Modulation of CD20 Expression in Rituximab-Sensitive (RSCL) and Rituximab Resistant Cell Lines (RRCL) Using IL-4 and Bryostatin-1.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2619-2619 ◽  
Author(s):  
Ping-Chiao Tsai ◽  
Francisco J. Hernandez-Ilizaliturri ◽  
Scott H. Olejniczak ◽  
Bangia Naveen ◽  
Myron S. Czuczman
Keyword(s):  
Transcription Factors ◽  
B Cell ◽  
Western Blotting ◽  
Flow Cytometric Analysis ◽  
Regulation Of Transcription ◽  
Bryostatin 1 ◽  
Down Regulation ◽  
Cd20 Antigen ◽  
Number Of Patients ◽  
Cd20 Expression

Abstract While the use of rituximab in combination with chemotherapy resulted in an improved survival among various subtypes of B-cell lymphomas, a significant number of patients fail to respond or relapse as a consequence of intrinsic or acquired resistance. A change in CD20 antigen density expression is a potential mechanism to explain rituximab resistance. Moreover, several groups of investigators are focused in understanding the mechanisms that regulate CD20 expression and to develop therapeutic strategies to up-regulate CD20 expression (i.e. IL-4, GM-CSF or Bryostatin-1). Up-regulation of CD20 in DB and Ramos cells by Bryostatin-1 was found to be PKC and Erk dependent. In an attempt to characterize the mechanisms responsible for rituximab resistance we developed several RRCL derived from rituximab-sensitive RL and Raji cells. We have demonstrated a significant down-regulation of CD20mRNA and CD20 surface antigen in RRCL when compared to RSCL. In our present work we evaluated the mechanisms involved in the mRNA down-regulation of CD20 and the potential of IL-4 and Bryostatin-1 in modulating CD20 antigen expression among a panel of RRCL. To this end, RSCL and RRCL were treated with either 5ng/ml of IL-4 or 3 different doses of Bryostatin-1(1, 3 or 5ng/ml). Nuclear and cytosolic extract were also obtained from RSCL RL and RRCL RL-4RH after 24, 48 and 74 hrs exposure to IL-4 (5ng/ml) or control. Differences in the expression of key regulatory transcription factors for B-cell lymphocyte development (PU.1, Oct-2, Pax5, E2A and EBF) were studied by Western Blotting. Previously, we found a significant down-regulation of CD20 antigen in the RRCL. An up-regulation of cytosolic and surface CD20 was detected in RRCL exposed to either IL-4 or Bryostatin by western blotting and flow cytometric analysis. Besides, a higher expression of Pax5, PU.1, EBF was found in nuclear fractions of RRCL when compared to RSCL. In vitro exposure of RRCL to IL-4 decreases the expression of PU.1, Oct-2, Pax5 and EBF in nuclear extracts from RSCL or RRCL when compared with controls treated cells. Our data suggest that rituximab resistance is associated with the up-regulation of transcription factors PU.1, Oct-2, Pax5 and EBF and concomitant suppression of CD20 antigen. Future study of how these agents induce CD20 expression in RRCL will provide a potential means to reversing resistance towards rituximab treatment.

Pax5 Activates Oncogenic Transcription Factors of the Ets Family by Repressing the mir-15a/16-1 microRNA Cluster.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 346-346
Author(s):  
Elaine Y. Chung ◽  
Diana Cozma ◽  
Duonan Yu ◽  
Michael Dews ◽  
Erik A. Wentzel ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cell ◽  
Western Blotting ◽  
Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Mrna Levels ◽  
Down Regulation ◽  
Protein Levels ◽  
Ets Family ◽  
Post Transcriptional Regulation

