combination effects between PI3K/Akt inhibition and Fas activation not only leads to rational design of effective combination therapy of PI3K/Akt inhibitors but also improve our knowledge about the impact of PI3K-Akt pathway on Fas signaling and the potential modulation of innate immune system by PI3K-Akt-targeting drugs in anticancer treatment.