Background and aims: GSK-3 is a serine-threonine kinase involved in metabolic regulation as well as in the control of many pathways associated to cancer development, including Notch Wnt/b-catenin, Hedgehog, and AKT. Association of GSK-3 inhibitors with All- trans-retinoic acid (ATRA) significantly improved ATRA-medi- ated differentiation and cell death of acute promyelocytic leukemia cells. However, little is currently known about the contribution of GSK-3 role to non-promyelocytic AML cell response to treatment with chemoterapeutic agents. Methods: GSK-3 expression was analyzed by Western blot or flow cytometry inAML cell lines (HL-60, THP1, U937) or primary non-promyelocytic AML blast cells (30 samples). AML cellscul- tured alone or in presence ofhuman bone marrow mesenchymal stromal cells (hBM-MSCs)were treated with GSK-3 inhibitors, including LiCL, AR-A014418, SB 216763, in association or not with Cytarabine (Ara-C) or Idarubicin. Cell proliferation and cell death were measured by CFSE dilution and TOPRO-3/Annexin-V stain- ing, respectively. Results: Flow cytometry and Western blot analysis in AML sam- ples revealed highexpression levels of all GSK-3 forms, including total GSK-3α, (Ser21) GSK-3α, total GSK-3b, and (Ser 21) GSK- 3b; theseforms were all down-modulated when AML cells were cul- tured in presence of hBM-MSCs, thus suggesting that GSK-3 plays an important role intransducting microenvironmental signals in AML cells interacting with bone marrow stroma. The treatment of AML cells with increasing concentrations of each GSK-3 inhibitors decreased AML cell viability in a dose-dependent manner; inter- estingly,hBM-MSCsor peripheral blood mononuclear cells were less sensitive to GSK-3inhibitors. The addition of each inhibitor increased dramatically the AML cell apoptotic rate induced by the addition of Ara-C or Idarubicinin vitro. Notably, LiCl and AR- A014418 were capable of abrogatinghBM-MSC-mediated AML cell resistance to apoptosis induced by Ara-C or Idarubicin. Conclusion: Our data clearly demonstrate that GSK-3 inhibition may improve the efficacy of some chemoterapeutic agents used in the treatment of non-promyelocytic AML.
Role of GSK-3 signalling in non-promyelocitic acute myeloid leukemia (AML) cell response to chemotherapy
Takam Kamga, Paul;Cassaro, Adriana;BASSI, Giulio;DAL COLLO, GIADA;ADAMO, ANNALISA;Gatti, Alessandro;MIDOLO, MARTINA;Carusone, Roberta;Di Trapani, Mariano;RESCI, FEDERICA;BONIFACIO, Massimiliano;KRAMPERA, Mauro
2016-01-01
Abstract
Background and aims: GSK-3 is a serine-threonine kinase involved in metabolic regulation as well as in the control of many pathways associated to cancer development, including Notch Wnt/b-catenin, Hedgehog, and AKT. Association of GSK-3 inhibitors with All- trans-retinoic acid (ATRA) significantly improved ATRA-medi- ated differentiation and cell death of acute promyelocytic leukemia cells. However, little is currently known about the contribution of GSK-3 role to non-promyelocytic AML cell response to treatment with chemoterapeutic agents. Methods: GSK-3 expression was analyzed by Western blot or flow cytometry inAML cell lines (HL-60, THP1, U937) or primary non-promyelocytic AML blast cells (30 samples). AML cellscul- tured alone or in presence ofhuman bone marrow mesenchymal stromal cells (hBM-MSCs)were treated with GSK-3 inhibitors, including LiCL, AR-A014418, SB 216763, in association or not with Cytarabine (Ara-C) or Idarubicin. Cell proliferation and cell death were measured by CFSE dilution and TOPRO-3/Annexin-V stain- ing, respectively. Results: Flow cytometry and Western blot analysis in AML sam- ples revealed highexpression levels of all GSK-3 forms, including total GSK-3α, (Ser21) GSK-3α, total GSK-3b, and (Ser 21) GSK- 3b; theseforms were all down-modulated when AML cells were cul- tured in presence of hBM-MSCs, thus suggesting that GSK-3 plays an important role intransducting microenvironmental signals in AML cells interacting with bone marrow stroma. The treatment of AML cells with increasing concentrations of each GSK-3 inhibitors decreased AML cell viability in a dose-dependent manner; inter- estingly,hBM-MSCsor peripheral blood mononuclear cells were less sensitive to GSK-3inhibitors. The addition of each inhibitor increased dramatically the AML cell apoptotic rate induced by the addition of Ara-C or Idarubicinin vitro. Notably, LiCl and AR- A014418 were capable of abrogatinghBM-MSC-mediated AML cell resistance to apoptosis induced by Ara-C or Idarubicin. Conclusion: Our data clearly demonstrate that GSK-3 inhibition may improve the efficacy of some chemoterapeutic agents used in the treatment of non-promyelocytic AML.
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