Expression profile of WNT, FZD and sFRP genes in human hematopoietic cells.

Identifying gene expression differences in the Wnt signaling pathway specific to leukemic cells can be hampered by the lack of verified knowledge on the expression of WNT genes in normal blood cells. In this study we aimed to determine the expression profile of human WNT, FZD and sFRP genes in normal adult bone marrow, T and B cells; along with the hematopoietic cell lines K562, HL60, Jurkat and Namalwa. Bone marrow and peripheral blood from 16 donors were evaluated and our results were compared with the GeneNote database expression arrays. In bone marrow, only WNT3, WNT10A, FZD3, FZD7 and sFRP1 genes are constitutively expressed. Lymphocytes express WNT7A, WNT9B, FZD6 and FZD7 in addition to the genes above, but T cells differ in that they lose sFRP1 expression and gain constitutive expression of WNT16. An established "normal" profile of the Wnt genes in various blood cells will provide a fundamental basis for research investigating hematopoiesis and cellular processes during leukemic transformation. (C) 2010 Elsevier Ltd. All rights reserved

sFRP1 promoter methylation is associated with persistent Philadelphia chromosome in chronic myeloid leukemia.

Epigenetic silencing of sFRP genes has been shown to lead to constitutive activation of the canonical Wnt-signaling pathway. The first description of deregulated Wnt-signaling activation in a hematological malignancy was reported in chronic myeloid leukemia (CML). To investigate whether epigenetic silencing of sFRP is responsible for the observed Wnt activation in CML, we studied the methylation and mutational status of the sFRP1 promoter in 48 chronic phase CML patients. Of the 48 CML patients 41 were shown to be unmethylated, 6 patients hemi-methylated and 1 patient fully methylated at the sFRP1 promoter. Albeit observed infrequently in chronic phase CML, sFRP1 promoter methylation correlated with primary cytogenetic resistance to imatinib mesylate. sFRP1 promoter methylation may indicate a genetically more unstable form of disease resistant to therapy and provide a key biological difference in therapy resistant patients, in addition to a possible mechanism for the observed activation of canonical Wnt signaling in CML. (C) 2008 Elsevier Ltd. All rights reserved

Beta-catenin mutations are not observed in chronic myeloid leukemia.

Aims and background. Studies reporting activated Wnt signaling in all stages of chronic myeloid leukemia (CML) have demonstrated that deregulation of the pathway plays a role in the pathogenesis of this disease. Several reports have suggested mechanisms for the deregulated Wnt signaling and beta-catenin stabilization observed in CML. One possible mechanism for beta-catenin stabilization could be the acquisition of mutations at its N-terminal domain, especially in the third exon where it is marked via phosphorylation for degradation. We sought to determine whether mutations in the third exon of the beta-catenin gene are responsible for the observed Wnt activation in CM

Dvl proteins regulate SMAD1, AHR, mTOR, BRD7 protein expression while differentially regulating canonical and non-canonical Wnt signaling pathways in CML cell lines

Dishevelled (Dvl) is a scaffold protein that transmits Wnt signals to downstream effectormolecules via both canonical and non-canonical Wnt signaling pathways. Deregulatedactivation of Dvl proteins has been reported in various solid tumors. However, it is not clearwhich pathway and proteins are responsible for observed aberrant activities and their relevancein disease prognosis. In addition, there is relatively limited knowledge on the role Dvl proteinsmay have in hematologic malignancy etiopathogenesis. In this study, we demonstrated that Dvlgenes are not expressed in normal bone marrow but are expressed at different levels in the bonemarrow of patients with chronic myeloid leukemia. We showed SMAD1, AHR, mTOR, BRD7protein expressions are significantly affected by Dvl silencing and overexpression in CML celllines. Wnt/-catenin and Wnt/PCP signaling pathway components are effectively repressedafter Dvl silencing in K562 cells, while regulator of Wnt/Ca2+ signaling showed increase inboth CML cell lines. Targeting Dvl proteins increases imatinib susceptibility of the K562 andMEG-01 cell lines. In light of our data, Dvl could be a potential therapeutic target in thetreatment of CML.</p

Induction of apoptosis increases expression of non-canonical WNT genes in myeloid leukemia cell lines

