Cell Lymphomagenesis Research Articles

PO-059 IBTK promotes B cell lymphomagenesis in Eμ-MYC transgenic mice conferring resistance to apoptosis

IntroductionAlbeit the advancement in breast cancer (BC) treatment, BC remains as the second leading cause of cancer death worldwide. Unlike the other BC subtypes, the lack of hormone receptor (HR) rendering the aggressive triple negative BC (TNBC) towards chemotherapy which is destructive to one’s quality of life. Tumour necrosis factor related apoptosis-inducing ligand (TRAIL) is a promising cancer therapeutic agent due to its selective killing of malignant cells leaving the normal cells unharmed. Despite being TRAIL-sensitive, the TNBC MDA-MB-231 cells develop resistance to TRAIL after repeated treatment. The epigenetic drugs zebularine (Zeb) and trichostatin A (TSA) were reported to increase the TRAIL-induced apoptosis. Therefore, we aim to study the combinational anti-cancer effect of Zeb and TSA followed by TRAIL (TZT treatment) and compare its effect with the standard chemotherapy drug doxorubicin (DOX).Material and methodsMDA-MB-231, MCF-7 (HR positive breast cancer cells) and MCF-10A (normal breast epithelial cells) were treated with TZT and DOX respectively and subjected to cell proliferation, cell cycle arrest and apoptosis analyses.Results and discussionsBased on the results obtained, TZT treatment hampered the cell proliferation in MDA-MB-231 and MCF-7 as compared to TRAIL treatment alone but not in MCF-10A. Nevertheless, the cell proliferation of both breast cancer and normal cells treated with DOX was significant reduced. Subsequently, the cell cycle distribution was evaluated via flow cytometry analysis. Higher induction of G1 cell cycle arrest was observed in the MDA-MB-231 treated with TZT (57.5%) as compared to TRAIL alone (52.2%) at 24 hours. In MCF-7, there is the minimal induction of G1 arrest with TZT treatment (71.2%) as compared to TRAIL alone (69.7%) at 24 hours. However, DOX treatment induced G2/M arrest in breast cancer and normal cells. Furthermore, the morphological features of apoptosis were determined by haematoxylin and eosin staining. Intriguingly, the hallmark of apoptosis was clearly observed in breast cancer treated with TZT compared to MCF-10A. The results suggest the significance of Zeb and TSA to foster the TRAIL-induced apoptosis in TNBC but not in normal breast epithelial cells, despite further molecular analyses are requisite to validate the enhancement of apoptosis induction.ConclusionThis research provides an insight of potential alternative of TZT in treating the TNBC to improve the patients’ prognosis in the near future.

Molecular Insights Into Pathogenesis of Peripheral T Cell Lymphoma: a Review

Peripheral T cell lymphoma (PTCL) is a heterogeneous group of lymphoproliferative neoplasms, with at least 29 distinct entities described in current WHO classification. Using present diagnostic approaches, more than a third of PTCL cases cannot be classified, hence designated as PTCL-not otherwise specified (PTCL-NOS). Herein, we summarize the current genomic findings and their role in the molecular pathogenesis in different PTCL entities. Gene expression profiling (GEP) studies have identified distinct molecular signatures for accurate diagnosis and elucidated oncogenic pathways enriched in major PTCL entities. Furthermore, genomic characterization has identified recurrent somatic mutations and potential therapeutic targets. Further efforts are underway to develop genetically faithful murine models. GEP studies have identified molecular subgroups of PTCL, characterized by distinct genetic and epigenetic alterations. Understanding the molecular mechanisms of T cell lymphomagenesis using in vivo model will help to reveal novel therapeutic targets.

