Oncogenic mutations of the receptor tyrosine kinase c-KIT play an important role in the pathogenesis of gastrointestinal stromal tumors, systemic mastocytosis, and some acute myeloid leukemias (AML). Although juxtamembrane mutations commonly detected in gastrointestinal stromal tumor are sensitive to tyrosine kinase inhibitors, the kinase domain mutations frequently encountered in systemic mastocytosis and AML confer resistance and are largely unresponsive to targeted inhibition by the existing agent imatinib. In this study, we show that myeloid cells expressing activated c-KIT mutants that are imatinib sensitive (V560G) or imatinib resistant (D816V) can inhibit the tumor suppressor activity of protein phosphatase 2A (PP2A). This effect was associated with the reduced expression of PP2A structural (A) and regulatory subunits (B55α, B56α, B56γ, and B56δ). Overexpression of PP2A-Aα in D816V c-KIT cells induced apoptosis and inhibited proliferation. In addition, pharmacologic activation of PP2A by FTY720 reduced proliferation, inhibited clonogenic potential, and induced apoptosis of mutant c-KIT + cells, while having no effect on wild-type c-KIT cells or empty vector controls. FTY720 treatment caused the dephosphorylation of the D816V c-KIT receptor and its downstream signaling targets pAkt, pSTAT5, and pERK1/2. Additionally, in vivo administration of FTY720 delayed the growth of V560G and D816V c-KIT tumors, inhibited splenic and bone marrow infiltration, and prolonged survival. Our findings show that PP2A inhibition is essential for c-KIT-mediated tumorigenesis, and that reactivating PP2A may offer an attractive strategy to treat drug-resistant c-KIT + cancers. Cancer Res; 70(13); 5438-47. ©2010 AACR.

The role of insulin-like growth factor-binding protein (IGFBP)-5 in human breast cancer cell growth is unclear. We determined the effects of IGFBP-5 expression on the growth of human breast cancer cell lines in vivo and in vitro. Expression of IGFBP-5, both by stable transfection and adenoviral-mediated infection, was inhibitory to the growth of MDA-MB-231 and Hs578T human breast cancer cells over a 13-day period. IGFBP-5 expression resulted in a G 2 /M cell cycle arrest and the induction of apoptosis in both cell lines, an effect that was abrogated in the presence of the broad-spectrum caspase inhibitor, z-VAD-fmk. IGFBP-5-induced apoptosis was associated with a transcriptional increase in expression of the proapoptotic regulator bax and decrease in the anti-apoptotic bcl-2 compared with vector controls. Secreted IGFBP-5 when added exogenously to breast cancer cells was not internalized and had no effect on cell growth or apoptosis, suggesting that IGFBP-5 may elicit its inhibitory effects via a novel, intracrine mechanism. In athymic nude mice, stable expression of IGFBP-5 significantly inhibited both the formation and growth of tumors derived from MDA-MB-231 cells. IGFBP-5-expressing tumors also had a significantly elevated level of bax mRNA and decreased levels of bcl-2 mRNA compared with vector tumors. These data suggest that IGFBP-5 is a potent growth inhibitor and proapoptotic agent in human breast cancer cells via modulation of cell cycle regulation and apoptotic mediators.The insulin-like growth factor-binding proteins (IGFBPs) 1 are a family of proteins that bind with high affinity to IGFs and modulate their mitogenic actions by regulating their ability to interact with their signaling receptor, the type I IGF receptor (IGFRI, reviewed in Ref. 1). However, it is now becoming clear that many IGFBPs have direct roles in the regulation of cell growth and cell death. For example, we and others have demonstrated the antiproliferative and proapoptotic effects of IGFBP-3 in breast (2-4) and prostate (4) cancer cell lines.The role of IGFBP-5 in cell growth is complex, with reports that it can either stimulate or inhibit cell proliferation in various experimental systems (5-7). There is also evidence that IGFBP-5 expression is up-regulated by antiproliferative agents such as retinoic acid (8), vitamin D-related compounds (9), and antiestrogen ICI 182780 (10), with some evidence that it may mediate their growth inhibitory effects (8, 10). Similarly, growth stimulation of human breast cancer cells by estradiol is associated with a down-regulation of IGFBP-5 expression (10), although exogenous IGFBP-5 had no effect on IGF-I-stimulated DNA synthesis in the breast cancer cell line, MCF-7 (11). They demonstrated that addition of exogenous IGFBP-5 to Hs578T cells protects these cells from ceramide-induced apoptosis, suggesting IGFBP-5 may have a survival function in response to apoptotic stimuli (20,21). A similar conclusion was reached by Roschier et al. (22) following their demonstration that induction of apoptosis...