Rapamycin and several analogs, such as CCI-779 and RAD001, are currently undergoing clinical evaluation as anticancer agents. In this study, we show that inhibition of mammalian target of rapamycin (mTOR) signaling by rapamycin leads to an increase of Akt phosphorylation in Rh30 and RD human rhabdomyosarcoma cell lines and xenografts, and insulin-like growth factor (IGF)-II-treated C2C12 mouse myoblasts and IGF-IIoverexpressing Chinese hamster ovary cells. RNA interference-mediated knockdown of S6K1 also results in an increase of Akt phosphorylation. These data suggest that mTOR/S6K1 inhibition either by rapamycin or small interfering RNA (siRNA) triggers a negative feedback loop, resulting in the activation of Akt signaling. We next sought to investigate the mechanism of this negative feedback regulation from mTOR to Akt. Suppression of insulin receptor substrate (IRS)-1 and tuberous sclerosis complex-1 by siRNAs failed to abrogate rapamycininduced upregulation of Akt phosphorylation in both Rh30 and RD cells. However, pretreatment with h7C10 antibody directed against insulin-like growth factor-1 receptor (IGF-1R) led to a blockade of rapamycin-induced Akt activation. Combined mTOR and IGF-1R inhibition with rapamycin and h7C10 antibody, respectively, resulted in additive inhibition of cell growth and survival. These data suggest that rapamycin mediates Akt activation through an IGF-1R-dependent mechanism. Thus, combining an mTOR inhibitor and an IGF-1R antibody/inhibitor may be an appropriate strategy to enhance mTOR-targeted anticancer therapy.

Obesity is becoming more prevalent worldwide and is a major risk factor for cancer development. Acute myeloid leukemia (AML), the most common acute leukemia in adults, remains a frequently fatal disease. Here, we investigated the molecular mechanisms by which obesity favors AML growth and uncovered the fatty acid binding protein 4 (FABP4) and DNA methyltransferase 1 (DNMT1) regulatory axis that mediates aggressive AML in obesity. We showed that leukemia burden was much higher in high-fat diet-induced obese mice, which had higher levels of FABP4 and IL-6 in sera. Upregulation of environmental and cellular FABP4 accelerated AML cell growth in both a cell-autonomous and cell-non-autonomous manner. Genetic disruption of FABP4 in AML cells or in mice blocked cell proliferation in vitro and induced leukemia regression in vivo. Mechanistic investigations showed that FABP4 upregulation increased IL-6 expression and STAT3 phosphorylation leading to DNMT1 overexpression and further silencing of the p15INK4B tumor suppressor gene in AML cells. Conversely, FABP4 ablation reduced DNMT1-dependent DNA methylation and restored p15INK4B expression, thus conferring substantial protection against AML growth. Our findings reveal the FABP4/DNMT1 axis in the control of AML cell fate in obesity, and suggest that interference with the FABP4/DNMT1 axis might be a new strategy to treat leukemia.