Abstract We have recently demonstrated that Pax5 promotes B-lymphomagenesis by upregulating key components of B-cell receptor signaling [Cozma et al, J Clin Inv, 117 (8), 2007]. Gene regulation by Pax5 often involves complex formation with other oncogenic transcription factors of the Ets family, namely Myb and Ets1. We determined that expression of these proteins themselves depends on the presence of Pax5, as seen in human diffuse large B-cell lymphomas with Pax5 knockdown and murine lymphomas with epigenetic silencing of Pax5 [Yu et al, Blood, 101:1950–1955, 2003; Johnson et al, Nat Immunol, 5:853–861, 2004]. Upon reconstitution with the Pax5 gene, Myb and Ets1 levels increase sharply. This occurs with little increase in steady-state mRNA levels, suggesting post-transcriptional regulation, possibly by microRNAs. To test this hypothesis, we compared miRNA profiles of Pax5-deficieint and sufficient cells and discovered that several miRNAs are indeed repressed by Pax5. Among them is the miR-15a/16-1 cluster whose predicted targets include both Myb and Ets1. Consistent with this prediction, forced expression of miR-15a/16 brings down Myb and Ets1 protein levels. This is accompanied by impaired Pax5 function and overall suppression of B-lymphomagenesis. Thus, Ets family members (along with previously identified bcl-2) are key targets of the miR-15a/16 locus, a tumor suppressor in chronic lymphocytic leukemia. Interplay between Pax5, Myb/Ets1, and miR-15a/16-1. (A) Upregulation of Myb and Ets 1 in tumors over-expressing Pax5ER fusion, as compared to control GFP-only neoplasms. (B) Down-regulation of Myb and Ets1 in Pax5 tumors engineered to over-express the miR-15a/16-1 cluster. All panels depict Western blotting. Interplay between Pax5, Myb/Ets1, and miR-15a/16-1. (A) Upregulation of Myb and Ets 1 in tumors over-expressing Pax5ER fusion, as compared to control GFP-only neoplasms. (B) Down-regulation of Myb and Ets1 in Pax5 tumors engineered to over-express the miR-15a/16-1 cluster. All panels depict Western blotting.

Impaired Ca++ mobilization in rituximab-resistant cells (RRCL) is associated with changes in the structure of CD20 antigen, down-regulation of Bax/Bak pro-apoptotic proteins and up-regulation of the endoplasmic reticulum (ER) Ca++ pump protein SERCA-3

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 2516-2516 ◽  
Author(s):  
F. J. Hernandez-Ilizaliturri ◽  
H. Kaur ◽  
A. Bhinder ◽  
S. Olejniczak ◽  
J. Knight ◽  
...  
Keyword(s):  
Western Blotting ◽  
Flow Cytometric Analysis ◽  
Acetoxymethyl Ester ◽  
Down Regulation ◽  
Raji Cells ◽  
External Domain ◽  
Cd20 Expression ◽  
Membrane Reorganization ◽  
Apoptotic Proteins

2516 Ca++ mobilization leading to apoptosis has been observed following rituximab biding to CD20 in lymphoma cells. Extending rituximab (R) exposure (e.g. as via R maintenance regimens) could potentially accelerate the emergence of resistance to rituximab. To define the molecular basis for rituximab resistance we developed several RRCL and demonstrated changes in CD20 expression/structure and membrane reorganization following rituximab therapy. Specifically, changes in the N-terminal and the C-terminal region of the internal domain of CD20 were observed in RRCL as determined by Western blotting using various antibodies recognizing epitopes located in the internal (GST77 and 1439) and external domain (B1) of CD20. In our current work we evaluated the effects that CD20 structure/expression changes have upon Ca++ mobilization. Extracellular and intracellular Ca++ mobilization was measured by flow cytometric analysis using FLUO-3 AM (acetoxymethyl ester). Optimization of the Ca++ indicator and calibration curves were performed for each cell line. Raji and RRCL were labeled under optimal conditions with FLUO-3 AM/Pluronic Acid F-127. Subsequently, cells were then re-suspended in HANKS media with or without Ca++ and exposed to rituximab or isotype (10μg/ml) ± human serum (25%). Surface CD20 expression was similar between Raji cells and RRCL. Expression of pro- or anti-apoptotic proteins and Ca++ regulatory proteins SERCA3, SERCA2 and Calreticulin was also determined by western blotting. In vitro exposure of Raji cells to rituximab + HS resulted in Ca++ mobilization even in Ca++ depleted media. Notably, Ca++ mobilization was impaired in RRCL when compared to Raji parental cells. In addition, down-regulation of Bax/Bak and up-regulation of SERCA3 was demonstrated in RRCL. Our data suggest that the acquirement of rituximab resistance is associated with changes in the intracellular domain of CD20 and in Ca++ regulator/pro-apoptotic proteins (SERCA3 and Bax/Bak) resulting in a decrease in the intracellular mobilization of Ca++. No significant financial relationships to disclose.

