scholarly journals The proliferative history shapes the DNA methylome of B-cell tumors and predicts clinical outcome

2020 ◽  
Author(s):  
Martí Duran-Ferrer ◽  
Guillem Clot ◽  
Ferran Nadeu ◽  
Renée Beekman ◽  
Tycho Baumann ◽  
...  
Keyword(s):  
Dna Methylation ◽  
B Cells ◽  
B Cell ◽  
De Novo ◽  
Genetic Alterations ◽  
Cell Tumor ◽  
Patient Specific ◽  
Maturation Stage ◽  
Cell Of Origin ◽  
Dna Methylome

AbstractWe report a systematic analysis of the biological and clinical implications of DNA methylation variability in five categories of B-cell tumors derived from B cells spanning the entire maturation spectrum. We used 2056 primary samples including training and validation series and show that 88% of the human DNA methylome is dynamically modulated under normal and neoplastic conditions. B-cell tumors display both epigenetic imprints of their cellular origin and de novo, disease-specific epigenetic alterations that in part are related to differential transcription factor binding. These differential methylation patterns were used by a machine-learning approach to create a diagnostic algorithm that accurately classifies 14 B-cell tumor entities and subtypes with different clinical management. Beyond this, we identified extensive patient-specific epigenetic variability targeting constitutively silenced chromatin regions, a phenomenon we could relate to the proliferative history of normal and neoplastic B cells. We observed that, depending on the maturation stage of the tumor cell of origin, mitotic activity leaves different imprints into the DNA methylome. Subsequently, we constructed a novel DNA methylation-based mitotic clock called epiCMIT (epigenetically-determined Cumulative MIToses), whose lapse magnitude represents a strong independent prognostic variable within specific B-cell tumor subtypes and is associated with particular driver genetic alterations. Our findings reveal DNA methylation as a holistic tracker of B-cell tumor developmental history, with implications in the differential diagnosis and prediction of the outcome of the patients.

The Chemokine Receptor Profile As Distinctive Criterion Between Normal B-Cell Subsets and As Potential Discriminative Marker to Identify the Cell of Origin in Patients with Chronic Lymphocytic Leukemia and Richter Syndrome

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3890-3890
Author(s):  
Katharina Troppan ◽  
Kerstin Wenzl ◽  
Peter Neumeister ◽  
Christine Beham-Schmid ◽  
Martina Przekopowitz ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Chemokine Receptor ◽  
Chemokine Receptors ◽  
De Novo ◽  
Memory B Cells ◽  
Cell Of Origin ◽  
Richter Syndrome ◽  
Receptor Profile ◽  
B Cell Subsets

