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Background Chronic lymphocytic leukemia (CLL) cells are often affected by genomic aberrations targeting key regulatory genes. Although fludarabine is the standard first line therapy to treat CLL, only few data are available about the resistance of B cells to this purine nucleoside analog in vivo. Here we sought to increase our understanding of fludarabine action and describe the mechanisms leading to resistance in vivo. We performed an analysis of genomic aberrations, gene expression profiles, and microRNAs expression in CLL blood B lymphocytes isolated during the course of patients' treatment with fludarabine. Results In sensitive patients, the differentially expressed genes we identified were mainly involved in p53 signaling, DNA damage response, cell cycle and cell death. In resistant patients, uncommon genomic abnormalities were observed and the resistance toward fludarabine could be characterized based on the expression profiles of genes implicated in lymphocyte proliferation, DNA repair, and cell growth and survival. Of particular interest in some patients was the amplification of MYC (8q) observed both at the gene and transcript levels, together with alterations of myc-transcriptional targets, including genes and miRNAs involved in the regulation of cell cycle and proliferation. Differential expression of the sulfatase SULF2 and of miR-29a, -181a, and -221 was also observed between resistant and sensitive patients before treatment. These observations were further confirmed on a validation cohort of CLL patients treated with fludarabine in vitro. Conclusion In the present study we identified genes and miRNAs that may predict clinical resistance of CLL to fludarabine, and describe an interesting oncogenic mechanism in CLL patients resistant to fludarabine by which the complete MYC-specific regulatory network was altered (DNA and RNA levels, and transcriptional targets). These results should prove useful for understanding and overcoming refractoriness to fludarabine and also for predicting the clinical outcome of CLL patients before or early during their treatment.

Multiple myeloma is, despite the emergence of new treatment strategies in recent years, still a lethal disease in 2005. Over the last several years, significant insights into the dysregulation of various signal transduction pathways of MM have emerged and have evolved the development of new agents. Histone deacetylase inhibitors as valproic acid are compounds that inhibit HDACs enzymes that, in conjunction with histone acetylases, regulate the acetylation state of histones and, by extension, the conformational status of chromatin. VPA is clinically known to be used in treatment of different types of epileptic disease. Effects and signal transduction pathways of cell death have been studied in cells harvested and purified from routine bone marrow aspirates of several patients with multiple myeloma, as well as on myeloma cell lines (OPM2, U266) treated with a physiologic dose range of valproic acid (1–4 mM). We observe significant in vitro toxicity starting at doses of 1 mM for 24h in cell lines as well as in patient cells using an XTT based cytotoxicity assay. The question we adressed was the mechanisms by which the MM died. Flow cytometric analysis with PI / Annexin V staining does not show apoptosis features, nor do TUNEL staining or DNA fragmentation assays, suggesting non activation of the intrinsic apoptosis pathway. In contrast, cell morphology of treated cells stained with May-Gruenwald-Giemsa staining show increase in multinucleated giant cells. Multinucleation has often been described in cells which die through mitotic catastrophe. Further experiments exploring this hypothesis will be conducted. Effect of 48h valproic ac on myeloma cell lines Effect of 48h valproic ac on myeloma cell lines Effect of 48h valproic ac on CD138 cells from myeloma patients Effect of 48h valproic ac on CD138 cells from myeloma patients