scholarly journals Common Gene Expression Signature of B-Cells of Waldenström Macroglobulinemia (WM) and IgM Monoclonal Gammopathies of Undetermined Significance (IgM MGUS) Compared to Healthy Subjects

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4317-4317
Author(s):  
Alessandra Trojani ◽  
Barbara Di Camillo ◽  
Luca Emanuele Bossi ◽  
Antonino Greco ◽  
Livia Leuzzi ◽  
...  
Keyword(s):  
Gene Expression ◽  
B Cells ◽  
B Cell ◽  
Signaling Pathway ◽  
Plasma Cells ◽  
Cell Lymphoma ◽  
Common Gene ◽  
Bcr Signaling ◽  
Unique Genes ◽  
Undetermined Significance

Abstract We performed a comparative gene expression profiling (GEP) study on B-cells and plasma cells of Waldenström Macroglobulinemia (WM), IgM monoclonal gammopathies of undetermined significance (IgMMGUS), and normal individuals (CTRLs) to identify GEP changes as reliable predictors of progression of IgMMGUS to WM. We analyzed bone marrow B-cells and plasma cells from 36 WM patients, 13 IgMMGUS subjects, and 7 CTRLs by Affymetrix microarray, respectively (Table 1). GEP experiments were performed on the CD19+ and CD138+ cells using GeneChip-HGU133 Plus 2.0. Data were preprocessed and normalized by Robust Multi-Array Average and ComBat. Selection of the different expressed genes was performed separately for CD19+ and CD138+ cells, using Significance Analysis of Microarrays (SAM) on the 3 groups and a false discovery rate threshold of 5%, followed, for significance comparisons, by a pair-wise SAM test corrected for multiple testing. We focused on the comparison of the CD19+ cells of WM vs. IgMMGUS vs. CTRLs which highlighted 2038 unique genes whereas the same comparison of the CD138+ cells determined 29 unique genes (Trojani et.al. Cancers 2021). Among the 2038 DEGs, 115 genes were grouped in KEGG pathways involved in Wnt-signaling, BCR-signaling, calcium signaling, hematopoietic cell antigens, cell adhesion, adherens junctions, coagulation cascade, platelet activation, cytokine receptor, and signaling pathways responsible for cell cycle, apoptosis, and survival. Interestingly, most of the 115 DEGs in B-cells were different expressed in WM vs. IgMMGUS and CTRLs. Only 9/115 DEGs were significantly different expressed in WM vs. CTRLs and in IgMMGUS vs. CTRLs, but no significant expression changes were noted between WM and IgMMGUS (Table 2). To further inspect the similarities and the differences among WM and IgMMGUS, we computed the Euclidean pair-wise distance between subjects and, using this distance as weight, constructed a minimum spanning tree (MST) (Figure 1). Considerably, four probesets identified ADRB2 (transmembrane Beta adrenergic receptor) which was up regulated in WM and IgMMGUS compared to CTRLs. The over expression of ADRB2 was also demonstrated in Mantle Cell Lymphoma cell lines and in Diffuse large B-cell lymphoma (DLBCL) lymphocytes compared to normal B-cells (doi10.1016/j.cellsig.2017.08.002), and in most malignancies (doi10.1007/s11033-021-06250-y) . As far as we know, ADAM23 (ADAM Metallopeptidase Domain23) has not been found in WM, whereas we suggest its possible role in WM patients with Sjogren's syndrome (SS). ADAM23 plays a role in the peripheral neuropathy by controlling the activity of potassium channels in SS (doi10.1007/s10067-016-3499-z). Some authors found that sensory/motor neuropathies were associated with MGUS patients (doi10.1017/s0317167100011483). We strongly believe that the down regulation of ADAM23 in WM and IgMMGUS has a good chance to be associated with clinical neuropathy in WM and IgMMGUS. RASGRP3 (RAS Guanyl Releasing Protein3) and PIK3AP1 (Phosphoinositide 3 Kinase Adaptor Protein1) play crucial roles in BCR signaling pathway: PIK3AP1 activates the PI3K-Akt signaling while RASGRP3 stimulates MAPK signaling pathway. The deregulation of LEF1 (Lymphoid Enhancer Binding Factor1) and genes of Wnt-pathway were previously demonstrated in B-cell disorders and multiple myeloma (doi10.1007/s00277-017-3207-3, doi10.1016/j.pathol.2019.09.009). According to these studies, we showed the under expression of LEF1 in WM and IgMMGUS compared to CTRLs. We identified the down regulation of EZH2 (Enhancer Of Zeste2 Polycomb Repressive Complex2Subunit) in WM and IgMMGUS compared to CTRLs. EZH2 is involved in Follicular lymphoma and DLBCL (doi10.1080/2162402X.2017.1321184). CDHR3 (Cadherin Related Family Member3), CHEK1 (Checkpoint Kinase1), and HIST1H1B (Histone-H1.5) were over expressed in CTRLs compared to IgMMGUS and WM. In conclusion, the common gene-set in WM and IgMMGUS could suggest two-hit hypothesis. First, the gene-set could play a role in the risk of progression of IgMMGUS to WM. Until now, all the IgMMGUS subjects have not been transformed in WM or other NHL, but they have been monitored every 6 months, and their possible transformation to lymphoma could highlight new insights. The second hypothesis suggests their involvement in the biological processes of leukemogenesis in WM and IgMMGUS which will be further investigated. Figure 1 Figure 1. Disclosures Tedeschi: AbbVie: Honoraria, Speakers Bureau; AstraZeneca: Honoraria, Speakers Bureau; Beigene: Honoraria, Speakers Bureau; Janssen: Honoraria, Speakers Bureau.