PO-130 SER235 residue drives eIF6 oncogenic activity in NPM-ALK induced T cell lymphomagenesis

Introduction Dysregulation of mRNA translational control in cancer leads to cell transformation, metabolic reprogramming and angiogenesis. eIF6 is an oncogenic translation factor, which regulates the initiation phase of translation acting on 60S availability in the cytoplasm and controlling active 80S complex formation. eIF6 activation is mTORC1-independent and driven by PKCβ mediated phosphorylation on Ser235. An increment of eIF6 expression is reported in several cancer cell lines and human tumours, due to amplification or overexpression. In mice, eIF6 haploinsufficiency blocks Myc-driven lymphomagenesis. Intriguingly, high levels of PKC and eIF6 are found in T-cell lymphomas. In particular, in Anaplastic Large Cell Lymphoma (ALCL) eIF6 is overexpressed and hyperactivated. Material and methods Here, we aimed to define the role of eIF6 phosphorylation in NPM-ALK mediated T-cell lymphomagenesis, combining multidisciplinary studies on murine and cellular models. We used a conditional eIF6 SA KI mouse model in which Ser235 is replaced by an Ala. Results and discussions First, we addressed the effect of eIF6 mutated protein expression in all tissues: homozygosity is lethal after gastrulation while heterozygous mice are viable but resistant to NPM-ALK driven lymphomagenesis. Then, we investigated the role of Ser235 phosphorylation specifically in T-cell lineage, crossing eIF6 SA KI mice with CD4-Cre mice. Physiological T-cell development and subsets composition are not affected by the eIF6 mutated protein. In cancer, eIF6 SA/SA CD4-Cre NPM-ALK mice have a significant increase in survival time, compared to wt with a delay in the appearance of lymphoma up to 6 months. Histological analysis and ex vivo cultures confirm the delay in disease development. eIF6 SA/SA CD4-Cre NPM-ALK thymocytes are smaller respect to wt counterparts and show a striking senescence-like phenotype in vitro . Similarly, in vitro generated eIF6 SA/SA MEFs show a markedly reduced proliferation and increased SA β-gal positivity. This phenotype is completely rescued by transducing eIF6 wild-type, but not by eIF6 SA . Currently, we are investigating the molecular mechanisms by which eIF6 phosphorylation affects ALK-induced malignancy and whether it may modulate premature cell senescence, thus establishing an effective barrier to T-cell lymphomagenesis. Conclusion Our work demonstrates for the first time that eIF6 phosphorylation plays an essential role in mammals development, cell homeostasis and is rate-limiting for T-cell lymphomagenesis in vivo .

Differential effects of Vav-promoter-driven overexpression of BCLX and BFL1 on lymphocyte survival and B cell lymphomagenesis.

Overexpression of BCLX and BFL1/A1 has been reported in various human malignancies and is associated with poor prognosis and drug resistance, identifying these prosurvival BCL2 family members as putative drug targets. We have generated transgenic mice that express human BFL1 or human BCLX protein throughout the haematopoietic system under the control of the Vav gene promoter. Haematopoiesis is normal in both the Vav‐BFL1 and Vav‐BCLX transgenic (TG) mice and susceptibility to spontaneous haematopoietic malignancies is not increased. Lymphoid cells from Vav‐BCLX TG mice exhibit increased resistance to apoptosis in vitro while most blood cell types form Vav‐BFL1 TG mice were poorly protected. Both transgenes significantly accelerated lymphomagenesis in Eμ‐MYC TG mice and, surprisingly, the Vav‐BFL1 transgene was the more potent. Unexpectedly, expression of transgenic BFL1 RNA and protein is significantly elevated in B lymphoid cells of Vav‐BFL1/Eμ‐MYC double‐transgenic compared to Vav‐BFL1 mice, even during the preleukaemic phase, providing a rationale for the potent synergy. In contrast, Vav‐BCLX expression was not notably different. These mouse models of BFL1 and BCLX overexpression in lymphomas should be useful tools for the testing the efficacy of novel human BFL1‐ and BCLX‐specific inhibitors.

The Future of Combination Therapies for Peripheral T Cell Lymphoma (PTCL).

