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HSP70 AND HSF1 GO HAND IN HAND AND HAVE A ROLE IN THE SURVIVAL OF CHRONIC LYMPHOCYTIC LEUKEMIA NEOPLASTIC B CELLS.
Author(s):
Flavia Raggi
,
Flavia Raggi
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Frezzato Federica
,
Frezzato Federica
Affiliations:
VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy;Dipartimento di Medicina,Università di Padova,Padova,Italy
Veronica Martini
,
Veronica Martini
Affiliations:
VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy;Dipartimento di Medicina,Università di Padova,Padova,Italy
Filippo Severin
,
Filippo Severin
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Valentina Trimarco
,
Valentina Trimarco
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Leonardo Martinello
,
Leonardo Martinello
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Andrea Visentin
,
Andrea Visentin
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Edoardo Scomazzon
,
Edoardo Scomazzon
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy
Silvia Imbergamo
,
Silvia Imbergamo
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy
Monica Facco
,
Monica Facco
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Francesco Piazza
,
Francesco Piazza
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Gianpietro Semenzato
,
Gianpietro Semenzato
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
Livio Trentin
Livio Trentin
Affiliations:
Dipartimento di Medicina,Università di Padova,Padova,Italy;VIMM Istituto Veneto di Medicina Molecolare,Padova,Italy
(Abstract release date: 05/18/17) EHA Library. Trentin L. 05/18/17; 182484; PB1770
Prof. Livio Trentin
Prof. Livio Trentin
Contributions
PDF Abstract
Abstract

Abstract: PB1770

Type: Publication Only

Background

B-cell Chronic Lymphocytic Leukemia (CLL) is a neoplastic disorder characterized by the accumulation of clonal B cells in peripheral blood, bone marrow and lymphoid tissues. CLL is a clinically and biologically heterogeneous disease. As a consequence, novel biological and cytogenetic features have become increasingly important in predicting prognosis at the time of diagnosis and the research for molecules involved in apoptosis resistance and increased survival of neoplastic B cells is still ongoing.

Aims

We recently found that the Heat Shock Protein of 70kDa (HSP70) is overexpressed in Chronic Lymphocytic Leukemia (CLL) B cells. Considering the pro-survival role of HSP70 in cancer, we were aimed at characterizing this protein and its master regulator, the Heat Shock Factor 1 (HSF1), within the pathogenetic mechanisms leading to CLL.

Methods

HSP70 and HSF1 expression levels were evaluated by Western blotting (WB) analysis in leukemic and normal B cells. HSP70 and HSF1 protein levels were correlated to IGHV mutational status and ZAP70 protein expression in CLL patients. HSP70 and HSF1 levels were also analyzed in neoplastic cells obtained from patients undergoing Ibrutinib based regimen by WB analysis. Moreover, HSP70 and HSF1 localization was analyzed by subcellular protein fractionation followed by WB analysis. The effects of HSP70 and HSF1 inhibition (by Zafirlukast and Fisetin) were evaluated by Annexin V/Propidium Iodide flow cytometry test and WB analysis of PARP cleavage.

Results

We demonstrated that HSP70 and HSF1 are overexpressed in leukemic vs normal B cells and their expression levels correlate to poor prognosis in CLL. We also analyzed HSP70 and HSF1 levels in patients following in vivo Ibrutinib based regimen, observing a positive correlation between these two protein expression levels and moreover we observed that these two protein levels decreased after therapy. We found that at steady state both HSP70 and HSF1 are localized in the nucleus of CLL B cells. HSP70 and HSF1 inhibition was proved to be effective in inducing a dose-dependent in vitro apoptosis of CLL B cells.