scholarly journals Ibrutinib in B-cell lymphoma: single fighter might be enough?

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Chao Xue ◽  
Xin Wang ◽  
Lingyan Zhang ◽  
Qingyuan Qu ◽  
Qian Zhang ◽  
...  
Keyword(s):  
B Cell ◽  
Signaling Pathway ◽  
Cell Lymphoma ◽  
Critical Role ◽  
B Cell Lymphoma ◽  
Cell Receptor ◽  
High Response Rate ◽  
Main Body ◽  
Combination Strategies ◽  
Bcr Signaling

Abstract Background In recent years, the B cell receptor (BCR) signaling pathway has become a “hot point” because it plays a critical role in B-cell proliferation and function. Bruton’s tyrosine kinase (BTK) is overexpressed in many subtypes of B-cell lymphoma as a downstream kinase in the BCR signaling pathway. Ibrutinib, the first generation of BTK inhibitor, has shown excellent antitumor activity in both indolent and aggressive B-cell lymphoma. Main body Ibrutinib monotherapy has been confirmed to be effective with a high response rate (RR) and well-tolerated in many B-cell lymphoma subgroups. To achieve much deeper and faster remission, combination strategies contained ibrutinib were conducted to evaluate their synergistic anti-tumor effect. Conclusions For patients with indolent B-cell lymphoma, most of them respond well with ibrutinib monotherapy. Combination strategies contained ibrutinib might be a better choice to achieve deeper and faster remission in the treatment of aggressive subtypes of B-cell lymphoma. Further investigations on the long-term efficacy and safety of the ibrutinib will provide novel strategies for individualized treatment of B-cell lymphoma.

scholarly journals Altered antigen receptor signaling and impaired Fas-mediated apoptosis of B cells in Lyn-deficient mice.

1996 ◽  
Vol 184 (3) ◽  
pp. 831-838 ◽  
Author(s):  
J Wang ◽  
T Koizumi ◽  
T Watanabe
Keyword(s):  
B Cells ◽  
B Cell ◽  
Protein Tyrosine Kinase ◽  
Plasma Cells ◽  
Cell Function ◽  
Antigen Receptor ◽  
Cd40 Ligand ◽  
Lymphoid Tissues ◽  
Cd40 Ligation ◽  
Bcr Signaling

Mice deficient in the src related protein tyrosine kinase, Lyn, exhibit splenomegaly and accumulate lymphoblast-like and plasma cells in spleen as they age, resulting in elevated levels of serum IgM (10-20-fold of control) and glomerulonephritis due to the presence of immune complexes containing auto-reactive antibodies. It remains unclear, however, how antibody-producing cells are accumulated in the lymphoid tissues of Lyn-/- mice. To elucidate the role of Lyn in B cell function, we have studied the proliferative responses to various stimuli and Fas-mediated apoptosis in B cells from young Lyn-/- mice which do not yet show apparent abnormality such as splenomegaly. Compared with control B cells, Lyn-/- B cells were hyper responsive to anti-IgM-induced proliferation and defective in Fc gamma RIIB-mediated suppression of B cell antigen receptor (BCR) signaling, indicating that Lyn is involved in the negative regulation of BCR signaling. In addition, the BCR-mediated signal in Lyn-/- B cells, unlike that in control B cells, failed to act in synergy with either CD40- or IL-4 receptor-triggered signal in inducing a strong proliferative response, suggesting that the BCR signaling pathway in Lyn-/- B cells is altered from that in control B cells. Furthermore, Lyn-/- B cells were found to be impaired in the induction of Fas expression after CD40 ligation and exhibited a reduced susceptibility to Fas-mediated apoptosis. Moreover, BCR cross-linking in Lyn-/- B cells suppressed Fas expression induced by costimulation with CD40 ligand and IL-4. Collectively, these results suggest that the accumulation of lymphoblast-like and plasma cells in Lyn-/- mice may be caused in part, by the accelerated activation of B cells in the absence of Lyn, as well as the impaired Fas-mediated apoptosis after the activation.

