ABSTRACT
PURPOSE: HSP90 targeting is a promising therapeutic approach in cancer. 17-AAG is an HSP90 inhibitor with completed Phase I trials in patients with advanced cancer and recently published Phase II trials. The aim of this work was to study the expression of HSP 90 and apoptotic proteins, the effects in culture of 17-AAG on cell survival and apoptosis and to compare Philadelphia-positive (Ph+) ALL to common B cell ALL, in ALL cell lines and in patients' cells collected at ALL diagnosis. METHODS: We analysed 2 ALL cell lines and 63 leukaemic samples from patients treated in our institution (44 common B cell ALL and 19 Ph+ ALL). We performed flow cytometry analysis of bone marrow aspiration and cell lines with a combination of anti-HSP90, Bax, Bcl-2 and Bcl-xl antibodies. Apoptosis after cell culture (in presence or not of 17-AAG) was assessed using Annexin V and activated caspase-3 staining. RESULTS: Ph+ ALL cells appeared to be more sensitive to 17-AAG cytotoxicity with a 100 % mortality rate after exposure to 10 µM for 24 h (vs. 62 % for B-common ALL). A high percentage of HSP90-positive cells (in Ph+ ALL samples) was associated with high sensitivity to 17-AAG. 17-AAG induced apoptosis in a dose-dependent manner and was associated with down-regulation of Bcl-2 and Bcl-Xl expression and up-regulation of Bax expression. CONCLUSION: Considering that Bcr-Abl constitutes HSP 90 substrates, HSP 90 inhibition could be of particular interest for Ph+ ALL disease, even in patients harbouring resistance to tyrosine kinase inhibitor therapy.
Subject(s)
Apoptosis/drug effects , Benzoquinones/pharmacology , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Lactams, Macrocyclic/pharmacology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Adolescent , Adult , Aged , Aged, 80 and over , Apoptosis/genetics , B-Lymphocytes/drug effects , B-Lymphocytes/metabolism , Caspase 3/genetics , Caspase 3/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Child , Child, Preschool , Flow Cytometry , HSP90 Heat-Shock Proteins/genetics , HSP90 Heat-Shock Proteins/metabolism , Humans , Infant , Middle Aged , Molecular Targeted Therapy/methods , Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics , Precursor Cell Lymphoblastic Leukemia-Lymphoma/metabolism , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Young Adult , bcl-2-Associated X Protein/genetics , bcl-2-Associated X Protein/metabolism , bcl-X Protein/genetics , bcl-X Protein/metabolismSubject(s)
Myelodysplastic Syndromes/metabolism , Stromal Cells/metabolism , Adult , Aged , Caspase 3 , Caspases/metabolism , Cell Division , Cells, Cultured , Cytokines/biosynthesis , Endothelial Growth Factors/biosynthesis , Female , Humans , Intercellular Signaling Peptides and Proteins/biosynthesis , Lymphokines/biosynthesis , Male , Middle Aged , Myelodysplastic Syndromes/diagnosis , Myelodysplastic Syndromes/pathology , Platelet Endothelial Cell Adhesion Molecule-1/analysis , Stromal Cells/chemistry , Stromal Cells/pathology , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
Excessive apoptosis is implicated in the pathogenesis of myelodysplastic syndromes (MDS). We assessed by flow cytometry the expression of several members of the Bcl-2 family in bone marrow mononuclear cells (BMMNC) of 168 MDS samples at diagnosis. The proteins studied were Bcl-2, Bcl-xL (anti-apoptotic), Bax, Bad, Bak, and Bcl-xS (pro-apoptotic). The percentage of BMMNC expressing Bcl-2 and Bcl-xL was higher in refractory anemia with excess of blasts (RAEB), RAEB in transformation (RAEB-T), and chronic myelomonocytic leukemia (CMML) than in refractory anemia (RA) and RA with ringed sideroblasts (RAS). Conversely pro-apoptotic proteins Bad, Bak, and Bcl-xS were detected in a higher percentage of cells in RA and RAS. RA and RAS were associated with an increased Bcl-xS/Bcl-xL ratio. The expression of anti-apoptotic proteins was also correlated with that of CD34 and P170 and with the percentage of blast cells. Two-color analyses demonstrated that CD34 and Bcl-2 were usually expressed in the same cells. No significant correlation was found with cytogenetic abnormalities. Higher expression of pro-apoptotic Bcl-2-family proteins (Bak, Bad, Bcl-xS) and higher Bcl-xS/Bcl-xL ratio were associated with longer survival and decreased risk of leukemic transformation in univariate analysis, whereas expression of anti-apoptotic proteins was associated with decreased survival. Consequently Bcl-2 proteins expression was well correlated with the International Prognostic Scoring System (IPSS). Our data confirm that the control of apoptosis is deregulated in MDS cells. Moreover, the study of markers such as CD34 (or Bcl-2), Bcl-xL, and Bcl-xS provides additional prognostic information.
