Development Of B-cell Lymphoma Research Articles

Pre-diagnostic blood immune markers, incidence and progression of B-cell lymphoma and multiple myeloma: Univariate and functionally informed multivariate analyses.

Recent prospective studies have shown that dysregulation of the immune system may precede the development of B‐cell lymphomas (BCL) in immunocompetent individuals. However, to date, the studies were restricted to a few immune markers, which were considered separately. Using a nested case–control study within two European prospective cohorts, we measured plasma levels of 28 immune markers in samples collected a median of 6 years before diagnosis (range 2.01–15.97) in 268 incident cases of BCL (including multiple myeloma [MM]) and matched controls. Linear mixed models and partial least square analyses were used to analyze the association between levels of immune marker and the incidence of BCL and its main histological subtypes and to investigate potential biomarkers predictive of the time to diagnosis. Linear mixed model analyses identified associations linking lower levels of fibroblast growth factor‐2 (FGF‐2 p = 7.2 × 10−4) and transforming growth factor alpha (TGF‐α, p = 6.5 × 10−5) and BCL incidence. Analyses stratified by histological subtypes identified inverse associations for MM subtype including FGF‐2 (p = 7.8 × 10−7), TGF‐α (p = 4.08 × 10−5), fractalkine (p = 1.12 × 10−3), monocyte chemotactic protein‐3 (p = 1.36 × 10−4), macrophage inflammatory protein 1‐alpha (p = 4.6 × 10−4) and vascular endothelial growth factor (p = 4.23 × 10−5). Our results also provided marginal support for already reported associations between chemokines and diffuse large BCL (DLBCL) and cytokines and chronic lymphocytic leukemia (CLL). Case‐only analyses showed that Granulocyte‐macrophage colony stimulating factor levels were consistently higher closer to diagnosis, which provides further evidence of its role in tumor progression. In conclusion, our study suggests a role of growth‐factors in the incidence of MM and of chemokine and cytokine regulation in DLBCL and CLL.

Coexistence of chronic myeloid leukemia and diffuse large B-cell lymphoma with antecedent chronic lymphocytic leukemia: a case report and review of the literature

BackgroundChronic lymphocytic leukemia and chronic myeloid leukemia are the most common types of adult leukemia. However, it is rare for the same patient to suffer from both. Richter’s transformation to diffuse large B-cell lymphoma is frequently observed in chronic lymphocytic leukemia. Purine analog therapy and the presence of trisomy 12, and CCND1 gene rearrangement have been linked to increased risk of Richter’s transformation. The coexistence of chronic myeloid leukemia and diffuse large B-cell lymphoma in the same patient is extremely rare, with only nine reported cases. Here, we describe the first reported case of concurrent chronic myeloid leukemia and diffuse large B-cell lymphoma in a background of chronic lymphocytic leukemia.Case presentationA 60-year-old Saudi man known to have diabetes, hypertension, and chronic active hepatitis B was diagnosed as having Rai stage II chronic lymphocytic leukemia, with trisomy 12 and rearrangement of the CCND1 gene in December 2012. He required no therapy until January 2016 when he developed significant anemia, thrombocytopenia, and constitutional symptoms. He received six cycles of fludarabine, cyclophosphamide, and rituximab, after which he achieved complete remission.One month later, he presented with progressive leukocytosis (mostly neutrophilia) and splenomegaly. Fluorescence in situ hybridization from bone marrow aspirate was positive for translocation (9;22) and reverse transcription polymerase chain reaction detected BCR-ABL fusion gene consistent with chronic myeloid leukemia. He had no morphologic or immunophenotypic evidence of chronic lymphocytic leukemia at the time. Imatinib, a first-line tyrosine kinase inhibitor, was started. Eight months later, a screening imaging revealed new liver lesions, which were confirmed to be diffuse large B-cell lymphoma.ConclusionsIn chronic lymphocytic leukemia, progressive leukocytosis and splenomegaly caused by emerging chronic myeloid leukemia can be easily overlooked. It is unlikely that chronic myeloid leukemia arose as a result of clonal evolution secondary to fludarabine treatment given the very short interval after receiving fludarabine. It is also unlikely that imatinib contributed to the development of diffuse large B-cell lymphoma; rather, diffuse large B-cell lymphoma arose as a result of Richter’s transformation. Fludarabine, trisomy 12, and CCND1 gene rearrangement might have increased the risk of Richter’s transformation in this patient.

