Background and purpose: ABC multidrug transporters (MDR-ABC proteins) cause multiple drug resistance in cancer and may be involved in the decreased anti-cancer efficiency and modified pharmacological properties of novel specifically targeted agents. It has been documented that ABCB1 and ABCG2 interact with several first-generation, small-molecule, tyrosine kinase inhibitors (TKIs), including the Bcr-Abl fusion kinase inhibitor imatinib, used for the treatment of chronic myeloid leukaemia. Here, we have investigated the specific interaction of these transporters with nilotinib, dasatinib and bosutinib, three clinically used, second-generation inhibitors of the Bcr-Abl tyrosine kinase activity. Experimental approach: MDR-ABC transporter function was screened in both membrane-and cell-based (K562 cells) systems. Cytotoxicity measurements in Bcr-Abl-positive model cells were coupled with direct determination of intracellular TKI concentrations by high-pressure liquid chromatography-mass spectrometry and analysis of the pattern of Bcr-Abl phosphorylation. Transporter function in membranes was assessed by ATPase activity. Key results: Nilotinib and dasatinib were high-affinity substrates of ABCG2, and this protein mediated an effective resistance in cancer cells against these compounds. Nilotinib and dasatinib also interacted with ABCB1, but this transporter provided resistance only against dasatinib. Neither ABCB1 nor ABCG2 induced resistance to bosutinib. At relatively higher concentrations, however, each TKI inhibited both transporters. Conclusions and implications: A combination of in vitro assays may provide valuable preclinical information for the applicability of novel targeted anti-cancer TKIs, even in multidrug-resistant cancer. The pattern of MDR-ABC transporter-TKI interactions may also help to understand the general pharmacokinetics and toxicities of new TKIs.

The tight control of proliferation, survival, and apoptosis of bone marrow progenitor cells has a major role in allowing normal hematopoiesis. A variety of cytokines, including GM-CSF 1 and IL-3, regulate viability, proliferation, differentiation, and function of hemopoietic cells (1). Hemopoietic cells undergo apoptotic cell death in the absence of growth factors, whereas an escape from this regulation may result in leukemogenesis.The human GM-CSF/IL-3-dependent myelo-erythroid cell line, TF-1, has been reported to undergo apoptosis upon hormone deprivation (2). Apoptosis of these cells can be suppressed by re-administration of GM-CSF, IL-3, or IL-4 (3) or overexpression of Bcl-2 (4). The alteration of intracellular Ca 2ϩ concentration control cellular processes as diverse as proliferation, development, contraction, or secretion. However, when exceeding its normal spatial and temporal boundaries, Ca 2ϩ can result in cell death through both necrosis and apoptosis (7).The purpose of this study was to delineate how Ca 2ϩ can have multiple roles in cell survival and growth. Ca 2ϩ functions directly through regulation of gene expression, e.g. c-fos (8) or via activating Ca 2ϩ -dependent enzymatic action, e.g. calcium/ calmodulin-dependent kinases II and IV in neurons, calcineurin in lymphocytes, and in conjunction with other signaling pathways such as those regulated through MAPK and PI3K * This work was supported by the Hungarian Research Foundation (Grant OTKA T029291) by the Scientific Research Council, Ministry of Health (Grant ETT 206/2001), and by NRDP 1/024, Ministry of Education, Hungary. 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.ʈ To whom correspondence should be addressed. Tel./Fax: 36-1-372-4353; E-mail: magocsi@biomembrane.hu.1 The abbreviations used are: GM-CSF, granulocyte-macrophage colony-stimulating factor; AP-1, activator protein-1; BAPTA-AM, 1,2-bis(o-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid tetra(acetoxymethyl)ester; [Ca 2ϩ ] i , cytoplasmic free calcium concentration; CPA, cyclopiazonic acid; CREB, cAMP response element binding protein; MAPK, mitogen-activated protein kinase; ERK1/2, extracellular signalregulated kinase; MEK, MAPK/ERK kinase; Egr-1, early growth response protein-1; Elk-1, Ets domain protein-1; p90RSK, 90-kDa ribosomal S6 kinase; PD98059, 2Ј-amino-3-methoxyflavone (MEK-1 inhibitor); UO126, 1,4-diamino-2,3-dicyano-1,4-bis[2-aminophenylthio]-butadiene (MEK1/2 inhibitor); PI3K, phosphoinositide 3-kinase; SRE, serum response element; STAT-5, signal transducer and activator of transcription-5; IL-3, interleukin-3; FACS, fluorescence-activated cell

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