Emergence of Rituximab-Fludarabine Resistance Is Associated with Changes in CD20 Antigen Expression and Improved Response to Almetuzumab Therapy in Patients with Chronic Lymphocytic Leukemia (CLL).

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2054-2054 ◽  
Author(s):  
Umeer Ashraf ◽  
Anjana N. Elefante ◽  
Raymond Cruz ◽  
Paul K. Wallace ◽  
Myron S. Czuczman ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Overall Survival ◽  
B Cell ◽  
Response Rate ◽  
Antigen Expression ◽  
Down Regulation ◽  
Cd20 Antigen ◽  
Cd20 Expression ◽  
Over Time

Abstract Cancer cells, including B-cell lymphoproliferative disorders, sculpt their phenotype in an attempt to escape not only immune-surveillance but also evade the anti-tumor activity of biological agents and/or chemotherapy drugs including rituximab. In an attempt to study the mechanisms that regulate the emergence of rituximab resistance, we developed several rituximab-resistant cell lines and demonstrated that the emergence of a rituximab-resistant phenotype was associated with global down regulation of CD20 antigen and unexpectedly an increase in CD52 antigen. Validation of our findings in more clinically relevant settings is important to support alemtuzumab-based clinical studies in rituximab-resistant B-cell malignancies. To this end, we retrospectively studied changes in CD20 expression over time in alemtuzumab treated-patients with rituximab-fludarabine (RF) refractory CLL. Patients were identified using the institute tumor registry and pharmacy electronic database. Demographic characteristics, treatment history, outcome data were obtained for each patient. In addition, CD20 expression in CLL cells obtained from flow cytometry analysis performed on peripheral blood, bone marrow and tissue was reviewed. A total of 16 patients with RF refractory CLL treated with alemtuzumab were included in the analysis, 13 males and 3 females, the median age at the time of diagnosis was 55.5 yrs +/− 10.47stv, and most of the patients had a good PS (0) at the time of treatment with alemtuzumab (81%). The response rate to alemtuzumab was 56.3%, with 7 (43.5%) patients achieving a complete response (CR). After a median follow up period of 88.5 months, 7 patients are still alive, 3 are free of disease. Complete flow cytometry data was available only for 9 patients. A down regulation of CD20 antigen expression was observed among 5/9 patients in blood, bone marrow and/or tissue over time when compared to baseline CD20 levels. Following alemtuzumab therapy, CR was achieved in 3/5 patients with CD20 down-regulation versus 1/4 patients with steady levels of CD20. Alemtuzumab-treated CLL patients with CD20 down-regulation had a longer overall survival (141 months) than alemtuzumab treated CLL patients with steady levels of CD20 antigen overtime (116 months, Log Rank P = 0.026). Our data suggest, that emergence of rituximab-fludarabine resistance is associated with changes in the expression of CD20 antigen. A response to alemtuzumab in this setting appears to be higher than historical controls (2% CR). Refractory CLL patients with decreased levels of surface CD20 appears to respond better to alemtuzumab and have a longer overall survival than patients with steady CD20 levels. Similarly to what we have observed in our pre-clinical models, an increase in CD52 expression in rituximab-fludarabine refractory patients with CD20 down regulation could explain the higher response rate observed in this small group of highly selected patients. Detailed monitoring of CD20 and CD52 levels in CLL patients before, during and after treatment with rituximab-based regimens is warranted in an attempt to identify patients that can benefit from almetuzumab-based therapies.

