PTEN Loss and Cyclin A2 Upregulation Define a PI3K/AKT Pathway Activation in Helicobacter pylori–induced MALT and DLBCL Gastric Lymphoma With Features of MALT

International audienceHelicobacter pylori infection is strongly associated with primary gastric diseases, such as extranodal mucosa-associated lymphoid tissue (MALT) lymphoma, diffuse large B-cell lymphoma (DLBCL) with histologic evidence of MALT origin, and gastric carcinoma. The cytotoxin-associated gene A (CagA) protein behaves as a bacterial oncoprotein, promoting tumorigenesis via dysregulation of the phosphatidylinositol 3-kinase/AKT pathway (PI3K/AKT). We investigated the molecular mechanisms of PI3K/AKT pathway dysregulation in H. pylori-induced MALT and DLBCL gastric lymphoma. Immunohistochemical assays for CagA, phospho(p)-S473-AKT, PTEN, SHIP, and cyclin A2 proteins were performed on samples from 23 patients with H. pylori-positive MALT lymphoma and 16 patients with H. pylori-positive gastric DLBCL. We showed that CagA localization is correlated with the activation of the AKT pathway in both MALT and DLBCL lymphoma cells. Interestingly, we found a close association between the loss of PTEN, the overexpression of cyclin A2, and the phosphorylation of AKT in gastric MALT and DLBCL tumor cells

5,6-Epoxycholesterol Isomers Induce Oxiapoptophagy in Myeloma Cells

International audienceMultiple myeloma (MM) is an incurable plasma cell malignancy with frequent patient relapse due to innate or acquired drug resistance. Cholesterol metabolism is reported to be altered in MM; therefore, we investigated the potential anti-myeloma activity of two cholesterol derivatives: the 5,6 α- and 5,6 β-epoxycholesterol (EC) isomers. To this end, viability assays were used, and isomers were shown to exhibit important anti-tumor activity in vitro in JJN3 and U266 human myeloma cell lines (HMCLs) and ex vivo in myeloma patients' sorted CD138+ malignant cells. Moreover, we confirmed that 5,6 α-EC and 5,6 β-EC induced oxiapoptophagy through concomitant oxidative stress and caspase-3-mediated apoptosis and autophagy. Interestingly, in combination treatment a synergistic interaction was observed between 5,6 α-EC and 5,6 β-EC on myeloma cells. These data highlight a striking anti-tumor activity of 5,6 α-EC and 5,6 β-EC bioactive molecules against human myeloma cells, paving the way for their potential role in future therapeutic strategies in MM

A lowered 26S proteasome activity correlates with mantle lymphoma cell lines resistance to genotoxic stress

International audienceBackground: Mantle cell lymphoma (MCL) is a B-cell hemopathy characterized by the t(11;14) translocation and the aberrant overexpression of cyclin D1. This results in an unrestrained cell proliferation. Other genetic alterations are common in MCL cells such as SOX11 expression, mutations of ATM and/or TP53 genes, activation of the NF-κB signaling pathway and NOTCH receptors. These alterations lead to the deregulation of the apoptotic machinery and resistance to drugs. We observed that among a panel of MCL cell lines, REC1 cells were resistant towards genotoxic stress. We studied the molecular basis of this resistance.Methods: We analyzed the cell response regarding apoptosis, senescence, cell cycle arrest, DNA damage response and finally the 26S proteasome activity following a genotoxic treatment that causes double strand DNA breaks.Results: MCL cell lines displayed various sensitivity/resistance towards genotoxic stress and, in particular, REC1 cells did not enter apoptosis or senescence after an etoposide treatment. Moreover, the G2/M cell cycle checkpoint was deficient in REC1 cells. We observed that three main actors of apoptosis, senescence and cell cycle regulation (cyclin D1, MCL1 and CDC25A) failed to be degraded by the proteasome machinery in REC1 cells. We ruled out a default of the βTrCP E3-ubiquitine ligase but detected a lowered 26S proteasome activity in REC1 cells compared to other cell lines.Conclusion: The resistance of MCL cells to genotoxic stress correlates with a low 26S proteasome activity. This could represent a relevant biomarker for a subtype of MCL patients with a poor response to therapies and a high risk of relapse

Additional file 3: Figure S2. of A lowered 26S proteasome activity correlates with mantle lymphoma cell lines resistance to genotoxic stress

A. REC1 cells were treated with vehicle or etoposide 4 μg/ml for 2–24 h and harvested. Cells (105 cells per spot) were cytospun on Superfrost glass slides, at 500 g for 3 min, then fixed in 4% paraformaldehyde (PFA) and permeabilized by incubation with 0.5% Triton-X100 (v/v) for 5 min. Slides were then stained with rabbit anti-cyclin D1 primary Ab (sc-718, Santa Cruz Biotech.) and AlexaFuor® 546 goat anti-rabbit IgG (Life Technologies) secondary Ab. DAPI (4′,6-diamidino-2-phenylindole dihydrochloride, Molecular Probes) served for nuclei counterstaining. Slides were mounted, and analyzed with a Fluoview FV 1000 confocal microscope and Fluoview Viewer software (Olympus). B. Cultured JeKo1 and REC1 cells were treated with vehicle (Ctrl) or doxorubicine (Dox 25 nM) for 24 h. Whole cell proteins were purified, separated by SDS-PAGE, and immunoblotted with the indicated antibodies. An anti-β-actin served as a control of charge and transfer. C. Cultured JeKo1 and REC1 cells were treated with vehicle (Ctrl) or bortezomib (10−1-104 nM) or sn38 (10−1-103 nM) for 24 h and then cell viability assessed by a MTS assay as described in the legend of the Fig. 1a. Dose-reponse curves were drawn with the PRISM® software (GraphPad, La Jolla, CA) and the IC50 were deduced from the data. (PPTX 5260 kb

Diagnostic value of CA19.9, circulating tumour DNA and circulating tumour cells in patients with solid pancreatic tumours

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