With the aim of determining the differential expression of WNT and FZD genes, before and after induction of apoptosis in BCR-ABL positive cells, we treated the myeloid cell line K562 and control cell line HL60 with imatinib mesylate and etoposide, and analyzed relative mRNA expression levels of WNT, FZD and sFRP genes under normal and apoptotic conditions by real-time RT-PCR. We observed marked increase in mRNA levels of FZD4, FZD5, FZD7 and WNT5b, correlating with apoptotic activity and independent of the agent or cell line used. Our results suggest the involvement of non-canonical Wnt signaling in executing programmed cell death in myeloid cell lines

Secreted Wnt antagonists in leukemia: A road yet to be paved

Wnt signaling has been a topic of research for many years for its diverse and fundamental functions in physiological (such as embryogenesis, organogenesis, proliferation, tissue repair and cellular differentiation) and pathological (carcinogenesis, congenital/genetic diseases, and tissue degeneration) processes. Wnt signaling pathway aberrations are associated with both solid tumors and hematological malignancies. Unregulated Wnt signaling observed in malignancies may be due to a wide spectrum of abnormalities, from mutations in the genes of key players to epigenetic modifications of Wnt antagonists. Of these, Wnt antagonists are gaining significant attention for their potential of being targets for treatment and inhibition of Wnt signaling. In this review, we discuss and summarize the significance of Wnt signaling antagonists in the pathogenesis and treatment of hematological malignancies

Forced expression of Wnt antagonists sFRP1 and WIF1 sensitizes chronic myeloid leukemia cells to tyrosine kinase inhibitors

Chronic myeloid leukemia is a clonal myeloproliferative disorder that arises from the neoplastic transformation of the hematopoietic stem cell, in which the Wnt/-catenin signaling pathway has been demonstrated to play an important role in disease progression. However, the role of Wnt signaling antagonists in therapy resistance and disease progression has not been fully investigated. We aimed to study the effects of Wnt/-catenin pathway antagonistssecreted frizzled-related protein 1 and Wnt inhibitory factor 1on resistance toward tyrosine kinase inhibitors in chronic myeloid leukemia. Response to tyrosine kinase inhibitors was analyzed in secreted frizzled-related protein 1 and Wnt inhibitory factor 1 stably transfected K562 cells. Experiments were repeated using a tetracycline-inducible expression system, confirming previous results. In addition, response to tyrosine kinase inhibitor treatment was also analyzed using the secreted frizzled-related protein 1 expressing, BCR-ABL positive MEG01 cell line, in the presence and absence of a secreted frizzled-related protein 1 inhibitor. Our data suggests that total cellular -catenin levels decrease in the presence of secreted frizzled-related protein 1 and Wnt inhibitory factor 1, and a significant increase in cell death after tyrosine kinase inhibitor treatment is observed. On the contrary, when secreted frizzled-related protein 1 is suppressed, total -catenin levels increase in the cell and the cells become resistant to tyrosine kinase inhibitors. We suggest that Wnt antagonists carry the potential to be exploited in designing new agents and strategies for the advanced and resistant forms of chronic myeloid leukemia

Forced expression of Wnt antagonists sFRP1 and WIF1 sensitizes chronic myeloid leukemia cells to tyrosine kinase inhibitors

Chronic myeloid leukemia is a clonal myeloproliferative disorder that arises from the neoplastic transformation of the hematopoietic stem cell, in which the Wnt/-catenin signaling pathway has been demonstrated to play an important role in disease progression. However, the role of Wnt signaling antagonists in therapy resistance and disease progression has not been fully investigated. We aimed to study the effects of Wnt/-catenin pathway antagonistssecreted frizzled-related protein 1 and Wnt inhibitory factor 1on resistance toward tyrosine kinase inhibitors in chronic myeloid leukemia. Response to tyrosine kinase inhibitors was analyzed in secreted frizzled-related protein 1 and Wnt inhibitory factor 1 stably transfected K562 cells. Experiments were repeated using a tetracycline-inducible expression system, confirming previous results. In addition, response to tyrosine kinase inhibitor treatment was also analyzed using the secreted frizzled-related protein 1 expressing, BCR-ABL positive MEG01 cell line, in the presence and absence of a secreted frizzled-related protein 1 inhibitor. Our data suggests that total cellular -catenin levels decrease in the presence of secreted frizzled-related protein 1 and Wnt inhibitory factor 1, and a significant increase in cell death after tyrosine kinase inhibitor treatment is observed. On the contrary, when secreted frizzled-related protein 1 is suppressed, total -catenin levels increase in the cell and the cells become resistant to tyrosine kinase inhibitors. We suggest that Wnt antagonists carry the potential to be exploited in designing new agents and strategies for the advanced and resistant forms of chronic myeloid leukemia