Peripheral T cell lymphoma is a rare heterogeneous group of diseases which are characterized by poor outcomes to treatment and short overall survival. In the past decade, several new therapies targeting T cell biology have been approved in the relapsed setting. These new therapies, such as pralatrexate, romidepsin, belinostat, and brentuximab vedotin, have begun to make their way into practice. Despite these advances, outcomes have not changed dramatically. In recent years, efforts have been made to incorporate these new therapies into combination strategies to treat this challenging disease entity. Herein we will review some of the latest developments. With the new WHO classification, discrete entities of PTCL are now being identified by molecular and phenotypic markers. This new classification is critical to our ability to define disease entities which may respond to certain classes of targeted therapy. Some such mutations include genes controlling epigenetics (TET2, IDH2, DNMT3A, RHOA, CD28). As such, epigenetic therapies such as histone deacetylase (HDAC) inhibitors have become the platform to which other novel therapies or chemotherapy has been added. Early phase clinical studies have demonstrated that combination therapy with romidepsin plus other agents known to have activity in T cell lymphoma have enhanced clinical benefit for this group of diseases. In addition, the antibody drug conjugate, brentuximab vedotin has been shown to have potent activity in T cell lymphomas expressing CD30. This drug is being studied as well with other targeted therapies and chemotherapy in an effort to improve response rates and progression-free survival. Although T cell lymphomas remain a highly challenging group of diseases to treat, new efforts to leverage drugs that discretely target the biology that drives T cell lymphomagenesis in combination provide hope that improved outcomes may be realized in the near future.

Checkpoint kinase 1 is essential for normal B cell development and lymphomagenesis

Checkpoint kinase 1 (CHK1) is critical for intrinsic cell cycle control and coordination of cell cycle progression in response to DNA damage. Despite its essential function, CHK1 has been identified as a target to kill cancer cells and studies using Chk1 haploinsufficient mice initially suggested a role as tumor suppressor. Here, we report on the key role of CHK1 in normal B-cell development, lymphomagenesis and cell survival. Chemical CHK1 inhibition induces BCL2-regulated apoptosis in primary as well as malignant B-cells and CHK1 expression levels control the timing of lymphomagenesis in mice. Moreover, total ablation of Chk1 in B-cells arrests their development at the pro-B cell stage, a block that, surprisingly, cannot be overcome by inhibition of mitochondrial apoptosis, as cell cycle arrest is initiated as an alternative fate to limit the spread of damaged DNA. Our findings define CHK1 as essential in B-cell development and potent target to treat blood cancer.

Loss of functional Miz-1 impairs C-Myc-dependent B cell lymphomagenesis by interfering with proteasome activity

Loss of functional Miz-1 impairs C-Myc-dependent B cell lymphomagenesis by interfering with proteasome activity

Adoptively transferred Vγ9Vδ2 T cells show potent antitumor effects in a preclinical B cell lymphomagenesis model.

A central issue for adoptive cellular immunotherapy is overcoming immunosuppressive signals to achieve tumor clearance. While γδ T cells are known to be potent cytolytic effectors that can kill a variety of cancers, it is not clear whether they are inhibited by suppressive ligands expressed in tumor microenvironments. Here, we have used a powerful preclinical model where EBV infection drives the de novo generation of human B cell lymphomas in vivo, and autologous T lymphocytes are held in check by PD-1/CTLA-4-mediated inhibition. We show that a single dose of adoptively transferred Vδ2+ T cells has potent antitumor effects, even in the absence of checkpoint blockade or activating compounds. Vδ2+ T cell immunotherapy given within the first 5 days of EBV infection almost completely prevented the outgrowth of tumors. Vδ2+ T cell immunotherapy given more than 3 weeks after infection (after neoplastic transformation is evident) resulted in a dramatic reduction in tumor burden. The immunotherapeutic Vδ2+ T cells maintained low cell surface expression of PD-1 in vivo, and their recruitment to tumors was followed by a decrease in B cells expressing PD-L1 and PD-L2 inhibitory ligands. These results suggest that adoptively transferred PD-1lo Vδ2+ T cells circumvent the tumor checkpoint environment in vivo.