FBXL10 contributes to the development of diffuse large B-cell lymphoma by epigenetically enhancing ERK1/2 signaling pathway

Epigenetic modifiers have emerged as critical factors governing the biology of different cancers. Herein we show that FBXL10 (also called KDM2B or JHDM1B), an important member of Polycomb repressive complexes, is overexpressed in human diffuse large B-cell lymphoma (DLBCL) tissues and the derived cell lines. Knocking down FBXL10 by specific short hairpin RNAs in DLBCL cells inhibits cell proliferation and induces apoptosis in vitro. Moreover, FBXL10 depletion in DLBCL cells abrogates tumor growth in mouse xenograft models. Through the analysis of RNA sequencing, we find that one of the key derepressed genes by depletion of FBXL10 is DUSP6, encoding a phosphatase for ERK1/2. Mechanistically FBXL10 maintains the silencing of DUSP6 expression via recruitment of Polycomb group proteins and deposition of repressive histone modifications at the DUSP6 promoter. Consistently, FBXL10 is required for ERK1/2 phosphorylation in DLBCL cells. Furthermore, we show that ERK1/2 activation and the proliferation rate of FBXL10-depleted cells can be rescued by downregulation of DUSP6 expression. These findings indicate that FBXL10 may be a promising therapeutic target in DLBCL and establish a link of epigenetic regulators to kinase signaling pathways.

Role of hypoxia in Diffuse Large B-cell Lymphoma: Metabolic repression and selective translation of HK2 facilitates development of DLBCL

Published molecular profiling studies in patients with lymphoma suggested the influence of hypoxia inducible factor-1 alpha (HIF1α) targets in prognosis of DLBCL. Yet, the role of hypoxia in hematological malignancies remains unclear. We observed that activation of HIF1α resulted in global translation repression during hypoxic stress in DLBCL. Protein translation efficiency as measured using 35S-labeled methionine incorporation revealed a ≥50% reduction in translation upon activation of HIF1α. Importantly, translation was not completely inhibited and expression of clinically correlated hypoxia targets such as GLUT1, HK2, and CYT-C was found to be refractory to translational repression under hypoxia in DLBCL cells. Notably, hypoxic induction of these genes was not observed in normal primary B-cells. Translational repression was coupled with a decrease in mitochondrial function. Screening of primary DLBCL patient samples revealed that expression of HK2, which encodes for the enzyme hexokinase 2, was significantly correlated with DLBCL phenotype. Genetic knockdown studies demonstrated that HK2 is required for promoting growth of DLBCL under hypoxic stress. Altogether, our findings provide strong support for the direct contribution of HK2 in B-cell lymphoma development and suggest that HK2 is a key metabolic driver of the DLBCL phenotype.

180 Sequential Development of Diffuse Large B-Cell Lymphoma in a Patient With Peripheral T-Cell Lymphoma: Challenges and Risk Factors

We report an extremely rare case of diffuse large B-cell lymphoma sequentially arising in a 61-year-old male patient with CD30-positive peripheral T-cell lymphoma, 9 months after his initial presentation. The patient presented to hospital with extensive pelvic retroperitoneal and inguinal lymphadenopathy. Histologic examination of the left inguinal lymph node excisional biopsy was interpreted as peripheral T-cell lymphoma, not otherwise specified, CD30 positive, Epstein-Barr virus (EBV) positive. The tumor cells were positive for CD3, CD4, CD5, CD30, CD20 (subset), and BCL6 (small subset), with partial loss of CD2 and near complete loss of CD7 and BCL2 by IHC. EBV encoding region in situ hybridization was positive in 10%-20% of lymphoma cells. Flow cytometry showed an abnormal T cell population with polyclonal B cell population. PET CT was positive for bone involvement. Cytogenetics showed a normal male karyotype, and no evidence of clonal T- or B-cell gene rearrangement was detected by PCR. The patient received six cycles of CHOEP (cyclophosphamide, doxorubicin, vincristine, etoposide, and prednisone) chemotherapy. Post chemotherapy, PET-CT revealed good response but with residual disease warranting a second biopsy from the left retroperitoneal lymph node, this time showing diffuse large B-cell lymphoma, germinal center type, CD30 negative, EBV negative. The tumor cells were positive for CD10, CD20, and PAX5, and negative for CD2, CD3, CD30, ALK1, and Cyclin D1. They were kappa restricted by flow cytometry. BCL2 gene rearrangement was detected by FISH. He was started on R-GDP (rituximab, gemcitabine, dexamethasone, cisplatin) and completed six cycles but never achieved complete remission. Immunohistochemistry, flow cytometry, molecular studies, and cytogenetics performed during the course of disease confirmed two clonally distinct T- and B-cell lymphomas. We discuss here this unique and challenging case along with the possible etiology and risk factors involved in its development.

