ABSTRACT
Focal adhesion kinase (FAK) activity contributes to many advanced cancer phenotypes, but little is known about its role in human acute myeloid leukemia (AML). Here, we show that FAK splice variants are abnormally expressed in the primitive leukemic cells of poor prognosis AML patients. In the CD34(+) 38(-) 123(+) long-term culture-initiating cell-enriched leukemic cells of these patients, FAK upregulates expression of Frizzled-4 and phosphorylates Pyk2 to enable the required association of Pyk2 with the Wnt5a/Frizzled-4/LRP5 endocytosis complex and downstream activation of ß-catenin, thereby replacing the Wnt3a-controlled canonical pathway used by normal hematopoietic stem cells. Transduction of primitive normal human hematopoietic cells with FAK splice variants induces a marked increase in their clonogenic activity and signaling via the Wnt5a-controlled canonical pathway. Targeting FAK or ß-catenin efficiently eradicates primitive leukemic cells in vitro suggesting that FAK could be a useful therapeutic target for improved treatment of poor prognosis AML cases.
Subject(s)
Focal Adhesion Protein-Tyrosine Kinases/metabolism , Leukemia, Myeloid, Acute/metabolism , Wnt Proteins/metabolism , Aged , Antigens, CD34/biosynthesis , Antigens, CD34/genetics , Antigens, CD34/metabolism , Disease-Free Survival , Female , Focal Adhesion Protein-Tyrosine Kinases/biosynthesis , Focal Adhesion Protein-Tyrosine Kinases/genetics , Frizzled Receptors/metabolism , Humans , Isoenzymes , Leukemia, Myeloid, Acute/enzymology , Leukemia, Myeloid, Acute/genetics , Low Density Lipoprotein Receptor-Related Protein-6/metabolism , Male , Middle Aged , Phosphorylation , Proto-Oncogene Proteins/metabolism , RNA, Small Interfering/administration & dosage , RNA, Small Interfering/genetics , Signal Transduction , Survival Analysis , Transcriptional Activation , Transfection , Wnt Proteins/genetics , Wnt-5a Protein , beta Catenin/biosynthesis , beta Catenin/genetics , beta Catenin/metabolismABSTRACT
In search for compounds able to reduce cell adhesion-mediated drug resistance (CAM-DR), we studied effects of Hammada scoparia extracts on leukemic cells adherent or in suspension. We show that H. scoparia flavonoidic fraction and its compound rutin induce apoptosis specifically in adherent leukemic cells and abolish CAM-DR. Importantly, rutin inhibited survival of adherent leukemic progenitors (CD34(+)38(-)123(+)) but spared normal progenitors (CD34(+)38(-)). The pro-apoptotic effects of rutin were correlated with a decrease of active GSK3ß and inhibitors of GSK3ß reproduced rutin-induced cytotoxicity. This study uncovers the potential of H. scoparia flavonoids and rutin to overcome CAM-DR in acute myeloid leukemia.
Subject(s)
Apoptosis/drug effects , Cell Adhesion/drug effects , Drug Resistance, Neoplasm/drug effects , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Phytotherapy , Rutin/pharmacology , Scoparia/chemistry , Antigens, CD34/metabolism , Blotting, Western , Cells, Cultured , Flavonoids/pharmacology , Flow Cytometry , Glycogen Synthase Kinase 3/antagonists & inhibitors , Glycogen Synthase Kinase 3/metabolism , Glycogen Synthase Kinase 3 beta , Hematopoietic Stem Cells/drug effects , Hematopoietic Stem Cells/metabolism , Humans , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
BACKGROUND: Cell survival mediated by integrin engagement has been implicated in cell adhesion-mediated drug resistance. We have recently demonstrated that the activation of glycogen synthase kinase 3 beta (GSK3beta) is a new pathway supporting the chemoresistance of leukemic cells adhered to fibronectin. METHODOLOGY AND PRINCIPAL FINDINGS: We show here that in conditions of serum starvation, the fibronectin receptor alpha(5)beta(1) integrin, but not alpha(4)beta(1), induced activation of GSK3beta through Ser-9 dephosphorylation in adherent U937 cells. The GSK3beta-dependent survival pathway occurred in adherent leukemic cells from patients but not in the HL-60 and KG1 cell lines. In adhesion, activated GSK3beta was found in the cytosol/plasma membrane compartment and was co-immunoprecipitated with alpha(5) integrin, the phosphatase PP2A and the scaffolding protein RACK1. PP2A and its regulatory subunit B' regulated the Ser-9 phosphorylation of GSK3beta. In adherent leukemic cells, alpha(5)beta(1) integrin but not alpha(4)beta(1) upregulated the resistance to TNFalpha-induced apoptosis. Both extrinsic and intrinsic apoptotic pathways were under the control of alpha(5)beta(1) and GSK3beta. CONCLUSIONS AND SIGNIFICANCE: Our data show that, upon serum starvation, alpha(5)beta(1) integrin engagement could regulate specific pro-survival functions through the activation of GSK3beta.