Control of B cell development and malignant transformation by microRNA-142

Abstract MicroRNA-142 (miR-142) is a versatile posttranscriptional regulator that plays a crucial role in the regulation of both innate and adaptive immune responses. Abrogation of miR-142 expression in mice results in a profound immunodeficiency that is characterized by hypoimmunoglobulinemia and failure to mount robust humoral immune responses to soluble antigen challenges. Paradoxically, miR-142−/− mice display a significant expansion of the B cell compartment due to the accumulation of immature and mature B lymphocytes. To better understand the role of miR-142 in B cell ontogenesis, we examined the early B cell development in miR-142−/− mice using the Hardy fraction analysis. We found that miR-142 deletion results in a significant increase in the frequency of the pro-B and large pre-B cells (Fractions B and C), while the pre-pro-B cell population (Fraction A) decreased. In contrast, miR-142 ablation had little effect on the frequency of small pre-B cell and immature B cell populations. Furthermore, our data suggest that the accumulation of pro-B and large pre-B cells in miR-142−/− bone marrow is linked to an increase in their survival capacity. Because genetic lesions that disrupt control of B cell maturation frequently contribute to the malignant transformation of B cells, we decided to examine the role of miR-142 dysregulation in B cell lymphomagenesis. To address this question, we bred miR-142−/− mice with Eμ-myc transgenic mice that spontaneously develop a mix of pre-B cell and mature B cell lymphomas. We found that miR-142 haploinsufficiency dramatically accelerated lymphoma development in this mouse model. Collectively, our results establish miR-142 as an essential negative regulator of B cell maturation and lymphomagenesis.

Role of Rab7 in activating NF-kB in B cells to mediate class-switching and plasma cell survival in antibody and autoantibody responses and to promote B cell lymphomagenesis

Abstract NF-kappaB activation mediates B cell class-switching to generate IgG, IgA and IgE as well as survival of plasma cells, as differentiated from B cells to secrete antibodies at a high rate. Here we have identified a B cell-intrinsic role of Rab7, a small GTPase located in endosomes, in activating NF-kappaB for class-switching and plasma cell survival. Mice that are deficient in Rab7 specifically in B cells or treated with CID 1067700, a Rab7-specific small molecule inhibitor, cannot mount T-independent or T-dependent switched antibody responses, despite normal B cell proliferation/survival and T cell/myeloid cells functions. Rab7 co-localizes, on endosomes, with the CD40 receptor that is internalized upon engagement and directly interacts with TRAF6, an adaptor of CD40 and TLR signaling for NF-kB activation, thereby forming complexes that transduce signals. Disruption of such complexes by CID 1067700 hampers NF-kappaB activation in normal B cells and abrogates NF-kappaB dysregulation in lupus B cells and B lymphoma cell lines, in which Rab7 is upregulated. Treatment with CID 1067700 reduces pathogenic IgG autoantibody responses in murine lupus models, leading to life-span extension, and prevents tumorigenesis in nude mice transplanted with B lymphoma cells. These, together with the rescue of class-switching/plasma cell survival in normal B cells and proliferation/survival of B lymphoma cells by enforced NF-kappaB activation, indicate that Rab7 plays an important role in NF-kappaB activation to mediate B cell differentiation and dysregulation and is a potential therapeutic target in lupus and B cell lymphomagenesis. Supported by NIH grants AI 079705, AI 105813/105813S1 (to P. C.) and AI 124172 (to Z. X.).

Enteropathy-associated T cell lymphoma subtypes are characterized by loss of function of SETD2.

