Imatinib Mesylate (STI-571) Reduces Bcr-Abl-Mediated Vascular Endothelial Growth Factor Secretion in Chronic Myelogenous Leukemia John M.L. Ebos, 1,2 Jennifer Tran, 1,3 Zubin Master, 1,2 Daniel Dumont, 1,2 Junia V. Melo, 4 Elisabeth Buchdunger, 5 and Robert S. Kerbel 1,2,3 1 Molecular and Cell Biology Research, Sunnybrook and Women’s College Health Sciences Centre, Toronto, Ontario, Canada M4N 3M5; 2 Department of Medical Biophysics and 3 Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada M4N 3M5; 4 Department of Haematology, Imperial College of Science, Technology and Medicine, Hammersmith Hospital, London, United Kingdom; and 5 Novartis Pharma AG, Oncology Research, Basel, Switzerland Abstract A large and diverse spectrum of oncogenes has been implicated as a contributor to angiogenesis in solid tumors based, in part, on its ability to induce proangio- genic growth factors such as vascular endothelial growth factor (VEGF), and the fact that various anti- oncogenic signaling inhibitor drugs have been shown to reverse such proangiogenic effects both in vitro and in vivo . Because leukemias are now also considered to be angiogenesis-dependent malignancies, we asked whether a similar paradigm might exist for the BCR-ABL oncogene and the Bcr-Abl targeting drug, STI-571 (imatinib mesylate), in the context of chronic myeloge- nous leukemia (CML) cells. We found that levels of VEGF expression in BCR-ABL -positive K562 cells were reduced in vitro by treatment with STI-571 in a dose- dependent fashion. Transfection of BCR-ABL into murine myeloid 32D and human megakaryocyte MO7e hematopoietic cells resulted in enhanced VEGF expres- sion, which could be further elevated by the exposure to cytokines such as interleukin 3 and granulocyte macro- phage colony-stimulating factor. We also found that conditioned media taken from 32D-p210-transfected cells could stimulate human umbilical vein endothelial cells by increasing phosphorylation of VEGF-R2/KDR and the downstream serine/threonine kinase PKB/Akt, an important regulator of endothelial cell survival. Moreover, amplification of BCR-ABL in STI-571-resistant cells was associated with elevated VEGF expression levels which could be reversed by treatment with higher concentrations of STI-571. Taken together, our results implicate BCR-ABL as a possible regulator of CML angiogenesis and raise the possibility that STI-571 could mediate some of its anti-CML properties in vivo through an angiogenesis-dependent mechanism. Introduction The angiogenic switch in tumors is thought to be governed by two major, complementary events: the induction or up- regulation of various proangiogenic growth factors such as vascular endothelial growth factor (VEGF), and the down- regulation or loss of expression of endogenous inhibitors of angiogenesis such as thrombospondin-1 (TSP-1) (1). A grow- ing body of evidence has further implicated both environ- mental/epigenetic alterations, e.g. , tumor hypoxia, as well as genetic changes being responsible for the tumor angiogenic switch (1), and it is thought that such genetic and epigenetic changes may work in a coordinated manner to stimulate an optimal angiogenic response (2). Relevant genetic changes include oncogene activation (3) and tumor suppressor gene mutation/inactivation events (e.g. , loss of p53, PTEN, or VHL function), both of which can strongly influence the expression of proangiogenic growth factors and angiogenesis inhibitors (1, 4). With respect to oncogenes, we (5) and Grugel et al. (6) first showed in 1995 that activated K- or H-ras was a powerful inducer of VEGF expression in transformed rat intestinal epithelial cells, human colorectal carcinoma cells, or mouse 3T3 fibroblasts. Since then, a broad spectrum of other (proto)- oncogenes, including ERBB-2/HER-2 , the epidermal growth factor receptor (EGFR ) tyrosine kinase, and MYC , among many others, has been shown to cause similar changes (3), including the down-regulation of TSP-1 (1). An important therapeutic implication of such findings is that many of the anti-cancer signal transduction inhibitor drugs, originally designed to block the activity of specific oncopro- teins, may additionally elicit antiangiogenic effects in vivo , which could conceivably contribute to their overall antitumor efficacy. Indeed, evidence in support of this possibility was first reported by us with respect to Ras farnesyltransferase inhibitors (Ras, FTIs) (5), Erbitux/C225, an EGFR blocking antibody (7), and Herceptin, the ErbB-2 blocking monoclonal antibody (7) as well as by other groups (8 – 10). An intriguing possibility that arises as a result of such findings is that the drug STI-571 (imatinib mesylate) that Received 09/09/02; revised 10/04/02; accepted 10/14/02. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Grant support: Studentships from the Canadian Institute for Health Research (CIHR) and National Cancer Institute of Canada (NCIC) (to Z.M. and J.T, respectively); CIHR (to R.S.K.). Requests for reprints: Robert S. Kerbel, Molecular and Cell Biology Research, Sunnybrook and Women’s College Health Sciences Centre, S-218 Research Building, 2075 Bayview Avenue, Toronto, Ontario, Canada M4N 3M5. Phone: (416) 480-5711; Fax: (416) 480-5703. E-mail: Robert.Kerbel@swchsc.on.ca Copyright D 2002 American Association for Cancer Research. 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