Gene Expression Profiling of Peripheral T-Cell Lymphoma (PTCL, NOS) and Comparison to Diffuse Large B-Cell Lymphoma (DLBCL).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2277-2277
Author(s):  
Daruka Mahadevan ◽  
Catherine Spier ◽  
Kimiko Della Croce ◽  
Susan Miller ◽  
Benjamin George ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lymph Node ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Gene Expression Profiling ◽  
Real Time ◽  
Expression Profiling ◽  
Cell Lymphoma ◽  
Rt Pcr

Abstract Background: WHO classifies NHL into B (~85%) and T (~15%) cell subtypes. Of the T-cell NHL, peripheral T-cell NHL (PTCL, NOS) comprises ~6–10% with an inferior response and survival to chemotherapy compared to DLBCL. Gene Expression Profiling (GEP) of DLBCL has provided molecular signatures that define 3 subclasses with distinct survival rates. The current study analyzed transcript profiling in PTCL (NOS) and compared and contrasted it to GEP of DLBCL. Methods : Snap frozen samples of 5 patients with PTCL (NOS) and 4 patients with DLBCL were analyzed utilizing the HG-U133A 2.0 Affymetrix array (~18,400 transcripts, 22,000 probe sets) after isolating and purifying total RNA (Qiagen, RNAeasy). The control RNA samples were isolated from normal peripheral blood (PB) B-cell (AllCell, CA), normal PB T-cell (AllCell, CA) and normal lymph node (LN). Immunohisto-chemistry (IHC) confirmed tumor lineage and quantitative real time RT-PCR was performed on selected genes to validate the microarray study. The GEP data were processed and analyzed utilizing Affymetrix MAS 5.0 and GeneSpring 5.0 software. Our data were analyzed in the light of the published GEP of DLBCL (lymphochip and affymtrix) and the validated 10 prognostic genes (by IHC and real time RT-PCR). Results : Data are represented as “robust” increases or decreases of relative gene expression common to all 5 PTCL or 4 DLBCL patients respectively. The table shows the 5 most over-expressed genes in PTCL or DLBCL compared to normal T-cell (NT), B-cell (NB) and lymph node (LN). PTCL vs NT PTCL vs LN DLVCL vs NB DLBCL vs LN COL1A1 CHI3L1 CCL18 CCL18 CCL18 CCL18 VNN1 IGJ CXCL13 CCL5 UBD VNN1 IGFBP7 SH2D1A LYZ CD52 RARRES1 NKG7 CCL5 MAP4K1 Of the top 20 increases, 3 genes were common to PTCL and DLBCL when compared to normal T and B cells, while 11 were common when compared to normal LN. Comparison of genes common to normal B-cell and LN Vs DLBCL or PTCL and normal T-cell and LN Vs PTCL or DLBCL identified sets of genes that are commonly and differentially expressed in PTCL and/or DLBCL. The 4 DLBCL patients analyzed express 3 of 10 prognostic genes compared to normal B-cells and 7 of 10 prognostic genes compared to normal LN and fall into the non-germinal center subtype. Quantitative real time RT-PCR on 10 functionally distinct common over-expressed genes in the 5 PTCL (NOS) patients (Lumican, CCL18, CD14, CD54, CD106, CD163, α-PDGFR, HCK, ABCA1 and Tumor endothelial marker 6) validated the microarray data. Conclusions: GEP of PTCL (NOS) and DLBCL in combination with quantitative real time RT-PCR and IHC have identified a ‘molecular signature’ for PTCL and DLBCL based on a comparison to normal (B-cell, T-cell and LN) tissue. The categorization of the GEP based on the six hallmarks of cancer identifies a ‘tumor profile signature’ for PTCL and DLBCL and a number of novel targets for therapeutic intervention.

Elevated Expression of GPR34 and Its Association with a Novel Translocation T(X;14)(p11;q32) Involving IGHS and GPR34 in MALT Lymphoma.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2251-2251 ◽  
Author(s):  
Anne Novak ◽  
Takashi Akasaka ◽  
Michelle Manske ◽  
Mamta Gupta ◽  
Thomas Witzig ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Signaling Pathway ◽  
Malt Lymphoma ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Fold Increase ◽  
Switch Region ◽  
Cell Control ◽  
Interphase Fish