Abstract 02: BCL-W significantly contributes to B-cell lymphoma survival and development

Abstract The ability of cells to undergo apoptosis in response to cellular stresses functions as a safeguard against tumorigenesis. Consequently, compromised apoptotic signaling must precede tumor development. To design effective therapies for cancer treatment, the underlying mechanisms that govern cell survival need to be understood. The anti-apoptotic proteins of the Bcl-2 family are key regulators of cell survival and are frequently overexpressed in malignancies, leading to increased cancer cell survival. Bcl-w, the closest anti-apoptotic relative to Bcl-2 and Bcl-xL, is essential for spermatogenesis, but was considered dispensable for other cell types. However, we recently uncovered a significant role for Bcl-w in B-cell survival and lymphomagenesis. Specifically, loss of Bcl-w conferred sensitivity to growth factor deprivation-induced B-cell apoptosis. In addition, apoptosis triggered by hyperproliferative signals from the Myc oncogene was accentuated by loss of Bcl-w. Consequences of elevated apoptosis were observed in vivo, as Bcl-w loss profoundly delayed Myc-mediated B-cell lymphoma development. We also determined Myc regulates Bcl-w expression through its transcriptional regulation of the miR-15 family. Importantly, evaluation of Burkitt lymphoma (BL) patient samples revealed BCL-W overexpression was highly selected for, and that targeting BCL-W using multiple methods induced apoptosis of BL cells. In addition, overexpression of BCL-W in BL cell lines conferred resistance to the BH3-mimetics ABT-737 and ABT-263 that inhibit anti-apoptotic BCL-2 family members. Furthermore, BCL-W was overexpressed in diffuse large B-cell lymphoma (DLBCL) and correlated with decreased patient survival. Analysis of multiple data sets of other aggressive and indolent B-cell lymphomas showed that BCL-W was frequently overexpressed in them as well. Collectively, our data indicate that BCL-W is an understudied contributor to B-cell lymphoma development and survival that appears to be as important as BCL-2 to B-cell lymphomas. In addition, BCL-W is a predictive biomarker that could improve prognostication and the development of more effective treatment strategies for B-cell lymphomas. Citation Format: Clare M. Adams, Ramkrishna Mitra, Jerald Z. Gong, Annette S. Kim, John K. Choi, Christine M. Eischen. BCL-W significantly contributes to B-cell lymphoma survival and development [abstract]. In: Proceedings of the Second AACR Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; May 6-9, 2017; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(24_Suppl):Abstract nr 02.

Outcomes among North American patients with diffuse large B-cell lymphoma are independent of tumor Epstein-Barr virus positivity or immunosuppression.

The prevalence, presenting clinical and pathological characteristics, and outcomes for patients with diffuse large B-cell lymphoma that is Epstein-Barr virus positive remain uncertain as does the impact of congenital or iatrogenic immunosuppression. Patients with newly diagnosed diffuse large B-cell lymphoma with available tissue arrays were identified from the University of Iowa/Mayo Clinic Molecular Epidemiology Resource. Patients infected with human immunodeficiency virus or who had undergone a prior organ transplant were excluded. Epstein-Barr virus-associated ribonucleic acid testing was performed on all tissue arrays. A history of significant congenital or iatrogenic immunosuppression was determined for all patients. At enrollment, 16 of the 362 (4.4%) biopsies were positive for Epstein-Barr virus. Thirty-nine (10.8%) patients had a significant history of immunosuppression. Patients with Epstein-Barr-positive diffuse large B-cell lymphoma had no unique clinical characteristics but on pathology exhibited a higher frequency of CD30 positivity (25.0% versus 8.1%, respectively; P<0.01), and non-germinal-center subtype (62.5% versus 34.1%, respectively; P<0.01). No baseline clinical characteristics were associated with a history of immunosuppression. With a median follow up of 59 months, and after adjustment for International Prognostic Index, there was no association of Epstein-Barr virus positivity or immunosuppression with event-free survival at 24 months (odds ratio=0.49; 95% confidence interval: 0.13–1.84 and odds ratio=0.81; 95% confidence interval: 0.37–1.77) or overall survival (hazard ratio=0.86; 95% confidence interval: 0.38–1.97 and hazard ratio=1.00; 95% confidence interval: 0.57–1.74). In contrast to non-Western populations, our North American population had a low prevalence of Epstein-Barr virus-positive diffuse large B-cell lymphoma that did not convey an adverse prognosis. A history of immunosuppression, while known to be a risk factor for the development of diffuse large B-cell lymphoma, did not affect subsequent prognosis.

SHIP1, but not an AML-derived SHIP1 mutant, suppresses myeloid leukemia growth in a xenotransplantation mouse model.

