ORIGINAL ARTICLE Autocrine insulin-like growth factor-I signaling promotes growth and survival of human acute myeloid leukemia cells via the phosphoinositide 3-kinase/Akt pathway KT Doepfner 1 , O Spertini 2 and A Arcaro 1 1 Division of Clinical Chemistry and Biochemistry, University Children’s Hospital Zurich, Zurich, Switzerland and 2 Service and Central Laboratory of Hematology, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland Insulin-like growth factor (IGF) signaling plays an important role in various human cancers. Therefore, the role of insulin- like growth factor I (IGF-I) signaling in growth and survival of acute myeloid leukemia (AML) cells was investigated. Expres- sion of the IGF-I receptor (IGF-IR) and its ligand IGF-I were detected in a panel of human AML blasts and cell lines. IGF-I and insulin promoted the growth of human AML blasts in vitro and activated the phosphoinositide 3-kinase (PI3K)/Akt and the extracellular signal-regulated kinase (Erk) pathways. IGF-I- stimulated growth of AML blasts was blocked by an inhibitor of the PI3K/Akt pathway. Moreover, downregulation of the class Ia PI3K isoforms p110b and p110d by RNA interference impaired IGF-I-stimulated Akt activation, cell growth and survival in AML cells. Proliferation of a panel of AML cell lines and blasts isolated from patients with AML was inhibited by the IGF-IR kinase inhibitor NVP-AEW541 or by an IGF-IR neutraliz- ing antibody. In addition to its antiproliferative effects, NVP- AEW541 sensitized primary AML blasts and cell lines to etoposide-induced apoptosis. Together, our data describe a novel role for autocrine IGF-I signaling in the growth and survival of primary AML cells. IGF-IR inhibitors in combination with chemotherapeutic agents may represent a novel approach to target human AML. Leukemia (2007) 21, 1921–1930; doi:10.1038/sj.leu.2404813; published online 21 June 2007 Keywords: acute myeloid leukemia; insulin-like growth factor; phosphoinositide 3-kinase; Akt, apoptosis Introduction Acute myeloid leukemia (AML) accounts for approximately 70–80% of acute leukemia in adults. Treatment and outcome of the disease depend on several factors, including leukemia karyotype, molecular alteration and patient age. The 5-year survival rate ranges from 65 to 15%, drastically decreasing with the age at diagnosis. Chromosomal translocations frequently result in dysfunction of transcription factors needed for normal hematopoietic development. Additional mutations have been described in the receptor tyrosine kinases FLT3, c-Kit and c-Fms, as well as in N- and K-Ras. 1,2 The manifestation of AML is a combination of mutations conferring proliferative advantage, impaired differentiation and apoptosis. Polypeptide growth factors have been shown to play a key role in AML proliferation and survival. 3–5 Human AML cells express a variety of growth factor and cytokine receptors that can be activated by mutation, overexpression and/or establish- ment of autocrine loops. Among these receptors are the polypeptide growth factor receptors FLT3, c-Kit, c-Fms, vascular endothelial growth factor receptor and fibroblast growth factor receptor. 3–8 Several potential anti-AML therapeutic approaches involving the FLT3 system have been reported. 9–11 Insulin-like growth factor (IGF) signaling plays a major role in various human malignancies, including breast, colon and prostate cancer. 12 In leukemia, IGF signaling has not yet been extensively studied, although expression of the IGF-I receptor (IGF-IR) was reported in human AML cells. 13,14 Autocrine IGF-I production has been suggested to play a role in drug resistance in an AML cell line. 15 In addition, IGF-I signaling has a crucial function in other hematological malignancies such as multiple myeloma, 16 and several anti-IGF-IR experimental therapies were shown to inhibit multiple myeloma proliferation in vitro and in vivo. 17 A critical intracellular signaling mediator of the IGF-IR is the phosphoinositide 3-kinase (PI3K)/Akt pathway. 18,19 Indeed, PI3K signaling is implicated in the control of cell proliferation, survival and motility/metastasis downstream of many different growth factor receptors. 20 The importance of PI3K signaling in human cancer is highlighted by the fact that mutations in the tumor suppressor gene PTEN occur frequently in human tumors. 20,21 PTEN is a phosphatase that antagonizes the action of PI3K by dephosphorylating the D-3 position of polyphos- phoinositides. 22 Moreover, recent reports have described activating mutations in the PIK3CA gene encoding the catalytic p110a isoform of class I A PI3K in a variety of human cancers, including, breast, colon and ovarian cancer. 23,24 Mutations in the PTEN gene have not been found in a high percentage of AML cases, although they were documented in AML cell lines. 21,25 Moreover, a recent screen comprising AML cases did not reveal any mutations in the gene encoding p110a, 26 suggesting a possible deregulation of other class I A PI3Ks, namely p110b and p110d in hematological malignancies. In support of this notion, constitutive activation of Akt/PKB has been reported by several studies in human AML blasts. 27,28 In the context of leukemia, altered PI3K signaling was also shown to play a role in the development of adult T-cell lymphoma. 29 In the present report, we have investigated the expression pattern and biological functions of components of the IGF-IR signaling system in human AML blasts and cell lines. Moreover, we have evaluated the potential of the novel IGF-IR kinase inhibitor NVP-AEW541 30 as an antitumor agent in AML. Finally, we have investigated whether targeting downstream signaling mediators of the IGF-IR could suppress growth and induce apoptosis in AML cell lines. Our findings describe for the first time a role for autocrine signaling by IGF-I and the IGF-IR in growth, survival and chemoresistance of AML cells, which involves the PI3K/Akt pathway. Received 31 January 2007; revised 23 May 2007; accepted 23 May 2007; published online 21 June 2007 Correspondence: Dr A Arcaro, Division of Clinical Chemistry and Biochemistry, University Children’s Hospital Zurich, Steinwiesstrasse 75, Zurich CH-8032, Switzerland. E-mail: Alexandre.Arcaro@kispi.unizh.ch Research Support: Krebsliga Zu ¨ rich. Leukemia (2007) 21, 1921–1930 & 2007 Nature Publishing Group All rights reserved 0887-6924/07 $30.00 www.nature.com/leu