Abstract MALT lymphoma is a genetically unique disorder and five mutually exclusive chromosomal translocations have been identified thus far in this disease: t(11;18), t(14;18), t(1;14), t(1;2) and t(3;14). Despite this genetic heterogeneity, all but one of the translocations affect the NF-κB signaling pathway, which is critical if not essential for antigen receptor mediated B- and T-cell activation and likely enhances MALT lymphoma growth. However, the known translocations are present in only 25% of cases suggesting that additional uncharacterized translocations exist. We used long-distance inverse polymerase chain reaction (LDI-PCR) technique to clone a novel IGH translocation partner in a 60-year-old female with a history of Sjogren’s syndrome and primary MALT lymphoma involving the parotid gland. The breakpoint on chromosome 14 occurred within the IGHSA2 switch region while the breakpoint on chromosome X fell within Xp11.4. The breakpoint on chromosome Xp11.2 fell between two genes, GPR82 and GPR34, both of which code for orphan G-protein coupled receptors (GPCRs). The breakpoint also fell within and disrupted a larger gene called CASK (a membrane-associated guanylate kinase-MAGUK). To determine the prevalence of this translocation in MALT lymphoma we performed interphase FISH studies on 64 MALT lymphomas using a breakapart probe for GPR82. Only the index case had an abnormal split signal pattern. We next designed primers to perform real-time quantitative RT-PCR for the genes located on Xp11 and found that GPR34 RNA was highly expressed, a 49-fold increase, in the patient carrying the t(X;14)(p11.4;q32) translocation compared to a normal splenic B cell control. Expression of GPR82 and CASK RNA was similar between normal B cells and the patient carrying the t(X;14)(p11.4;q32) translocation. These data suggest that the GPR34 gene is dysregulated upon its translocation to the IGHSA2 switch region. We then measured GPR34 RNA expression in a panel of MALT lymphomas (n=12) and found that GPR34 was expressed at levels higher than that seen in normal B cells with an average increase of 11 fold, 9/11 of which had an expression lever greater than 3-fold over normal splenic B cells. In a gastric MALT lymphoma specimen arising in a 67-year-old male, we saw a 64 fold increase in GPR34 expression. Interphase FISH studies performed on this specimen showed an extra intact GPR34 signal but no translocation involving IGH or GPR34, suggesting that other mechanisms, including gene dosage effect, can upregulate GPR34. GPR34 RNA was also detected in other normal B cell populations and histologic subtypes of NHL but not to the extent seen in MALT lymphoma; values represent the expression of GPR34 normalized to β-actin and a normal B cell control (value of 1.0): t(X;14)(p11.4;q32) specimen, 49.0; MALT lymphoma, 11.0 (n=12); peripheral blood B cells, 0.48 (n=2); normal bone marrow B cells, 0.97 (n=3); follicular lymphoma, 2.47 (n=3); marginal zone B cell lymphoma, 2.47 (n=3); diffuse large cell lymphoma, 0.36, (n=3); mantle cell lymphoma, 2.67 (n=3); and multiple myeloma 0.52 (n=6). The receptor encoded by GPR34 is most similar to the PY2 receptor subfamily of GPCR and GPR34 mRNA transcripts are particularly abundant in mast cells while lower levels were detected in other immune cells including B cells. However, little is known about its natural ligand, biologic function, or the signaling cascades activated by its engagement. Because the NF-κB signaling pathway has been shown to be a common downstream target of MALT lymphoma translocations we first examined the impact of GPR34 expression on phosphorylation of Iκ-Bα. Transient expression of a YFP-GPR34 expression plasmid in HeLa cells results in increased phosphorylation of Iκ-Bα compared to the YFP control. Additionally, we observed increased phosphorylation of ERK1/2 in GPR34-expressing cells, however no change in phosphorylation of GSK3β was detected. In summary, these data identify a novel IGHS translocation partner in MALT lymphoma and suggest that dysregulation of GPR34 is commonly found in MALT lymphoma. Overexpression of GPR34 results in activation of the NF-κB and MAP kinase pathways and may be a novel mechanism by which MALT lymphoma occurs.

Defining The Role Of Microrna-146a In B Cell Lymphomagenesis

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3805-3805
Author(s):  
Jorge Contreras ◽  
Jayanth Kumar Palanichamy ◽  
Tiffany Tran ◽  
Dinesh S. Rao
Keyword(s):  
Gene Expression ◽  
B Cells ◽  
B Cell ◽  
Conflicts Of Interest ◽  
Cell Lymphoma ◽  
Expression Patterns ◽  
Significant Proportion ◽  
B Cell Lymphoma ◽  
Gene Expression Patterns