Constitutive activation of the PI3K/AKT signaling pathway is found in ~50-70% of AML patients. The SH2-containing inositol 5-phosphatase 1 (SHIP1) is a negative regulator of PI3K/AKT signaling in hematopoietic cells. SHIP1 knockout mice develop a myeloproliferative syndrome and concomitant deletion of SHIP1 and the tumor suppressor PTEN leads to the development of lethal B-cell lymphomas. In the study presented here, we investigated the role of SHIP1 as a tumor suppressor in myeloid leukemia cells in an in vivo xenograft transplantation model. NSG Mice transplanted with UKE-1 cells derived from a secondary AML showed a significantly extended lifespan after lentiviral-mediated overexpression of SHIP1 in comparison to the vector control cohort. In contrast, the AML-derived SHIP1Y643H mutant, which has a strongly reduced enzymatic activity showed a significant reversion of the SHIP1-induced prolongation of the survival time. In addition, the analysis of 290 AML patients revealed a correlation between expression of SHIP1 and overall survival of the AML patients. These results indicate that SHIP1 can act as a tumor suppressor in acute myeloid leukemia cells and that higher SHIP1 expression is associated with prolonged overall survival in AML patients. SHIP1 may be an interesting candidate for gene therapy.

IL4 and IL21 cooperate to induce the high Bcl6 protein level required for germinal center formation.

Bcl6 (B-cell lymphoma 6) is a transcriptional repressor and critical mediator of the germinal center reaction during a T-cell-dependent antibody response, where it enables somatic hypermutation of immunoglobulin genes and inhibits terminal differentiation via repression of Blimp1. It can also contribute to the development of diffuse large B-cell lymphoma when expressed inappropriately. Bcl6 regulation is mediated both at the transcriptional and post-transcriptional levels, and in particular a strong signal through the B-cell receptor causes rapid proteasomal degradation of Bcl6. Despite the importance of Bcl6 in both immunity and cancer, little is known about how other extrinsic factors regulate Bcl6 in B cells. Here we show that Bcl6 is indeed highly unstable in B cells after a B-cell receptor (BCR) signal, but that the T-cell-derived cytokines interleukin 4 (IL4) and IL21 counteract BCR-mediated degradation, preserving Bcl6 protein levels. Stat6, downstream of IL4, can induce Bcl6 transcription directly. In vivo, B-cell intrinsic loss of IL4 or IL21 signaling reduces the magnitude or duration of the GC response, respectively, while their combined loss almost completely eliminates the GC response. This work provides key insights into the effect mediated by T-follicular helper cytokines on Bcl6 regulation.

An Unexpected Cause of Iron Deficiency Anemia: Synchronous Development and Diagnosis of a Rare Gastric Collision Tumor

A 75-year-old man with diabetes and hypertension presented with chest pain and was found to have a ST-elevation myocardial infarction. He was taken to the cardiac catheterization lab for coronary angiography. Following the procedure, he described a several week history of post-prandial epigastric discomfort, for which he was taking naproxen. He denied symptoms of gastrointestinal bleeding. Laboratory data showed iron deficiency anemia (ferritin: 15 ng/mL, hemoglobin: 11.3 g/dL). Esophagogastroduodenoscopy (EGD) demonstrated an irregular, linear, non-bleeding 3 cm ulcer in the stomach body extending into the antrum with a hard consistency (Figure 1), as well as an irregular, cratered, non-bleeding 3 cm ulcer in the gastric cardia with heaped-up margins (Figure 2). Biopsies of the ulcer in the body revealed an invasive moderately differentiated gastric adenocarcinoma (Figure 3, top left and right). Biopsies of the ulcer in the cardia revealed diffuse large B-cell lymphoma (Figure 3, bottom left and right). Histopathology was negative for Helicobacter pylori (H. pylori). The patient underwent subsequent coronary artery bypass grafting and is following with an oncologist for further management. The EGD and histopathologic findings demonstrated synchronous development of a rarely reported collision tumor: gastric adenocarcinoma and diffuse large B-cell lymphoma, each having a distinct endoscopic appearance. Synchronous tumors in the stomach are uncommon. The majority of synchronous malignancies are gastric adenocarcinomas and mucosa associated lymphoid tissue (MALT) lymphomas.1 There have been less than 10 known published reports of the synchronous development of gastric adenocarcinoma and diffuse large B-cell lymphoma.1 This case is also unusual, as H. pylori was not detected on histology. This vignette is significant both for the rarity of the final diagnoses, as well as the reminder that the practicing gastroenterologist provide a thorough endoscopic examination and perform appropriate mucosal sampling, even in the presence of an already identified abnormality. These malignancies would likely have been diagnosed at a later stage had the patient not presented with acute coronary syndrome and undergone endoscopic evaluation for iron deficiency anemia.Figure: Gastric Body Adenocarcinoma.Figure: Gastric Cardia Diffuse Large B-Cell Lymphoma.Figure: Clockwise, from Top Left: Gastric Body Adenocarcinoma (H&E stain, 40x), Invasive Neoplastic Glands (H&E stain, 200x), Gastric Cardia Large Cell Lymphoma (H&E stain, 400x), Diffuse B-Cell Infiltrates Entrapping Benign Gastric Glands (H&E stain, 40x).