Enteropathy-associated T cell lymphoma (EATL) is a lethal, and the most common, neoplastic complication of celiac disease. Here, we defined the genetic landscape of EATL through whole-exome sequencing of 69 EATL tumors. SETD2 was the most frequently silenced gene in EATL (32% of cases). The JAK-STAT pathway was the most frequently mutated pathway, with frequent mutations in STAT5B as well as JAK1, JAK3, STAT3, and SOCS1 We also identified mutations in KRAS, TP53, and TERT Type I EATL and type II EATL (monomorphic epitheliotropic intestinal T cell lymphoma) had highly overlapping genetic alterations indicating shared mechanisms underlying their pathogenesis. We modeled the effects of SETD2 loss in vivo by developing a T cell-specific knockout mouse. These mice manifested an expansion of γδ T cells, indicating novel roles for SETD2 in T cell development and lymphomagenesis. Our data render the most comprehensive genetic portrait yet of this uncommon but lethal disease and may inform future classification schemes.

Role of MYC in B Cell Lymphomagenesis.

B cell lymphomas mainly arise from different developmental stages of B cells in germinal centers of secondary lymphoid tissue. There are a number of signaling pathways that affect the initiation and development of B cell lymphomagenesis. The functions of several key proteins that represent branching points of signaling networks are changed because of their aberrant expression, degradation, and/or accumulation, and those events determine the fate of the affected B cells. One of the most influential transcription factors, commonly associated with unfavorable prognosis for patients with B cell lymphoma, is nuclear phosphoprotein MYC. During B cell lymphomagenesis, oncogenic MYC variant is deregulated through various mechanisms, such as gene translocation, gene amplification, and epigenetic deregulation of its expression. Owing to alterations of downstream signaling cascades, MYC-overexpressing neoplastic B cells proliferate rapidly, avoid apoptosis, and become unresponsive to most conventional treatments. This review will summarize the roles of MYC in B cell development and oncogenesis, as well as its significance for current B cell lymphoma classification. We compared communication networks within transformed B cells in different lymphomas affected by overexpressed MYC and conducted a meta-analysis concerning the association of MYC with tumor prognosis in different patient populations.

BCL-W has a fundamental role in B cell survival and lymphomagenesis.

Compromised apoptotic signaling is a prerequisite for tumorigenesis. The design of effective therapies for cancer treatment depends on a comprehensive understanding of the mechanisms that govern cell survival. The antiapoptotic proteins of the BCL-2 family are key regulators of cell survival and are frequently overexpressed in malignancies, leading to increased cancer cell survival. Unlike BCL-2 and BCL-XL, the closest antiapoptotic relative BCL-W is required for spermatogenesis, but was considered dispensable for all other cell types. Here, however, we have exposed a critical role for BCL-W in B cell survival and lymphomagenesis. Loss of Bcl-w conferred sensitivity to growth factor deprivation-induced B cell apoptosis. Moreover, Bcl-w loss profoundly delayed MYC-mediated B cell lymphoma development due to increased MYC-induced B cell apoptosis. We also determined that MYC regulates BCL-W expression through its transcriptional regulation of specific miR. BCL-W expression was highly selected for in patient samples of Burkitt lymphoma (BL), with 88.5% expressing BCL-W. BCL-W knockdown in BL cell lines induced apoptosis, and its overexpression conferred resistance to BCL-2 family-targeting BH3 mimetics. Additionally, BCL-W was overexpressed in diffuse large B cell lymphoma and correlated with decreased patient survival. Collectively, our results reveal that BCL-W profoundly contributes to B cell lymphoma, and its expression could serve as a biomarker for diagnosis and aid in the development of better targeted therapies.

Development of Mouse Model Systems of Germinal Center Lymphomas.

Germinal centers (GC) are the predominant origin of human B cell lymphomagenesis. Transgenic mice in which gene expression is altered specifically in GC B cells have broadened our knowledge about the mechanisms of malignant transformation. However, extensive resources are needed due to the genetic complexity of these mouse models. Thus, bone marrow (BM)-derived chimerism is an attractive approach to study GC B cell derived lymphomagenesis, as it allows for an efficient allocation of resources and reduces the number of animals used.