The Hexosamine Biosyntheic Pathway and Platinum-Ethylenedicysteine-Glucosamine (Pt-ECG) for Targeted Therapy in Diffuse Large B-Cell Lymphoma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 587-587
Author(s):  
Lan V Pham ◽  
Jerry Bryant ◽  
Archito T. Tamayo ◽  
Richard Mendez ◽  
Edna Chum ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cell ◽  
Cancer Biology ◽  
Biosynthetic Pathway ◽  
B Cell Lymphoma ◽  
Regulation Of Transcription ◽  
Growth And Survival ◽  
Lead Compounds ◽  
Hexosamine Pathway ◽  
Large B Cell

Abstract Abstract 587 Aggressive non-Hodgkin lymphomas (NHL), such as diffuse large B cell lymphomas (DLBCL), are very common in the US with increasing incidences. Although these lymphomas are now potentially curable, almost half the treated patients still develop relapsed/refractory disease with poor survival outcomes, indicating an urgent need for better therapeutic approaches with improved efficacy. The hexosamine signaling pathway terminating in O-linked N-acetyl glucosamine (O-GlcNAc) cycling has been implicated in cellular signaling cascades and regulation of transcription factors involved in cancer biology. Biological functions of the hexosamine biosynthetic signaling pathways need elucidation, to determine whether altered O-GlcNAc metabolism plays a significant role in hematologic tumors such as DLBCL, and utilize this bifunctional pathway as a targeted therapeutic strategy in DLBCL. We have identified key enzymes of the hexosamine biosynthetic pathways to be highly-expressed in DLBCL cell lines and patient tumor cells. In contrast to normal circulating and tonsillar B cells, DLBCL cells expressed high levels of the rate limiting enzyme glutamine: fructose-6-phosphate amidotransferase (GFAT) as well as terminating enzyme O-GlcNAc transferase (OGT). We discovered that several key growth and survival transcription factors, such as NF-kB and NFAT, known to be highly-activated in DLBCL, are linked to the hexoasmine biosynthetic pathway. We demonstrated that both NF-kB (p65) and NFATc1 directly associated with OGT, and down-regulation of OGT by siRNA inhibits these transcription factors activation, suggesting that both NF-kB-p65 and NFATc1 require O-GlcNAc glycosylation by OGT for their activation. ChiP on Chip analysis on NFATc1 indicated that this transcription factor regulates a set of genes involved in glucose metabolism, including hexokinase and GFAT. These results suggest that the hexosamine pathway is highly active and utilized in DLBCL, and that exploiting this bi-functional pathway(s) as a therapeutic approach is feasible. We have previously developed an imaging agent, 99mTc-ethylenedicysteine-glucosamine (99mTc-EC-G) because EC-G mimics phosphorylated N-acetylglucosamine. ECG treatment in DLBCL cells enhances p65 and NFATc1 nuclear translocation. For therapeutic strategies, we developed metallic unlabeled Platinum (Pt) derivatives-EC-G as potential therapeutic agents. Pre-clinical in vitro studies have shown that our two lead compounds, Pt- and Pt-(DACH)-EC-G effectively inhibit lymphoma cell growth and induce apoptosis. These lead compounds can also induce DNA damage in DLBCL cells, through the up-regulation of phosphorylated histone 2AX (pH2AX), leading to the disruption of p65 and NFATc1 binding to DNA. This data importantly demonstrates that the hexosamine biosynthetic pathway is linked to key growth and survival pathways involved in the pathophysiology of DLBCL. Targeting these pathways with novel platinum EC-G compounds as a theranostic approach should lead to new, more effective treatments and diagnosis for DLBCL, particularly for relapsed/refractory DLBCL. Disclosures: Rollo: Cell Point: Employment.

