Gene expression profiling of T cells of CLL patients, stimulated with antiCD3 antibody in presence or not of idelalisib
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ABSTRACT: The aim of the study is to identify genes modulated by idelalisib in T cells of CLL patients, that could suggest an impairment of T cell-mediated immunity caused by the drug. We found 61 genes downregulated by idelalisib in stimulated T cells with Anova p≤0.01 and [FC]≥2 and the most represented categories were chemokines involved in T cell migration (as CXCL9-10-11), inflammatory and T helper cytokines (as IL-2, TNFα, IFNg, IL-13, IL-21), mitogenic signal transducers (as IRF4, STAT1, EGR1-2-3-4) and receptors (as CD69, CD25) commonly induced during activation of T cells.
Project description:Idelalisib is a phosphatidylinositol 3-delta kinase inhibitor that has shown good efficacy in treating some hematologic malignancies. Rare, but potentially serious liver toxicity was associated with idelalisib use in clinical trials. The objective of this study was to evaluate the idelalisib-induced activation of stress response pathways in human hepatocytes to inform mechanisms of liver injury observed in the clinic Primary human hepatocytes from three donors were sandwich cultured and assayed for global gene expression across 5 concentrations of idelalisib after a 24 h exposure. The most significantly enriched pathway among genes upregulated in response to idelalisib was “Endoplasmic Reticulum Stress”. These data suggest that oxidative stress is a dominant mechanism contributing to liver injury associated with idelalisib.
Project description:RNAseq profile of TMD8 cell lines resistant to Idelalisib treatment. Idelalisib resistant TMD8 cells were generated by continuous passage in the presence of 1 μM idelalisib for 8 weeks until stable resistance to idelalisib was established. Parallel cultures were grown in the presence of 0.1% DMSO as passage-matched, drug-sensitive control lines. Sensitive and resistant TMD8 cells were clonally isolated through two rounds of single cell limiting dilution
Project description:Follicular Lymphoma primary cells (FL) were treated with the PI3K delta inhibitor idelalisib (500nM, 48h) in the presence or absence of follicular dendritic cells We used microarrays to uncover the mechanisms underlying mechanism of resistance and sensitivity of FL to idelalisib
Project description:Idelalisib was the first-in-class PI3Kδ inhibitor and additional compounds are undergoing clinical investigation. To identify modalities to overcome resistance to these agents, we have developed idelalisib-resistant model derived from marginal zone lymphoma cell line (VL51). Cells were kept under idelalisib until acquisition of resistance (VL51-RER) or with no drug (parental). In this experiment we identified the modulated genes between the resistant cell line and the parental counterpart (sensitive to the drug) We invastigated the transcriptomic profiles of parental K1718 B-cell lymphoma cell lines and the same cell line made resistant to Idelalisib. in addition two B-Cells (SUDHL2 and SUDHL4) are made resistant to IMGN529 (Naratuximab emtansine) an antibody-drug conjugate (ADC) incorporating an anti-CD37 monoclonal antibody conjugated to the maytansinoid DM1 as payload.
Project description:BCR pathway inhibitors idelalisib and ibrutinib are the first small molecule targeted agents for B-cell malignancies. In spite of encouraging response rates in various forms of B cell diseases, patients will eventually develop relapse due to the emergence of resistant cells. To better identify the possible mechanisms of resistance we developed and characterized idelalisib- and ibrutinib-resistant variants of the human non Hodgkin’s lymphoma cell lines DoHH2 and Daudi. These resistant variants displayed a cross-resistance profile limited to PI3K inhibitors, BTK inhibitors and a SYK inhibitor but not to unrelated agents. A number of alterations were observed in the resistant lines, including a strong reduction of the Akt3 protein. Resistant lines tended to express larger amounts of CD38 and CD52 on their cell membrane and were found to display enhanced sensitivity to anti-CD38 antibodies. These results identify potential novel mechanisms of resistance to idelalisib and ibrutinib and raise the possibility that cells resistant to BCR pathway inhibitors might possess enhanced sensitivity to anti-CD38 antibodies.
Project description:Interleukin-21 (IL-21) is a pleiotropic cytokine that induces expression of transcription factor BLIMP1 (encoded by Prdm1), which regulates plasma cell differentiation and T cell homeostasis. We identified an IL-21 response element downstream of Prdm1 that binds the transcription factors STAT3 and IRF4, which are required for optimal Prdm1 expression. Genome-wide ChIP-Seq mapping of STAT3- and IRF4-binding sites showed that most regions with IL-21-induced STAT3 binding also bound IRF4 in vivo, and furthermore, revealed that the noncanonical TTCnnnTAA GAS motif critical in Prdm1 was broadly used for STAT3 binding. Comparing genome-wide expression array data to binding sites revealed that most IL-21-regulated genes were associated with combined STAT3-IRF4 sites rather than pure STAT3 sites. Correspondingly, ChIP-Seq analysis of Irf4_/_ T cells showed greatly diminished STAT3 binding after IL-21 treatment, and Irf4_/_ mice showed impaired IL- 21-induced Tfh cell differentiation in vivo. These results reveal broad cooperative gene regulation by STAT3 and IRF4. Affymetrix expression data: Prepare CD4+ T cells from spleen. CD4+ T cells were preactivated, rested, and treated with IL-21 for 1, 6, and 24 hours. ChIP-seq data: Profiling of IRF4 and Stat3 binding with and without IL-21 stimulation in wild type and IRF4 KO mice.
Project description:Gene expression profiling of murine irf4-/- and irf4+/+ splenic B cells identifies genes regulated by the transcription factor IRF4 in CD40+IL-4 activated mature B cells. B cells from 12-week old irf4-/- and irf4+/+ littermate mice were isolated by magnetic depletion of non-B cells from splenic mononuclear cells and cultures with mitogens in vitro. RNA was isolated. Labeled cRNA was hybridized to microarrays and genes specifically expressed in irf4-/- vs. irf4+/+ samples.