AICDA Introduces Epigenetic Plasticity in Germinal Center-Derived Lymphomas and Accelerates Lymphomagenesis

Diffuse large B-cell lymphomas (DLBCLs) are aggressive tumors derived from germinal center B cells (GC) B cells. Previous work from our group established that inferior outcome in DLBCL is associated with higher degrees of intra-tumor and inter-tumor cytosine methylation heterogeneity, although the molecules driving this epigenetic perturbation are unknown. We investigated the contribution of activation-induced cytidine deaminase (AICDA) to cytosine methylation heterogeneity in DLBCLs. AICDA is highly expressed in GC B cells where it drives somatic hypermutation (SHM) of immunoglobulin (Ig) and non-Ig genes. AICDA is also expressed in DLBCLs and high level of AICDA in CHOP-treated DLBCL patients is associated with unfavorable prognosis. In addition to mutagenesis, AICDA can cause DNA demethylation through the removal of modified cytosine residues. Indeed, we previously reported that GC B cells manifest a marked AICDA-mediated DNA hypomethylation signature as well as a high degree of epigenetic heterogeneity as compared to other B-cell subsets. Thus, we hypothesized that AICDA could contribute to the progression of GC-derived lymphomas by facilitating epigenetic plasticity through the redistribution of cytosine methylation. We overexpressed AICDA in VavP-Bcl2 transgenic mice -which develop B cell lymphomas of GC origin- using an AICDA-expressing retroviral vector (VavP-Bcl2+Aicda) vs empty vector (VavP-Bcl2) as control and transplanted them into lethally irradiated recipient mice. We observed more aggressive lymphomas based on greater disruption of the splenic architecture and higher degree of B cell infiltration in organs such as lung, liver and kidney in VavP-Bcl2+Aicda mice (n=7) compared to VavP-Bcl2 mice (n=6). Notably, the overexpression of AICDA reduced significantly the lifespan of the mice (Log-rank test p=0.0289) with a median survival of 214 days for VavP-Bcl2+Aicda mice (n=10) and 293 days for VavP-Bcl2 animals (n=9). Neoplastic B cells from VavP-Bcl2+Aicda and VavP-Bcl2 micedisplayed similar SHM rate in Ig genes (JH4 and Sm) and AICDA off-targets (Bcl6, Cd83 and Pax5), suggesting that the more aggressive phenotype is not likely due to increased mutagenesis. We then profiled the DNA methylation landscape of these lymphomas using enhanced reduced representation bisulfite sequencing. We analyzed the DNA methylation level and heterogeneity of all represented CpGs by calculating the mean methylation difference and interquartile range (IQR) between VavP-Bcl2+Aicda and VavP-Bcl2 tumors. A principal component analysis of all CpGs, each one represented by its mean methylation and IQR differences across replicates, revealed methylation loss and increased intertumor heterogeneity within VavP-Bcl2+Aicda methylomes compared to VavP-Bcl2. AICDA-perturbed CpGs (n=49,750) in particular showed significant intratumor heterogeneity (p<1e-300) in AICDA overexpressing tumors. These altered CpGs were depleted in promoters and enriched in introns and intergenic regions. We observed a remarkably similar pattern of hypomethylation and increased methylation heterogeneity in Aicda+/+ GC B cells compared to Aicda-/- GC B cells (AICDA-perturbed CpGs n=64,323) and also in primary DLBCLs with high AID compared to low AID expression (AICDA-perturbed CpGs n=37,557), suggesting a conserved epigenetic function of AICDA in GC B cells and human and mouse GC-derived lymphomas. We performed RNA sequencing on BCL2-driven lymphomas and found that expressed genes containing AICDA-perturbed CpGs displayed significantly lower expression in VavP-Bcl2+Aicda compared to VavP-Bcl2 tumors (gene set enrichment analysis (GSEA) NES=-1.68, FDR<0.001). Pathway analysis revealed that these AICDA-epigenetic target genes were associated with MAPK- and KMT2D-signaling pathways, which are related to B cell activation and lymphomagenesis. Similarly to mouse tumors, genes enriched for AICDA-perturbed CpG in human lymphomas exhibited lower expression in AID high DLBCLs compared to AICDA low DLBCLs (GSEA NES=-1.65, FDR<0.001) and were associated with MAPK and KMT2D pathways. These results demonstrate that AICDA acts as a methylome modifier in GC-derived lymphomas, introducing epigenetic plasticity and leading to gene expression changes, thus contributing to clonal evolution and higher adaptability to an evolving environment. DisclosuresMelnick:Janssen: Research Funding.