Abstract Diffuse large B cell lymphoma (DLBCL) is one of the most common Non-Hodgkin lymphomas among adults. It is a heterogeneous disease characterized by multiple mutations and translocations. Gene expression profiling studies have revealed several characteristic gene expression patterns, with two main patterns emerging, namely Germinal Center(GC) type, and Activated B Cell (ABC) type. ABC-type DLBCL shows gene expression patterns that resemble activated B-cells, with increased expression of anti-apoptotic, and pro-proliferative genes. Critically, upregulation of the NF-κB the pathway is a hallmark of ABC-type DLBCL and has been shown to be necessary for survival, and is caused by several different mutations at different levels within the pathway. Recent work has revealed the critical importance of a new class of small RNA molecules, namely microRNAs, in gene regulation. Of these, microRNA-146a (miR-146a) was discovered as an NF-κB induced microRNA that plays a role as a negative feedback regulator of this pathway by targeting adaptor proteins. To further characterize miR-146a, mice deficient for this miRNA were created, and were found to develop lymphadenopathy, splenomegaly, and myeloid proliferation. As expected, immune cells in these mice have an upregulated NF-κB pathway and many of the phenotypes can be ameliorated by inhibition of the NF-κB pathway. Importantly, a significant proportion of the animals develop B-cell lymphoma at older ages. In this study, we examined the role of miR-146a in the development of malignancy in B-cells. To accelerate the role of miR-146a in tumor formation we overlaid the miR-146a deficient allele onto the Eμ-Myc like mouse model. Eμ-Myc mice develop tumors on average by 14weeks of age. The transgenic status of animals was verified by genotyping, RNA and protein expression analyses. miR-146a sufficient and deficient animals on the Eμ-Myc background were followed for tumor latency by peripheral blood analysis and careful physical examination. Based on approved humane criteria for animal discomfort, animals were sacrificed and hematopoietic tissue was harvested for analysis. Mice deficient for miR-146a had a statistically reduced survival in comparison with miR-146a sufficient animals with a p-value of .0098 (Kaplan Meir survival analysis). Complete Blood Count of animals at time of death revealed an increase leukemia presentation in the miR-146a deficient background. FACS analysis of tumor tissue from both groups revealed an increase in the number of IgM positive tumors in the miR-146a-deficient background indicating skewing towards more mature B cell neoplasms when miR-146a is lacking. Lineage analysis of tumors verified them to be of B cell origin although a subset of miR-146a sufficient tumors had higher numbers of infiltrating myeloid cells compared to deficient animals. Furthermore, histologic analysis of hematopoietic organs showed that while infiltration remained similar in kidneys and liver, more spleens in the miR-146a deficient background tended to be less involved. Our extensive histopathologic and immunophenotypic analyses indicate that miR-146a deficiency drives a more aggressive malignant phenotype in the B-cell lineage. In keeping with this, our profiling studies of human DLBCL suggest that a subset of DLBCL show decreased expression of miR-146a. We are currently examining the status of NF-κB in the murine tumors and using high throughput sequencing approaches to delineate gene expression differences between miR-146a sufficient and deficient tumors. We anticipate the discovery of novel gene targets of miR-146a and expect that these studies will lead to improved diagnostic and therapeutic options for patients of B-cell malignancies. Disclosures: No relevant conflicts of interest to declare.

The Role of PIM1 in the Ibrutinib-Resistant ABC Subtype of Diffuse Large B-Cell Lymphoma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 699-699 ◽  
Author(s):  
Hsu-Ping Kuo ◽  
Sidney Hsieh ◽  
Karl J. Schweighofer ◽  
Leo WK Cheung ◽  
Shiquan Wu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Tumor Growth ◽  
B Cell ◽  
Repeated Measures ◽  
Cell Lymphoma ◽  
Xenograft Model ◽  
B Cell Lymphoma ◽  
Bcr Signaling

