Mammalian target of rapamycin (mTOR) inhibitors, such as rapamycin and CCI-779, have shown preclinical potential as therapy for multiple myeloma. By inhibiting expression of cell cycle proteins, these agents induce G1 arrest. However, by also inhibiting an mTOR-dependent serine phosphorylation of insulin receptor substrate-1 (IRS-1), they may enhance insulin-like growth factor-I (IGF-I) signaling and downstream phosphatidylinositol 3-kinase (PI3K)/AKT activation. This may be a particular problem in multiple myeloma where IGF-I-induced activation of AKT is an important antiapoptotic cascade. We, therefore, studied AKT activation in multiple myeloma cells treated with mTOR inhibitors. Rapamycin enhanced basal AKT activity, AKT phosphorylation, and PI3K activity in multiple myeloma cells and prolonged activation of AKT induced by exogenous IGF-I. CCI-779, used in a xenograft model, also resulted in multiple myeloma cell AKT activation in vivo. Blockade of IGF-I receptor function prevented rapamycin's activation of AKT. Furthermore, rapamycin prevented serine phosphorylation of IRS-1, enhanced IRS-1 association with IGF-I receptors, and prevented IRS-1 degradation. Although similarly blocking IRS-1 degradation, proteasome inhibitors did not activate AKT. Thus, mTOR inhibitors activate PI3-K/AKT in multiple myeloma cells; activation depends on basal IGF-R signaling; and enhanced IRS-1/IGF-I receptor interactions secondary to inhibited IRS-1 serine phosphorylation may play a role in activation of the cascade. In cotreatment experiments, rapamycin inhibited myeloma cell apoptosis induced by PS-341. These results provide a caveat for future use of mTOR inhibitors in myeloma patients if they are to be combined with apoptosis-inducing agents.