Early loss of Crebbp confers malignant stem cell properties on lymphoid progenitors.

Loss-of-function mutations of cyclic-AMP response element binding protein, binding protein (CREBBP) are prevalent in lymphoid malignancies. However, the tumour suppressor functions of CREBBP remain unclear. We demonstrate that loss of Crebbp in murine haematopoietic stem and progenitor cells (HSPC) leads to increased development of B-cell lymphomas. This is preceded by accumulation of hyperproliferative lymphoid progenitors with a defective DNA damage response (DDR) due to a failure to acetylate p53. We identify a pre-malignant lymphoma stem cell population with decreased H3K27ac, which undergoes transcriptional and genetic evolution due to the altered DDR, resulting in lymphomagenesis. Importantly, when Crebbp is lost later in lymphopoiesis, cellular abnormalities are lost and tumour generation attenuated. We also document that CREBBP mutations may occur in HSPC from patients with CREBBP-mutated lymphoma. These data suggest that earlier loss of Crebbp is advantageous for lymphoid-transformation and inform the cellular origins and subsequent evolution of lymphoid malignancies.

MiRNAs in B-cell lymphoma: Molecular mechanisms and biomarker potential

MicroRNAs (miRNAs) are small non-coding RNAs that regulate many human genes including those involved in normal B-cell development. When these miRNAs are aberrantly expressed in B-cells they play key pathogenetic roles in the development and maintenance of B-cell lymphomas and by association may serve as useful biomarkers. In this review, we provide an overview of the importance of miRNAs to B-cell lymphomagenesis, as well as considering their use as biomarkers, and their potential usefulness for the clinic.

RETRACTED: BEBT-908: A novel potent PI3K/HDAC inhibitor against diffuse large B-cell lymphoma

The PI3K pathway and epigenetic regulation have been shown to play a pivotal role in the development and progression of diffuse large B-cell lymphoma (DLBCL). In the clinic, existing PI3K and HDAC inhibitors show limited efficacy as single agents toward DLBCL. However, in preclinical studies, the synergistic effects of PI3K inhibitors and HDAC inhibitors on DLBCL have sparked the enthusiasm of researchers to target both PI3K and HDAC. We hypothesized that a novel dual PI3K/HDAC inhibitor, BEBT-908, would display improved pharmacologic effects on DLBCL. We analyzed the anti-DLBCL activity of BEBT-908 in a comprehensive manner, demonstrating its role in the suppression of invitro cell proliferation, blockade of PI3K and HDAC activities, inhibition of multiple signaling pathways, and promotion of apoptosis and cell cycle arrest. BEBT-908 showed potent PI3K/HDAC inhibition, with nanomolar IC50 values against DLBCL cell lines. Moreover, BEBT-908 inhibited multiple pathways, including JAK/STAT3, MAPK/ERK and NF-κB, and induced apoptosis and cell cycle arrest at G1 phase in these cells. Additionally, dual PI3K/HDAC inhibition was superior to the inhibition of PI3K or HDAC alone. The dual inhibitor BEBT-908 is a promising lead compound for developing novel targeted therapeutic agents against DLBCL.

Abstract 2655: Drug resistant B-cell tumors eliminated by novel therapeutic antibodies in vivo