MYC selects against reduced BCL2A1/A1 protein expression during B cell lymphomagenesis

Rearrangements of MYC or ABL proto-oncogenes lead to deregulated expression of key-regulators of cell cycle and cell survival, thereby constituting important drivers of blood cancer. Members of the BCL-2 family of apoptosis regulators contribute to oncogenic transformation downstream of these oncogenes, but the role of anti-apoptotic BCL2A1/A1 in transformation and drug resistance caused by deregulation of these oncogenes remains enigmatic. Here we analyzed the role of A1 in MYC as well as ABL kinase-driven blood cancer in mice, employing in vivo RNAi. We report that overexpression of either oncogene leads to a significant increase in A1 protein levels in otherwise A1-negative B cell progenitors, indicating a key role downstream of these oncogenes to secure survival during transformation. Knockdown of A1 by RNAi, however, did not impact on tumor latency in v-Abl-driven pre-B-ALL. In contrast, A1 knockdown in premalignant Eμ-MYC mice caused a significant reduction of transgenic pre-B cells without impacting on tumor latency as the emerging lymphomas escaped silencing of A1 expression. These findings identify A1 as a MYC target that can be induced prematurely during B cell development to aid expansion of otherwise cell-death-prone MYC transgenic pre-B cells. Hence, A1 should be considered as a putative drug target in MYC-driven blood cancer.

Gata3 restrains B cell proliferation and cooperates with p18INK4c to repress B cell lymphomagenesis.

GATA3, a lineage specifier, controls lymphoid cell differentiation and its function in T cell commitment and development has been extensively studied. GATA3 promotes T cell specification by repressing B cell potential in pro T cells and decreased GATA3 expression is essential for early B cell commitment. Inherited genetic variation in GATA3 has been associated with lymphoma susceptibility. However, it remains elusive how the loss of function of GATA3 promotes B cell development and induces B cell lymphomas. In this study, we found that haploid loss of Gata3 by heterozygous germline deletion increased B cell populations in the bone marrow (BM) and spleen, and decreased CD4 T cell populations in the thymus, confirming that Gata3 promotes T and suppresses B cell development. We discovered that haploid loss of Gata3 reduced thymocyte proliferation with induction of p18Ink4c (p18), an inhibitor of CDK4 and CDK6, but enhanced B cell proliferation in the BM and spleen independent of p18. Loss of p18 partially restored Gata3 deficient thymocyte proliferation, but further stimulated Gata3 deficient B cell proliferation in the BM and spleen. Furthermore, we discovered that haploid loss of Gata3 in p18 deficient mice led to the development of B cell lymphomas that were capable of rapidly regenerating tumors when transplanted into immunocompromised mice. These results indicate that Gata3 deficiency promotes B cell differentiation and proliferation, and cooperates with p18 loss to induce B cell lymphomas. This study, for the first time, reveals that Gata3 is a tumor suppressor specifically in B cell lymphomagenesis.

MS-analysis of SILAC-labeled MYC-driven B lymphoma cells overexpressing miR-17-19b.