Abstract Introduction: Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma (NHL), accounting for roughly 30% of newly diagnosed cases in the United States (US). DLBCL is a heterogeneous lymphoma, including the activated B cell-like (ABC) and germinal center B cell-like (GCB) subtypes, which have different gene expression profiles, oncogenic aberrations, and clinical outcomes (Alizadeh, Nature 2000; Staudt, Adv Immunol 2005). ABC-DLBCL is characterized by chronic active B-cell receptor (BCR) signaling (Davis, Nature 2010), which is required for cell survival. Thus, the BCR signaling pathway is an attractive therapeutic target in this type of B-cell malignancy. Bruton's tyrosine kinase (BTK), which plays a pivotal role in BCR signaling, is covalently bound with high affinity by ibrutinib, a first-in-class BTK inhibitor approved in the US for mantle cell lymphoma and chronic lymphocytic leukemia (CLL) patients (pts) who have received at least one prior treatment, CLL with del17p, and WaldenstršmÕs macroglobulinemia. A recent phase 2 clinical trial of single-agent ibrutinib in DLBCL pts revealed an overall response rate of 40% for ABC-DLBCL (Wilson, Nat. Med 2015); however, responses to single kinase-targeted cancer therapies are often limited by the cellÕs ability to bypass the target via alternative pathways or acquired mutations in the target or its pathway (Nardi, Curr Opin Hematol 2004; Gazdar, Oncogene 2009). The serine/threonine-protein kinase PIM1 is one of several genes exhibiting differential expression in ibrutinib-resistant ABC-DLBCL cells compared with wild-type (WT) cells. We identified and report herein the role of PIM1 in ABC-DLBCL ibrutinib-resistant cells. Methods: PIM1 gene expression was analyzed by RT-qPCR. In vitro, cell viability was assessed in the human ABC-DLBCL cell line HBL-1 after treatment with ibrutinib and/or a pan-PIM inhibitor for 3 days, and the effect on colony formation was determined 7 days post-treatment. PIM1 mutational analysis was performed with clinical tumor biopsy samples from 2 studies, PCYC-04753 (NCT00849654) and PCYC-1106-CA (NCT01325701). PIM1 protein stability was analyzed by treating cells with cycloheximide and examining protein levels at different time points up to 8 hours. Results: Gene expression profiling of ibrutinib-resistant ABC-DLBCL cells revealed an upregulation of PIM1 (15-fold increase compared with WT cells) as well as PIM2 and PIM3. We also found that, compared with single-drug treatment, in vitro cell growth could be synergistically suppressed with a combination of ibrutinib and a pan-PIM inhibitor. This effect was observed in both WT (combination index (C.I.) = 0.25; synergy score = 3.18) and ibrutinib-resistant HBL-1 cells (C.I. = 0.18; synergy score = 4.98). In HBL-1 cells, this drug combination reduced colony formation and suppressed tumor growth in a xenograft model (Figure 1). In 48 DLBCL patient samples with available genomic profiling, PIM1 mutations appeared more frequently in pts diagnosed with ABC-DLBCL compared with GCB-DLBCL (5 out of 6 DLBCL pts with PIM1 mutations were ABC-subtype). 4 of these 5 pts exhibited a poor clinical response to ibrutinib, ie, 80% of ABC-DLBCL pts with PIM1 mutations had progressive disease, compared with only 13 of 26 (ie, 50%) ABC-DLBCL pts without PIM1 mutations. Subsequent characterization of the mutant PIM1 proteins (L2V, P81S, and S97N) confirmed that they were more stable than WT PIM1, suggesting increased protein levels by 2 potential mechanisms (WT PIM1 gene up-regulation or increased mutant PIM1 protein half-life). The impact of these mutations on PIM1 function and ibrutinib sensitivity is under investigation. Conclusions: Ibrutinib-resistant ABC-DLBCL cells have increased PIM1 expression, and synergistic growth suppression was observed when ibrutinib was combined with a pan-PIM inhibitor. PIM1 mutations identified in ABC-DLBCL pts with poor responses to ibrutinib contributed to increased PIM1 protein stability. A better understanding of the role of PIM1 in ibrutinib-resistant ABC-DLBCL tumors could provide a rationale for the design of combination therapies. Figure 1. Combination of ibrutinib and a pan-PIM inhibitor in the HBL-1 xenograft model. Ibrutinib and PIM inhibitor treatment suppressed tumor growth by 62% compared with the vehicle-treated group (*p < 0.01, repeated measures MANOVA adjusted univariate F-test). Figure 1. Combination of ibrutinib and a pan-PIM inhibitor in the HBL-1 xenograft model. Ibrutinib and PIM inhibitor treatment suppressed tumor growth by 62% compared with the vehicle-treated group (*p < 0.01, repeated measures MANOVA adjusted univariate F-test). Disclosures Kuo: Pharmacyclics LLC, an AbbVie Company: Employment. Hsieh:pharmacyclics LLC, an AbbVie Company: Employment. Schweighofer:Pharmacyclics LLC, an AbbVie Company: Employment. Cheung:Pharmacyclics LLC, an AbbVie Company: Employment. Wu:Pharmacyclics LLC, an AbbVie Company: Employment. Apatira:Pharmacyclics LLC, an AbbVie Company: Employment. Sirisawad:Pharmacyclics LLC, an AbbVie Company: Employment. Eckert:Pharmacyclics LLC, an AbbVie Company: Employment. Liang:Pharmacyclics LLC, an AbbVie Company: Employment. Hsu:Pharmacyclics LLC, an AbbVie Company: Employment. Chang:Pharmacyclics LLC, an AbbVie Company: Employment.

scholarly journals A Signaling Pathway Mediating Downregulation of BCL6 in Germinal Center B Cells Is Blocked by BCL6 Gene Alterations in B Cell Lymphoma

Cancer Cell ◽  
2007 ◽  
Vol 12 (3) ◽  
pp. 280-292 ◽  
Author(s):  
Masumichi Saito ◽  
Jie Gao ◽  
Katia Basso ◽  
Yukiko Kitagawa ◽  
Paula M. Smith ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Signaling Pathway ◽  
Germinal Center ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Gene Alterations ◽  
Germinal Center B Cells

scholarly journals IgM and IgD B cell receptors differentially respond to endogenous antigens and control B cell fate