Abstract Chemokine receptors are G-protein-coupled cell surface receptors, which dissociate upon activation by their ligands and cause downstream signaling. Several studies have revealed the crucial contribution of chemokine receptors and their ligands in normal B-cell differentiation and development of hematopoietic malignancies. The Richter syndrome (RS) represents the clinico-pathologic transformation of chronic lymphocytic leukaemia (CLL) to an aggressive lymphoma, most commonly diffuse large B-cell lymphoma (DLBCL). Due to the lack of knowledge on the chemokine receptor, we aimed to investigate their expression profile in patients with CLL and Richter syndrome. Therefore, we investigated the mRNA expression levels of 18 known chemokine receptors (CCR1-CCR9, CXCR1-CXCR7, XCR1, CX3CR1) by using semi-quantitative real-time PCR on seven samples of paired (CLL and transformed DLBCL) RS samples, additionally four CLL samples -all of them subsequently transformed into DLBCL-, and eight transformed DLBCL samples originating from CLL. Additionally, 30 samples of de-novo DLBCL, including 10 germinal center B-cell (GCB) lymphomas, 12 non-germinal center B-cell lymphomas (non-GCB), and 8 unclassified DLBCL were included. Four samples of naïve B-cells (CD5 neg), CD5+ naïve B-cells and CD27+ memory B-cells (n=12) served as non-neoplastic controls. No differences in the chemokine receptor profile were detected between CD5+ and negative naïve B-cells. When comparing CD27+ memory B-cells to naïve B-cells a significant lower expression level was found for CCR7 (7-fold), CXCR4 (4-fold), and CXCR5 (1.5 fold). CCR7 (5-fold) and CXCR4 (5-fold) were also lower expressed in CD27+ memory B-cells compared to CD5+ naïve B-cells. Five out of 18 chemokine receptors were differentially expressed comparing the distinct normal B-cell subsets with RS samples. Comparing CLL samples and RS samples to CD5+ naïve B-cells, CXCR4 (12-fold for CLLs and 10-fold for RS samples) and CXCR5 (2-fold for CLLs and 2.4-fold for RS samples) were lower expressed, whereas CXCR3 (10-fold for CLLs and 8.5-fold for the transformed samples) was higher expressed and CCR5 de-novo expressed. Compared to naïve B-cells, the same chemokine receptors were deregulated: CXCR4 (10-fold for CLLs and 8.5-fold for the RS samples) and CXCR5 (2-fold for CLLs and 2.4-fold for the transformed samples) were lower expressed, CXCR3 (45-fold for CLLs and 30-fold for the transformed samples) was higher expressed and CCR5 was de-novo expressed. Comparing CLL samples and transformed RS samples to CD27+ memory B-cells, CCR5 (5.1-fold for CLLs and 4.3-fold for the RS samples) and CCR7 (8.7-fold for CLLs and 10-fold for the transformed samples) were higher expressed in both malignancies. Only one chemokine receptor was found to be differentially expressed in our seven paired RS samples: CCR6 showed a trend of a higher expression (1.4-fold) in CLL components. Considering RS and GCB DLBCL, CCR1, CCR5, and CXCR6 were found to be significantly down-regulated in RS (at least 4-fold), in contrast to CCR7 and CXCR4, which showed higher expression levels in RS (6-fold). CCR1 and CCR5 were lower expressed comparing RS and non-GCB DLBCL (25-fold and 8-fold), whereas CCR7 again, together with CXCR7, was higher expressed (3- fold and 6-fold respectively). Our data indicate a difference in the chemokine receptor profile within normal B-cell subsets. These differences are also reflected in the different expression profile of low and high aggressive component of CLL/RS compared to the distinct B cell subtypes. Hence, in future these multiple deregulated CC and CXC receptors might serve as a further hint in identifying the cell of origin of different B-cell malignancies. Disclosures No relevant conflicts of interest to declare.

scholarly journals Whole Genome DNA Methylation Analysis of Commpass Identifies Biomarkers of Multiple Myeloma Survival

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3174-3174
Author(s):  
Benjamin G Barwick ◽  
Daniel Auclair ◽  
Alex Blanski ◽  
Meghan Kirchhoff ◽  
Brianne Docter ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Multiple Myeloma ◽  
B Cells ◽  
B Cell ◽  
Prognostic Value ◽  
Bisulfite Sequencing ◽  
Plasma Cells ◽  
Whole Genome ◽  
Dna Methylome