Micro RNAs (miRNAs) are small non-coding RNAs, which dampen gene expression by repressing translation and/or inducing degradation of target-mRNAs. Although the role of miR-17-19b (a truncated version of miR-17-92 cluster) is well documented in MYC-driven B cell lymphomagenesis, little is known about the function of the cluster in the maintenance of full-blown lymphomas. We employed SILAC-based quantitative proteomics to identify miR-17-19b targets upon a mild overexpression of the cluster in B cell lymphomas, established from λ-MYC transgenic mice. The proteomics data described in detail in this study, whose follow up analysis with MaxQuant algorithm is part of the recent publication (Mihailovich et al., 2015) [1], are deposited to the ProteomeXchange Consortium via the PRIDE partner repository, with the accession code PRIDE: PXD002810.

Tetraspanin CD37 protects against the development of B cell lymphoma.

Worldwide, B cell non-Hodgkin lymphoma is the most common hematological malignancy and represents a substantial clinical problem. The molecular events that lead to B cell lymphoma are only partially defined. Here, we have provided evidence that deficiency of tetraspanin superfamily member CD37, which is important for B cell function, induces the development of B cell lymphoma. Mice lacking CD37 developed germinal center-derived B cell lymphoma in lymph nodes and spleens with a higher incidence than Bcl2 transgenic mice. We discovered that CD37 interacts with suppressor of cytokine signaling 3 (SOCS3); therefore, absence of CD37 drives tumor development through constitutive activation of the IL-6 signaling pathway. Moreover, animals deficient for both Cd37 and Il6 were fully protected against lymphoma development, confirming the involvement of the IL-6 pathway in driving tumorigenesis. Loss of CD37 on neoplastic cells in patients with diffuse large B cell lymphoma (DLBCL) directly correlated with activation of the IL-6 signaling pathway and with worse progression-free and overall survival. Together, this study identifies CD37 as a tumor suppressor that directly protects against B cell lymphomagenesis and provides a strong rationale for blocking the IL-6 pathway in patients with CD37- B cell malignancies as a possible therapeutic intervention.

CXCL13 and CCL11 Serum Levels and Lymphoma and Disease Activity in Primary Sjögren's Syndrome.

Non-Hodgkin's lymphoma (NHL) is a severe complication of primary Sjögren's syndrome (SS). Ectopic germinal centers (GCs) in the salivary glands are predictors of the occurrence of NHL. Given the association between CCL11 and CXCL13 and ectopic GCs, we assessed the link between these chemokines and NHL, as well as the association between these chemokines and disease activity, in patients with primary SS. Serum levels of CCL11 and CXCL13 were evaluated by multiplex assay in 385 patients included in the Assessment of Systemic Signs and Evolution of Sjögren's Syndrome (ASSESS) cohort. The association between chemokine levels, B cell biomarkers, and patient subsets was assessed using Spearman's test for continuous data and the nonparametric Mann-Whitney U test for categorical data. Multivariate analyses were performed to identify parameters associated with lymphoma and disease activity. Seventeen patients had a history of lymphoma, and 5 of them had developed NHL during followup. The median serum levels of CCL11 and CXCL13 in the total cohort were 106.48 pg/ml (interquartile range 69.33-149.85) and 108.31 pg/ml (interquartile range 58.88-200.13), respectively. Patients with lymphoma had higher levels of CXCL13 than did patients without lymphoma (P = 0.006) and a trend toward a higher level of CCL11 (P = 0.056). Low C4 and high BAFF levels were associated with NHL on multivariate analysis (P = 0.01 and P = 0.0002, respectively). CCL11 and CXCL13 levels correlated positively with the rheumatoid factor titer, the κ-to-λ free light chain ratio, and the β2 -microglubulin level. CXCL13 was the only parameter associated with disease activity on multivariate analysis. These findings demonstrate a link between CXCL13 and CCL11 and disease activity and lymphoma. This highlights the continuum between chronic B cell activation, disease activity, and lymphomagenesis in patients with primary SS.