Conclusion
HSP70 and HSF1 overexpression and correlation with poor prognosis in CLL patients underline their pivotal role in the regulation of leukemic B cell survival. HSP70 and HSF1 both correlation and reduction in CLL patients following in vivo Ibrutinib regimen let us hypothesize a role of these proteins in the progression of the disease. In normal B cells HSP70 and HSF1 are both localized into the nucleus after stress conditions, however we found both HSP70 and HSF1 localized into the nucleus of CLL B cells at steady state, suggesting a constitutive activation of these proteins in CLL. Although HSP70 has been extensively linked to cancer, little progresses have been made in bringing HSP70 inhibitors to the clinic, because of their potential off-target effects. For this reason we tried an alternative approach by targeting the HSP70 major regulator, HSF1. We observed that both inhibitors, Zafirlukast and Fisetin, lead to an in vitro dose dependent B cell apoptosis. These data demonstrate HSP70 and HSF1 involvement in the pathogenesis of CLL and identify HSP70/HSF1 axis as a target for new therapeutic strategies.

Session topic: 5. Chronic lymphocytic leukemia and related disorders - Biology

Keyword(s): Heat shock protein, Chronic Lymphocytic Leukemia

Abstract: PB1770

Type: Publication Only

Background

B-cell Chronic Lymphocytic Leukemia (CLL) is a neoplastic disorder characterized by the accumulation of clonal B cells in peripheral blood, bone marrow and lymphoid tissues. CLL is a clinically and biologically heterogeneous disease. As a consequence, novel biological and cytogenetic features have become increasingly important in predicting prognosis at the time of diagnosis and the research for molecules involved in apoptosis resistance and increased survival of neoplastic B cells is still ongoing.

Aims

We recently found that the Heat Shock Protein of 70kDa (HSP70) is overexpressed in Chronic Lymphocytic Leukemia (CLL) B cells. Considering the pro-survival role of HSP70 in cancer, we were aimed at characterizing this protein and its master regulator, the Heat Shock Factor 1 (HSF1), within the pathogenetic mechanisms leading to CLL.

Methods

HSP70 and HSF1 expression levels were evaluated by Western blotting (WB) analysis in leukemic and normal B cells. HSP70 and HSF1 protein levels were correlated to IGHV mutational status and ZAP70 protein expression in CLL patients. HSP70 and HSF1 levels were also analyzed in neoplastic cells obtained from patients undergoing Ibrutinib based regimen by WB analysis. Moreover, HSP70 and HSF1 localization was analyzed by subcellular protein fractionation followed by WB analysis. The effects of HSP70 and HSF1 inhibition (by Zafirlukast and Fisetin) were evaluated by Annexin V/Propidium Iodide flow cytometry test and WB analysis of PARP cleavage.

Results

We demonstrated that HSP70 and HSF1 are overexpressed in leukemic vs normal B cells and their expression levels correlate to poor prognosis in CLL. We also analyzed HSP70 and HSF1 levels in patients following in vivo Ibrutinib based regimen, observing a positive correlation between these two protein expression levels and moreover we observed that these two protein levels decreased after therapy. We found that at steady state both HSP70 and HSF1 are localized in the nucleus of CLL B cells. HSP70 and HSF1 inhibition was proved to be effective in inducing a dose-dependent in vitro apoptosis of CLL B cells.

Conclusion
HSP70 and HSF1 overexpression and correlation with poor prognosis in CLL patients underline their pivotal role in the regulation of leukemic B cell survival. HSP70 and HSF1 both correlation and reduction in CLL patients following in vivo Ibrutinib regimen let us hypothesize a role of these proteins in the progression of the disease. In normal B cells HSP70 and HSF1 are both localized into the nucleus after stress conditions, however we found both HSP70 and HSF1 localized into the nucleus of CLL B cells at steady state, suggesting a constitutive activation of these proteins in CLL. Although HSP70 has been extensively linked to cancer, little progresses have been made in bringing HSP70 inhibitors to the clinic, because of their potential off-target effects. For this reason we tried an alternative approach by targeting the HSP70 major regulator, HSF1. We observed that both inhibitors, Zafirlukast and Fisetin, lead to an in vitro dose dependent B cell apoptosis. These data demonstrate HSP70 and HSF1 involvement in the pathogenesis of CLL and identify HSP70/HSF1 axis as a target for new therapeutic strategies.

Session topic: 5. Chronic lymphocytic leukemia and related disorders - Biology

Keyword(s): Heat shock protein, Chronic Lymphocytic Leukemia

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