Abstract B-cell malignancies have been successfully targeted in the clinic by therapies such as anti-CD20 antibody rituximab or Bruton’s tyrosine kinase inhibitor ibrutinib. However, leukemias and lymphomas remain incurable due to primary or acquired resistance, ultimately leaving patients without an effective treatment option. We sought to circumvent this drug resistance by pursuing an alternative target known as B cell activating factor receptor (BAFF-R). Despite past limited success, BAFF-R remains a prime target for B-cell lymphoma and leukemia therapeutic antibody development due to its key role in B-cell proliferation and development. We report the development of two novel monoclonal antibodies (mAbs) against human (h) BAFF-R. The mAbs were generated by immunizing mice with (h)BAFF-R-expressing mouse fibroblast cells presenting a natively folded, cell-surface immunogen. The two mAbs presented unique complementarity determining regions that specifically bound (h)BAFF-R-expressing mouse fibroblast cells but not the parental counterpart. Furthermore, the antibodies were specific to B-cell containing organs such as tonsil and spleen, by immunohistochemical staining and without detectable reactivity in heart, lung, brain, liver, and kidney tissues. To tailor the antibodies for clinical application, a human IgG1 Fc capable of eliciting an immune response was substituted, creating chimeric versions. We showed that both chimeric mAbs bound with high affinity to human B-cell lymphoma cell lines including JeKo-1 (mantle cell lymphoma; MCL), SU-DHL-6 (diffuse large B cell lymphoma; DLBCL), Raji (Burkitt lymphoma), and RL (follicular lymphoma). The chimeric antibodies also elicited antibody-dependent cell-mediated cytotoxicity (ADCC) with primary human natural killer (NK) cells in vitro against these tumor lines as well as primary lymphoma samples (n=5) from patients who progressed after rituximab exposure. Most notably, the antibodies demonstrated efficacy in two in vivo drug resistant lymphoma models we developed, a rituximab-resistant CD20 genomic knockout variant of JeKo-1 and the naturally ibrutinib-resistant Z-138. Using these lymphomas lines for xenogeneic tumor models in NOD scid gamma (NSG) mice, we found our antibodies significantly inhibited tumor growth, conferring long-term and tumor free survival on the mice. Our in vitro and in vivo results robustly demonstrate the high specificity and significant anti-tumor effects of our anti-BAFF-R antibodies against a broad variety of B-cell malignancies, especially against cases of rituximab and ibrutinib resistance. This successful development of novel anti-BAFF-R therapeutic antibodies warrants support for further translational development for clinical use in light of current resistance cases. Citation Format: Hong Qin, Guowei Wei, Ippei Sakamaki, Zhenyuan Dong, Wesley A. Cheng, Diane L. Smith, Feng Wen, Han Sun, Soung-chul Cha, Sattva S. Neelapu, Larry W. Kwak. Drug resistant B-cell tumors eliminated by novel therapeutic antibodies in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2655. doi:10.1158/1538-7445.AM2017-2655

A novel in vivo model for studying conditional dual loss of BLIMP-1 and p53 in B-cells, leading to tumor transformation.

The tumor suppressors B-lymphocyte-induced maturation protein-1 (BLIMP-1) and p53 play a crucial role in B-cell lymphomas, and their inactivation contributes to the pathogenesis of a wide spectrum of lymphoid malignancies, including diffuse large B-cell lymphomas (DLBCLs). Patients with activated B-cell-like (ABC) DLBCL may present with loss of BLIMP-1, c-Myc over-expression, decreased p53, and poor prognosis. Nevertheless, there is a lack of in vivo models recapitulating the biology of high-grade ABC DLBCL. We therefore aimed to develop an in vivo model aiming to recapitulate the phenotype observed in this cohort of patients. A Cre-Lox approach was used to achieve inactivation of both p53 and BLIMP-1 in murine B-cells. Contextual ablation of BLIMP-1 and p53 led to development of IgM-positive B-cell lymphoma with an aggressive phenotype, supported by c-Myc up-regulation, and accumulation of somatic mutations, as demonstrated by whole exome sequencing. Sensitivity of B-tumor cells to BTK inhibition was demonstrated. This model mirrors what reported in patients with ABC DLBLC, and therefore represents a novel model for studying the biology of ABC-DLBCL harboring the dual loss of BLIMP-1/p53 and c-Myc over-expression.

Clinicopathologic characteristics and outcomes of transformed diffuse large B-cell lymphoma in hepatitis C virus-infected patients: A case-control study of 84 patients.