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Mark Noviski ◽  
James L Mueller ◽  
Anne Satterthwaite ◽  
Lee Ann Garrett-Sinha ◽  
Frank Brombacher ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Fate ◽  
Plasma Cells ◽  
Autoreactive B Cells ◽  
Binding Domains ◽  
Bcr Signaling ◽  
And Control

Naive B cells co-express two BCR isotypes, IgM and IgD, with identical antigen-binding domains but distinct constant regions. IgM but not IgD is downregulated on autoreactive B cells. Because these isotypes are presumed to be redundant, it is unknown how this could impose tolerance. We introduced the Nur77-eGFP reporter of BCR signaling into mice that express each BCR isotype alone. Despite signaling strongly in vitro, IgD is less sensitive than IgM to endogenous antigen in vivo and developmental fate decisions are skewed accordingly. IgD-only Lyn−/− B cells cannot generate autoantibodies and short-lived plasma cells (SLPCs) in vivo, a fate thought to be driven by intense BCR signaling induced by endogenous antigens. Similarly, IgD-only B cells generate normal germinal center, but impaired IgG1+ SLPC responses to T-dependent immunization. We propose a role for IgD in maintaining the quiescence of autoreactive B cells and restricting their differentiation into autoantibody secreting cells.

scholarly journals Co-Expression of SYK and ZAP70 Subverts Negative B-Cell Selection and Enables Oncogenic Signaling in Multiple B-Cell Malignancies

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 295-295
Author(s):  
Teresa Sadras ◽  
Mickaël Martin ◽  
Lauren Kim-Sing ◽  
Jevon Cutler ◽  
Gal Lenz ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
B Cells ◽  
B Cell ◽  
Cell Lymphoma ◽  
Cell Selection ◽  
B Cell Malignancies ◽  
Close Proximity ◽  
Mantle Cell ◽  
Bcr Signaling

B-cells are under intense selective pressure to eliminate autoreactive or premalignant clones. B-cell receptor (BCR) signals are required for survival, however, BCR-signaling exceeding maximum thresholds often reflects signaling from an autoreactive BCR or a transforming oncogene and triggers negative selection and cell death. The tyrosine kinase SYK initiates BCR-downstream signaling in B-cells while its close relative ZAP70 is almost exclusively expressed in T-cells. Interestingly, the segregation of SYK to B-cells and ZAP70 to T-cells is less confined in malignant lymphopoiesis suggesting that the balance of these related kinases may alter signaling output in disease and contribute to development of leukemia. As previously shown in B-cell chronic lymphocytic leukemia (B-CLL), we identified aberrant ZAP70 expression as a frequent feature in multiple other B-cell malignancies that depend on survival signals from a functional (pre-) BCR (E2A-PBX1+ pre-B ALL, and mantle cell lymphoma) or harbor oncogenic mimics of the BCR (BCR-ABL1+ B-ALL). Studying SYK and ZAP70 expression by single-cell Western blot, co-expression of the two tyrosine kinases was extremely rare in normal B- and T-cell populations. In contrast, &gt;50% of tumor B-cells in mantle cell lymphoma, pre-B ALL and CLL co-expressed SYK and ZAP70. Despite their structural similarities, genetic deletion and engineered reconstitution of SYK and ZAP70 in human B-cell lymphoma cells revealed striking functional differences. Proximity-dependent biotin identification (BioID) analyses identified that SYK, but not ZAP70, engaged the PI3K pathway via interaction with CD19. Consistent with this, reconstitution with SYK and SYK-ZAP70 but not ZAP70 alone promoted survival and proliferation. Detailed analysis of BCR-mediated cascades in lymphoma cells expressing SYK, ZAP70 or SYK-ZAP70 established that ZAP70 is only weakly efficient at propagating BCR-mediated calcium and downstream pathway activation in B-cells. Strikingly, co-expression of ZAP70 with SYK resulted in re-wired BCR-signaling of intermediate strength: compared to cells expressing only SYK, SYK-ZAP70 co-expressing cells had markedly reduced activation of the BLNK-BTK-PLCγ pathway, further reflected in BCR-induced Ca2+ signaling with delayed onset, lower amplitude but longer duration. In this way, we speculated that SYK and ZAP70 may be present within close proximity at the apex of BCR-initiated interactions, and hence compete for downstream substrates resulting in a re-wiring of classic signaling programs propagated normally by SYK. To explore this, we utilized proximity ligation assays (PLA) to monitor the proximity of SYK and ZAP70 in resting or BCR-stimulated B-cells, and found that SYK and ZAP70 co-exist within close proximity consistent with the view that varying levels of these kinases may alter B-cell signaling output. Functional experiments further showed that phosphomimetic activation of SYK, but not ZAP70, induced hyperactivation of PI3K-signaling and acute BTK-mediated cell death in pre-B ALL cells. In line with altered BCR-signaling strength and quality in SYK and ZAP70 co-expressing cells, over-expression of Zap70 in pre-B ALL cells rescued auto-immune checkpoint activation induced by hyper-activation of BCR-associated signaling. To study functional consequences of SYK-ZAP70 co-expression during normal B-cell development, we generated a novel knock in Zap-70+/Mb1-Cre+mouse model, to induce conditional expression of Zap70 in the B cell compartment from the proB stage. Consistent with compromised central tolerance checkpoints, Syk-Zap70 co-expressing pro/pre-B and immature B-cells had reduced spontaneous apoptosis rates and gave rise to autoantibody production against multiple self-antigens. Importantly, our findings highlight a previously unrecognized role for ZAP70 in oncogenic BCR-signaling and we conclude that the co-expression of ZAP70 mitigates the ability of SYK, downstream of an autoreactive BCR or a transforming oncogene, to trigger negative B-cell selection and cell death (Figure 1). Disclosures Weinstock: Celgene: Research Funding. Meffre:AbbVie: Consultancy, Other: Grant.