Trispecific CD19-CD20-CD22–targeting duoCAR-T cells eliminate antigen-heterogeneous B cell tumors in preclinical models

2021 ◽  
Vol 13 (586) ◽  
pp. eabc6401
Author(s):  
Dina Schneider ◽  
Ying Xiong ◽  
Darong Wu ◽  
Peirong Hu ◽  
Leah Alabanza ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Intracellular Signaling ◽  
B Cell Lymphoma ◽  
Target Antigen ◽  
Down Regulation ◽  
T Cell Therapy ◽  
Number Of Patients ◽  
Antigen Loss

A substantial number of patients with leukemia and lymphoma treated with anti-CD19 or anti-CD22 monoCAR-T cell therapy relapse because of antigen loss or down-regulation. We hypothesized that B cell tumor antigen escape may be overcome by a chimeric antigen receptor (CAR) design that simultaneously targets three B cell leukemia antigens. We engineered trispecific duoCAR-T cells with lentiviral vectors encoding two CAR open reading frames that target CD19, CD20, and CD22. The duoCARs were composed of a CAR with a tandem CD19- and CD20-targeting binder, linked by the P2A self-cleaving peptide to a second CAR targeting CD22. Multiple combinations of intracellular T cell signaling motifs were evaluated. The most potent duoCAR architectures included those with ICOS, OX40, or CD27 signaling domains rather than those from CD28 or 4-1BB. We identified four optimal binder and signaling combinations that potently rejected xenografted leukemia and lymphoma tumors in vivo. Moreover, in mice bearing a mixture of B cell lymphoma lines composed of parental triple-positive cells, CD19-negative, CD20-negative, and CD22-negative variants, only the trispecific duoCAR-T cells rapidly and efficiently rejected the tumors. Each of the monoCAR-T cells failed to prevent tumor progression. Analysis of intracellular signaling profiles demonstrates that the distinct signaling of the intracellular domains used may contribute to these differential effects. Multispecific duoCAR-T cells are a promising strategy to prevent antigen loss–mediated relapse or the down-regulation of target antigen in patients with B cell malignancies.

Recurrent 11q24.3 Gain Contributes to the Pathogenesis of Diffuse Large B Cell Lymphoma (DLBCL) by Deregulating ETS1 and FLI1

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1386-1386
Author(s):  
Paola Bonetti ◽  
Monica Testoni ◽  
Marta Scandurra ◽  
Maurilio Ponzoni ◽  
Roberto Piva ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
B Cell ◽  
Germinal Center ◽  
Conflicts Of Interest ◽  
Cell Lymphoma ◽  
Functional Characterization ◽  
B Cell Lymphoma ◽  
Transcriptional Program ◽  
Down Regulation

Abstract Abstract 1386 DLBCL represents the most common form of B-cell non-Hodgkin lymphoma (B-NHL). It is an aggressive and heterogeneous disease, comprising at least three distinct subtypes based on gene expression profile analysis: germinal center B cell-like DLBCL (GCB), activated B cell-like DLBCL (ABC) and primary mediastinal B-cell lymphoma (PMBL). These subtypes are supposed to derive from B cells at different stages of differentiation. Normal germinal center (GC) B-cell differentiation requires a complex transcriptional program and alterations of genes involved in this process are relevant for DLBCL pathogenesis. Identification and functional characterization of new genetic lesions would provide critical information to better understand the pathogenesis of DLBCL. With this aim, we studied the genomic profiles of 166 DLBCL patients, identified and characterized a recurrent gain mapping to chromosome 11q24.3. Methods. Genomic profiles were obtained from 166 Affymetrix 250K SNP arrays and integrated with gene expression data (GeneChip U133 plus 2.0) in 54 cases. Data were validated by PCR and immunohistochemistry. Gene silencing experiments were done with shRNA. Results. A minimal common region 11q24.3 gain was present in 26% of DLBCL samples and it encompassed six genes (ETS1, FLI1, KCNJ1, KCNJ5, P53AIP1, RICS). Samples with the 11q24.3 gain were significantly associated with high expression of the transcription factors ETS1 and FLI1. Data were confirmed by real-time PCR and by immunohistochemical analysis. Gene expression analysis showed 228 transcripts with a significantly different expression between cases with or without the lesion (p<0.01, >2-fold change): 215 genes were up-regulated in the patients with the gain and 13 were down-regulated, suggesting that this lesion has an impact on the transcriptional program of the tumor cells. To study the biological meaning of the lesion, ETS1 and FLI1 expression was down-regulated in a DLBCL cell line bearing the same lesion observed in clinical specimens (OCI-Ly7). Results showed that ETS1 and FLI1 down-regulation caused a reduced proliferation rate and activation of apoptosis leading to cell death. Concomitant ETS1 and FLI1 down-regulation resulted in a more severe phenotype. Only FLI1 was confirmed to be essential for cell viability in other DLBCL cell lines (SUDHL4, VAL, U2932), whereas ETS1 did not, suggesting a distinct role of the two transcription factors in different DLBCL samples. Preliminary results showed that down-regulation of ETS1 affected the transcriptional program of GC B-cell terminal differentiation causing an up-regulation of BLIMP1, the master regulator of plasma cells differentiation. Conclusions. In DLBCL, a recurrent gain at 11q24.3 determines the over-expression of the transcription factors ETS1 and FLI1. Functional experiments showed that the lesion might sustain DLBCL proliferation and viability, and contribute to a differentiation blockade of the GC B-cell towards a plasma cell lineage by negatively regulating BLIMP1. Disclosures: No relevant conflicts of interest to declare.