7553 Background: Chronic hepatitis C virus (HCV) infection is associated with development of marginal zone lymphoma (MZL) and diffuse large B-cell lymphoma (DLBCL). Preliminary data showed particular characteristics of HCV-associated DLBCL; such as frequent transformation from indolent lymphomas and aggressive oncologic courses. We studied herein the clinicopathologic characteristics and outcomes of HCV-infected patients (pts) with transformed DLBCL. Methods: In this case control study, the medical records and pathology reports of HCV-infected (cases) and HCV-uninfected (controls) pts with transformed DLBCL seen at our institution (6/2004 - 5/2015) were reviewed. Included pts had a concomitant or a history of a low grade lymphoma. To determine their clinicopathologic characteristics, cases were compared to controls at a ratio of 1:3. To determine predictors of oncologic relapse, we compared pts with DLBCL relapse after first line chemotherapy to those who did not relapse in univariate and logistic regression analyses. Results: Twenty-one cases were compared to 63 controls. Compared to controls, cases were younger (median year age [interquartile range], 54 [49-62] v 62 [53-66], P = .01), had advanced Ann Arbor stages, 3-4 (95% v 76%, P = .05), and upper gastro-intestinal involvement (48% v 25%, P = .05). Immunophenotypically, cases had more CD10-negative B-cells (76% v 43%, P = .008) and CD5-positive B-cells (39% v 7%, P = .004) compared to controls, consistent with an activated B-cell phenotype. A comparison between pts who relapsed after first line chemotherapy (n = 42) and those who did not (n = 40) revealed that having CD5-positive B-cells was the only factor associated with DLBCL relapse in univariate (24% v 6%, P = .03) and multivariate analyses (OR = 10.7, P= .02). Conclusions: HCV-infected pts with transformed DLBCL are younger, present with advanced stages, are more commonly CD5-positive, and have an activated B-cell phenotype, suggesting more frequent transformation from MZL compared to HCV-uninfected pts. The higher frequency of CD5-positive B-cells might explain the higher DLBCL relapses previously described in HCV-infected pts.

TET2 mutations in B cells of patients affected by angioimmunoblastic T‐cell lymphoma

Angioimmunoblastic T-cell lymphomas (AITLs) frequently carry mutations in the TET2 and IDH2 genes. TET2 mutations represent early genetic lesions as they had already been detected in haematopoietic precursor cells of AITL patients. We show by analysis of whole-tissue sections and microdissected PD1+ cells that the frequency of TET2-mutated AITL is presumably even higher than reported (12/13 cases in our collection; 92%). In two-thirds of informative AITLs (6/9), a fraction of B cells was also TET2-mutated. Investigation of four AITLs by TET2 and IGHV gene sequencing of single microdissected B cells showed that between 10% and 60% of polyclonal B cells in AITL lymph nodes harboured the identical TET2 mutations of the respective T-cell lymphoma clone. Thus, TET2-mutated haematopoietic precursor cells in AITL patients not only give rise to the T-cell lymphoma but also generate a large population of mutated mature B cells. Future studies will show whether this is a reason why AITL patients frequently also develop B-cell lymphomas. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

The Bruton's tyrosine kinase inhibitor ibrutinib exerts immunomodulatory effects through regulation of tumor-infiltrating macrophages.

The Bruton's tyrosine kinase (Btk) inhibitor ibrutinib has demonstrated promising efficacy in a variety of hematologic malignancies. However, the precise mechanism of action of the drug remains to be fully elucidated. Tumor-infiltrating macrophages presented in the tumor microenvironment have been shown to promote development and progression of B-cell lymphomas through crosstalk mediated by secreted cytokines and chemokines. Because Btk has been implicated in Toll-like receptor (TLR) signaling pathways that regulate macrophage activation and production of proinflammatory cytokines, we investigated the immunomodulatory effects of Btk inhibitor on macrophages. Our results demonstrate that Btk inhibition efficiently suppresses production of CXCL12, CXCL13, CCL19, and VEGF by macrophages. Furthermore, attenuated secretion of homeostatic chemokines from Btk inhibitor-treated macrophages significantly compromise adhesion, invasion, and migration of lymphoid malignant cells and even those not driven by Btk expression. The supernatants from Btk inhibitor-treated macrophages also impair the ability of endothelial cells to undergo angiogenic tube formation. Mechanistic analysis revealed that Btk inhibitors treatment downregulates secretion of homeostatic chemokines and cytokines through inactivation of Btk signaling and the downstream transcription factors, NF-κB, STAT3, and AP-1. Taken together, these results suggest that the encouraging therapeutic efficacy of Btk inhibitor may be due to both direct cytotoxic effects on malignant B cells and immunomodulatory effects on macrophages present in the tumor microenvironment. This novel mechanism of action suggests that, in addition to B-cell lymphomas, Btk inhibitor may also have therapeutic value in lymphatic malignancies and solid tumors lacking Btk expression.

Fbw7 regulates apoptosis in activated B-cell like diffuse large B-cell lymphoma by targeting Stat3 for ubiquitylation and degradation