Gene Expression Profiling of Mantle Cell Lymphoma in the Leukemic Phase Reveals Aberrant Expression of Genes from the TGF-β Signaling Pathway.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2048-2048
Author(s):  
Edgar G. Rizzatti ◽  
Rodrigo A. Panepucci ◽  
Rodrigo Proto-Siqueira ◽  
Wilma T. Anselmo-Lima ◽  
Oswaldo K. Okamoto ◽  
...  
Keyword(s):  
Gene Expression ◽  
B Cells ◽  
Cyclin D1 ◽  
Signaling Pathway ◽  
Peripheral Blood ◽  
Cell Lymphoma ◽  
Aberrant Expression ◽  
Mantle Cell ◽  
Leukemic Phase ◽  
Activin Receptors

Abstract Mantle cell lymphoma (MCL) is a distinctive subtype of B-cell lymphoma associated with the t(11;14)(q13;32) and consequent ectopic overexpression of cyclin D1 in the tumor cells. Disease is predominantly disseminated at diagnosis and a frank leukemic phase is detected in one fourth of patients. Ontogenetically, MCL is considered the malignant counterpart of pre-germinal-center naive B-cells. Although the overexpression of cyclin D1 plays a pivotal role on the pathogenesis of MCL, studies with transgenic mice have shown that it is not sufficient by itself to cause lymphoma, and a better understanding of the molecular genetics of this disease may provide insights toward a potentially curable therapy. To address this issue, we compared the gene expression profile of MCL and normal naive B-cells using oligonucleotide microarrays representing 10,000 genes. MCL cells and naive B-cells (IgD+CD38±CD27−) were highly purified, by magnetic activated cell sorting, from the peripheral blood of patients with MCL in the leukemic phase and from tonsils of normal controls, respectively (purity > 95% in all samples). Three individuals were selected for each group and experiments were performed in replicates using the Amersham CodeLink Human UniSet I Bioarrays. For validation purposes, the expression of 10 selected genes (6 overexpressed and 4 underexpressed in lymphoma cells) was quantified by TaqMan real-time RT-PCR in non-purified peripheral blood samples from 25 patients with MCL in the leukemic phase and compared with normal naive B-cells, with fully concordant results. Data mining from our microarray results revealed an aberrant expression of several genes from the TGF-β signaling pathway in MCL (p<0.01): ACVR1 (fold change = 2.5), ACVR2 (2.9), ACVR2B (16.3), BMP4 (11.8), TGIF (4.0), Smad2 (3.4) and Smad6 (0.6). Except for TGIF and Smad6, all other genes induce the TGF-β signaling pathway. Although TGIF was overexpressed, it depends on the relative levels of Smad co-repressors or co-activators to exert its inhibitory activity; whereas Smad6, which is also an inhibitory mediator, was underexpressed. The activin receptors ACVR1, ACVR2 and ACVR2B are receptors of the TGF-β superfamily, which consists of TGF-β, activins, bone morphogenic proteins (BMPs) and others. Upon ligand binding, activin receptors induce anti-proliferative and pro-apoptotic responses, acting as tumor suppressors in early tumorigenesis. In advanced cancer, however, there is a loss of growth-inhibitory responsiveness downstream the core TGF-β signaling pathway, and it may be used as a tumor-progression factor by inducing immune supression, angiogenesis, epithelial-mesenchymal transdifferentiation and increased potential for metastasis. Interestingly, the cyclin D1/TGF-β double transgenic liver model in mice (Deane et al. Cancer Res.2004; 64:1315) showed enhanced tumor formation when compared with its single transgenic littermates. Our results suggest an activation of the TGF-β signaling pathway in MCL, and point toward potential new therapeutic targets for this yet incurable lymphoma.

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