Abstract Multiple myeloma is a malignancy of terminally differentiated, antibody secreting B cells known as plasma cells. Normal B cell differentiation and cell fate are coupled to epigenetic and transcriptional reprogramming, including a proliferation-dependent global loss of DNA methylation (Barwick et al., 2016, 2018). However, relatively little is known about the epigenetic changes that underlie myelomagenesis and how these may contribute to pathogenesis. To this end, we are analyzing the DNA methylome of myeloma specimens from the MMRF CoMMpass trial (NCT01454297), which has already characterized the mutational, structural, and transcriptional landscape of nearly 1,000 myelomas from newly diagnosed patients. CoMMpass specimens were obtained from a centralized biobank with approval from the CoMMpass Tissue Use Committee and Emory IRB. DNA isolated from CD138+ myeloma specimens was subjected to reduced representation bisulfite sequencing (RRBS) or whole genome bisulfite sequencing (WGBS). In total, DNA methylation was derived for over 24 million CpGs with an average of 18x coverage. WGBS data from normal B cells and plasma cells was obtained with permission from the BluePrint project (Agirre et al., 2015) via the European Genome Archive. DNA methylation levels were associated with PFS and OS using a cox proportional regression. We have determined the DNA methylome for 36 primary myeloma specimens and an additional 84 specimens are currently being sequenced. Relative to normal B cells that had an average DNA methylation level of 89.1%, plasma cells and myelomas exhibited a progressive demethylation with mean levels of 71.3% and 43.7%, respectively. While this is consistent with previous observations (Agirre et al., 2015; Salhia et al., 2010), WGBS revealed that myeloma in particular was characterized by large hypomethylated domains. These large hypomethylated domains encompassed genes that were devoid of gene expression whereas DNA methylation remained unchanged in the bodies of genes that were highly expressed. Although the majority of these hypomethylated domains were common across myelomas, there existed many regions where methylation levels varied between myelomas and these differences commonly corresponded with local gene expression differences. To understand if these specific patterns of DNA methylation were indicative of disease pathogenesis, DNA methylation levels were compared to PFS and OS. This identified 2,594 regions where the level of DNA methylation was prognostic of outcome (P≤0.001). Reduced DNA methylation corresponded with poor outcome at 88.5% (N=2,298) of these regions, which included loci proximal to cell cycle genes such as MYC, E2F3, CCND1, and CCNE1. Only 11.5% (N=296) of regions associated with outcome had higher levels of DNA methylation associated with poor prognosis. These regions tended to be proximal to genes involved in B cell receptor signaling, such as PLCG2 and VAV2. Although the expression of several of these genes was also prognostic of survival, the majority were not, indicating that the epigenetic state contains a unique prognostic value. These data indicate that myeloma undergoes profound epigenetic remodeling that is co-ordinate with changes in gene expression. Perhaps the most striking feature were megabase domains of hypomethylation. That DNA methylation was preferentially retained in the bodies of expressed genes suggests that a molecular mechanism and/or cellular selection occurs to maintain methylation at genes whose expression is required for myeloma cell survival. Despite the small number (N=36) of myeloma specimens analyzed thus far, the large number of regions associated with survival indicates the potential prognostic value of DNA methylation in myeloma. Furthermore, DNA methylation indicative of outcome only partially overlapped with the prognostic value of gene expression, indicating DNA methylation has independent value as a biomarker of outcome in myeloma. This may be due, in part, to the fact that DNA methylation is a very stable modification that not only reflects the current gene expression program, but is also indicative of the cell history and potential. Integrative genetic, epigenetic, and transcriptional analysis from WGBS of 120 CoMMpass myeloma specimens will be presented, including matched baseline and relapsed specimens from 25 patients. Disclosures Lonial: Amgen: Research Funding. Boise:Abbvie: Consultancy; AstraZeneca: Honoraria.

Two-Track Epigenetic Remodeling and Backtracking to Embryonic Stem Cell Bivalency in B-Cell Acute Lymphoblastic Leukemias

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3557-3557
Author(s):  
Seung-Tae Lee ◽  
Marcus O. Muench ◽  
Marina Fomin ◽  
Jianqiao Xiao ◽  
Mi Zhou ◽  
...  
Keyword(s):  
Dna Methylation ◽  
B Cells ◽  
B Cell ◽  
De Novo ◽  
Embryonic Stem ◽  
Cell Leukemia ◽  
Childhood All ◽  
B Cell Leukemia ◽  
Functional Consequences ◽  
De Novo Methylation

Abstract Purpose: We sought to define the drivers and functional consequences of DNA methylation changes in childhood pre-B cell leukemia. A comprehensive analysis of such DNA methylation changes will reveal aspects of the etiology and pathogenesis of leukemias, and suggest potential therapeutic modalities. Background: The epigenome is extensively altered in carcinogenesis, but the proximate causes, functional consequences, and overall patterning of DNA methylation changes in the pediatric leukemias is not well defined. Methods: We bisulfite sequenced at single base pair resolution two common pre-B cell leukemia case DNAs and a pre-B cell control (CD19+/CD34+ pre-B cells from normal marrow), and analyzed DNA methylation by high definition microarray in another 227 subjects. RNA expression was available for 82 samples, including the sequenced pre-B cells and leukemias. Results: Epigenetic alteration of B-ALLs occurred in two tracks: de novo methylation of small functional compartments and demethylation of large inter-compartmental backbones. The de novo methylation occurred preferentially at polycomb targets and binding sites for the transcriptional co-repressor CtBP2. DNA methylation of ETV6-AML1 leukemia was overall equivalent to the control, with high hyperdiploid methylation levels being 4.4% lower. Hierarchical clustering revealed four groups, with ETV6-AML1 and hyperdiploid leukemias enriched within two separate groups. DNA methylation deviations were exaggerated in lamina-associated domains, with differences corresponding to these methylation clusters and/or cytogenetic groups. DNA methylation changes were correlated with gene expression changes, and a key polycomb expression signature was reversed when DNA methylation marks were pharmacologically removed in leukemia cell lines. Discussion: While leukemia cells retain the majority of their developmentally determined DNA methylation patterns, key modifications with leukemogenesis revealed both broad patterns and functional targets with some dependence on known characteristics of pre-B cell leukemias. Our data suggested a pivotal role of polycomb and CTBP2 in de novo methylation, which may be traced back to bivalency status of embryonic stem cells. Driven by these potent epigenetic modulations, suppression of polycomb target genes was observed along with disruption of developmental fate and cell cycle and mismatch repair pathways and altered activities of key upstream regulators. An appreciation and validation of the observed changes will lead to new insights into etiology, pathogenesis, and treatments of childhood ALL. Disclosures No relevant conflicts of interest to declare.