The Human Pre-B Cell Receptor Signaling Cascade Is Regulated Via PI-3Kinase and MAPK Pathway

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1314-1314
Author(s):  
Kolandaswamy Anbazhagan ◽  
Vincent Fuentes ◽  
Eliane Bissac ◽  
Remy Nyga ◽  
Naomi Taylor ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cells ◽  
B Cell ◽  
Nuclear Translocation ◽  
Accessory Pathway ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Signaling Cascade ◽  
Down Regulation ◽  
Bcr Signaling

Abstract Abstract 1314 Background: Pre-B cell receptor (pre-BCR) constitutes a major check point in the early steps of mouse and human B cell development. Several functions have been attributed to this receptor which include a delivery of proliferation and survival signals, increased sensitivity to interleukin-7 (IL-7) and down modulation of recombinase activating genes (RAG) and surrogate light chain (SLC) encoding genes. Pre-BCR is also involved in shaping the VH repertoire and preventing autoimmunity. Finally, there is increasing evidence that pre-BCR might be implicated in leukemogenesis. Most of the functions of pre-BCR have been predicted based on studies in knockout mice and leukemic cell lines. In a previous study we have shown that pre-BCR aggregation resulted in the activation of src and Syk kinases which in turn activated the PI-3K/Akt, Btk, PLCγ-2 and Ras/MAPK. In this study, we examined the pre-BCR signalling cascade using human normal primary pre-B cells with a particular focus on transcription factors activation and Rag modulation and their regulatory aspects. Methods: Pre-B cells were sorted from adult human bone marrow samples, treated or not with inhibitors of Syk (BAY61–3606), Akt (LY294002) and MEKK1 (UO126) prior to crosslink the pre-BCR by means of F(ab')2 anti-μHC. The effect of Pre-BCR signaling was examined by quantifying the transcript levels of Rag1, Rag2, E2A, EBF1, Pax5, FoxO1 and FoxO3, IRF4/8. Activation of transcription factors such as NF-κB p50, c-Fos, IRF4 and FoxO3A, was assessed by analyzing their nuclear translocation by immunofluorescence microscopy. Results: We show that NF-κB p50 is translocated into nucleus within 3h after pre-BCR stimulation. Crosslinking of pre-BCR also resulted in an enhancement of nuclear c-Fos translocation. BAY61-3606 (Syk inhibitor) treatment resulted in complete apoptosis (100 % cell death within 48h). Although treatment of normal pre-B cells with LY294002 or U0126 did not alter cell survival, nuclear translocation of pre-BCR-induced p50 NF-κB was prevented by former and enhanced by later. Conversely, c-Fos nuclear expression was inhibited by U0126 and slightly but consistently enhanced by LY294002 in association with a decrease in its cytoplasmic location. Pre-BCR stimulation also induced IRF4 translocation to the nucleus. Pre-BCR stimulation also resulted in the down regulation of Rag1 (− 48 %, P<0.01), Pax5 (− 40%, P<0.01) and E2A (− 35 %, P< 0.01) transcripts, whereas EBF1 and FoxO1 and 3 expression remained unchanged. In LY294002-treated cells, Rag1/Rag2 expression was up regulated (+130%, P< 0.01 and +251%, P< 0.01, respectively) following pre-BCR crosslinking, whereas in the presence of U0126 the pre-BCR induced Rag1/Rag2 down modulation remained unchanged. Conclusion: Our results indicate that the pre-BCR has the potential to promote pre-B cell proliferation, survival and differentiation by activating NF-kB, c-Fos and IRF4. It also has the ability to protect pre-B cells from genome instability by down-regulating Rag1/2, probably through down modulation of Pax5 and E2A. We bring evidence that PI-3 K/Akt pathway plays a crucial role in the regulation of the pre-BCR signaling cascade and that Akt-mediated NF-kB and c-Fos activation is antagonized by MAPK. Up-regulation of Rag transcripts upon Akt inhibition suggests either a feed-back negative loop or a dual effect of pre-BCR on Rag expression with an Akt-dependent Rag down regulation and an accessory pathway that enhances Rag expression. Disclosures: No relevant conflicts of interest to declare.