BackgroundThe ubiquitin-ligase Fbw7 acts as a tumor suppressor, targeting lots of proto-oncogenes for proteolysis. However, the exact role of Fbw7 in diffuse large B-cell lymphoma (DLBCL) development remains unclear.MethodsWe evaluated Fbw7 expression in patient samples of DLBCL using immunohistochemical staining. The effect of Fbw7 overexpression on cell viability and apoptosis was investigated using activated B-cell (ABC) like DLBCL cell lines. The mechanism of Fbw7 activity in DLBCL was investigated using immunoprecipitation, ubiquitination, western blot and qualitative analyses.ResultsThe non-germinal center B-cell-like subtype of DLBCL showed reduced Fbw7 expression compared with the germinal center B-cell (GBC) subtype, and low Fbw7 expression was associated with a worse prognosis. Fbw7 overexpression caused decreased cell viability and increased apoptosis rates in the ABC-DLBCL cell lines SU-DHL-2 and OCI-LY-3. Importantly, Stat3 and phospho-Stat3Tyr705 stability were reduced following Fbw7 overexpression in ABC-DLBCL cell lines. In HEK293T and SU-DHL-2 cells, we demonstrated that Fbw7 interacts with Stat3 and pStat3Tyr705 to regulate their ubiquitylation and degradation. Downstream anti-apoptotic target genes of activated Stat3, including Myc, Survivin, Mcl-1, Pim-1, Bcl-2 and Bcl-xl showed decreased mRNA expression following exogenous Fbw7 overexpression. The negative relationship between Fbw7 and pStat3Tyr705 levels was also confirmed in DLBCL patient samples.ConclusionThe ubiquitin-ligase Fbw7 mediates apoptosis through targeting Stat3 for ubiquitylation and degradation in ABC-DLBCL. Thus, our study may offer a promising approach for ABC-DLBCL therapy through Stat3 inhibition.

Dynamics of Soluble B-Cell Activation Markers sCD27 and sCD30 and Risk of B-Cell Lymphoma: A Prospective Study Using Repeated Plasma Samples Donated up to 25 Years Pre-Diagnosis

Background: We studied changes in blood concentrations of the soluble B-cell activation markers sCD27 and sCD30 in relation to future lymphoma risk, by investigating repeated pre-diagnostic plasma samples from the same individuals. High levels of sCD27 and sCD30 have been associated with lymphoma risk in previous prospective studies based on a single pre-diagnostic blood sample per participant. These studies, however do not inform about the dynamics of the marker-disease association on an individual level hampering its interpretation and potential use in lymphoma diagnostics and disease monitoring.Methods: In the Northern Sweden Health and Disease Study cohort we identified 170 individuals who had donated two pre-diagnostic blood samples and subsequently developed B-cell lymphoma. Blood samples were donated 13.2±4.4 and 5.5±4.0 years (mean±SD) prior to diagnosis, respectively. Cancer-free controls from the same cohort were individually matched to cases on a 1:1 ratio on sex, age and blood draw dates. We investigated associations between lymphoma risk by subtypes and marker concentrations; as measured separately at baseline, at time of the repeated sample, as well as the association with rate of change (slope) while adjusting for the baseline marker concentration.Findings: We observed strong associations between B-cell lymphoma risk and sCD27 and sCD30 concentrations, both at baseline and at time of the repeated blood sample. Associations were evident in samples collected up to 25 years before diagnosis. Blood concentrations of sCD27 and sCD30 increased significantly closer to diagnosis among future B-cell lymphoma cases, while they remained temporally stable among controls. Modeling measures of baseline and slope simultaneously with multivariable conditional logistic regression to estimate odds ratios (ORs), revealed significant associations between both slope and baseline measures and lymphoma risk; sCD27 (ORBaseline=7.2, Ptrend=2.8 x 10-4; ORSlope=2.8, Ptrend=2.2 x 10-4), and sCD30 (ORBaseline=2.9, Ptrend=3.7 x 10-4; ORSlope=2.9, Ptrend=0.001) (ORs 4th vs. 1st quartile; P for linear trend based on median values of analyte quartiles used as a continuous variable). Subtype specific analyses showed that the association between lymphoma risk and slope was restricted to indolent subtypes and mainly driven by chronic lymphocytic leukemia; sCD27 (ORSlope=6.7, Ptrend=1.3 x 10-5), and sCD30 (ORSlope=5.9, Ptrend=4.8 x 10-5), while associations with baseline marker concentration were evident among different subtypes, including diffuse large B-cell lymphoma.Interpretation: B-cell lymphoma risk seems to be influenced by increased sCD27 and sCD30 blood concentrations more than two decades before diagnosis. Increased concentrations of these B-cell activation markers among different future lymphoma subtypes early in life may reflect a constitutional predisposition, suggesting a role of B-cell activation in lymphoma development at early stages across subtypes. The observed increase of sCD27 and sCD30 concentrations closer to diagnosis however, seems to be subtype-specific and restricted to indolent lymphoma cases. Therefore, these markers may reflect early progression of undiagnosed disease and could potentially be utilized to improve lymphoma diagnostics and disease monitoring in indolent lymphoma. DisclosuresNo relevant conflicts of interest to declare.