Quinalizarin inhibits adipogenesis through down-regulation of transcription factors and microRNA modulation

2017 ◽  
Vol 1861 (12) ◽  
pp. 3272-3281 ◽  
Author(s):  
Lisa Schwind ◽  
Lisa Nalbach ◽  
Andreas D. Zimmer ◽  
Katja B. Kostelnik ◽  
Jennifer Menegatti ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Regulation Of Transcription ◽  
Down Regulation

Down-regulation of CD20 expression in B-cell lymphoma cells after treatment with rituximab-containing combination chemotherapies: its prevalence and clinical significance

Blood ◽  
2009 ◽  
Vol 113 (20) ◽  
pp. 4885-4893 ◽  
Author(s):  
Junji Hiraga ◽  
Akihiro Tomita ◽  
Takumi Sugimoto ◽  
Kazuyuki Shimada ◽  
Masafumi Ito ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Surface Expression ◽  
University Hospital ◽  
Cell Surface Expression ◽  
Phenotypic Change ◽  
Down Regulation ◽  
Cd20 Expression

Although rituximab is a key molecular targeting drug for CD20-positive B-cell lymphomas, resistance to rituximab has recently been recognized as a considerable problem. Here, we report that a CD20-negative phenotypic change after chemotherapies with rituximab occurs in a certain number of CD20-positive B-cell lymphoma patients. For 5 years, 124 patients with B-cell malignancies were treated with rituximab-containing chemotherapies in Nagoya University Hospital. Relapse or progression was confirmed in 36 patients (29.0%), and a rebiopsy was performed in 19 patients. Of those 19, 5 (26.3%; diffuse large B-cell lymphoma [DLBCL], 3 cases; DLBCL transformed from follicular lymphoma, 2 cases) indicated CD20 protein-negative transformation. Despite salvage chemotherapies without rituximab, all 5 patients died within 1 year of the CD20-negative transformation. Quantitative reverse-transcription–polymerase chain reaction (RT-PCR) showed that CD20 mRNA expression was significantly lower in CD20-negative cells than in CD20-positive cells obtained from the same patient. Interestingly, when CD20-negative cells were treated with 5-aza-2′-deoxycytidine in vitro, the expression of CD20 mRNA was stimulated within 3 days, resulting in the restoration of both cell surface expression of the CD20 protein and rituximab sensitivity. These findings suggest that some epigenetic mechanisms may be partly related to the down-regulation of CD20 expression after rituximab treatment.

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