scholarly journals Extensive RAG-Mediated Rearrangements and Mutations in BCR-ABL1 and BCR-ABL1-like Adult Acute Lymphoblastic Leukemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4067-4067
Author(s):  
Mathijs A. Sanders ◽  
Anikó Szabó ◽  
Carla Exalto ◽  
Remco Hoogenboezem ◽  
Annelieke Zeilemaker ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
B Cells ◽  
B Cell ◽  
De Novo ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Cell Development ◽  
B Cell Development ◽  
Genetic Events

Abstract BCR-ABL1 and BCR-ABL1-like acute lymphoblastic leukemia (ALL) are two major pre-B cell acute leukemia subtypes characterized by genetic alterations affecting lymphoid-specific transcription factors. Studies examining the chain of genetic events necessary to develop leukemia established that the BCR-ABL1 fusion gene and kinase-activating BCR-ABL1-like lesions are initiating events, however, insufficient for leukemia development. Secondary genetic events targeting B cell development genes are therefore an essential requirement for overt ALL. A recent study (Papaemmanuil et al, Nat. Genet., 2014) revealed that illegitimate RAG-mediated recombination is the predominant mutational mechanism establishing these secondary genetic events in ETV6-RUNX1 ALL. Of note, ETV6-RUNX1ALL is mainly restricted to pediatric cases and it remains unanswered whether this mutational process also plays a prominent role in adult ALL pathogenesis. We carried out a detailed genomic characterization to determine whether aberrant RAG activity is also a prominent mutational driver in certain adult B cell ALL (B-ALL) subtypes. Diagnostic material of 53 unselected B-ALL cases and matched remission specimens were characterized using DNA mapping arrays to discern copy number alterations (CNAs). We observed multiple BCR-ABL1/BCR-ABL1-like patients with abundant genetic lesions and selected 5 cases for targeted sequencing of CNA boundaries to determine whether these lesions were driven by RAG-mediated recombination. Whole genome sequencing (WGS) for a single BCR-ABL1-like patient was used to asses this mutational mechanism genome-wide. In total 64 structural variants (SVs) could be analyzed at base-pair level. De novo motif detection on breakpoint sequences revealed the prominence of the heptamer CACAGTG (E-value=5.68x10-91), a constituent of the recombination signal sequence (RSS), present in 121 out of 128 breakpoints (94.5%). RSS detection revealed that 58 out of 64 SVs (90.6%) had a cryptic RSS (cRSS) on one or both sides of the lesion. Incorporation of non-templated sequences was observed for 54 out of the 64 (84.4%) SVs. Superimposition of breakpoints on chromatin marks revealed a strong enrichment for active promoters and enhancers (p < 2.2x10-16). WGS data revealed cRSS motifs and incorporation of non-templated sequences for 23 out of 26 SVs (88.5%). Integrative analysis of all 6 cases confirmed 125 unique SV breakpoints strongly enriched for the active chromatin marks H3K4me3 and H3K27ac. STAT5 binding, a postulated regulator of V(D)J recombination, is similarly enriched at the breakpoints. Promiscuous binding of RAG1 and RAG2 was previously noted in human thymocytes and murine pre-B cells (Teng et al, Cell, 2015). Strikingly, the breakpoints are frequently bound by RAG2 in human thymocytes. In total 66 out of 125 breakpoints could be translated to the murine genome and revealed a strong enrichment of RAG1 and RAG2 binding at homologous positions in murine pre-B cells. Exhaustive mutation detection revealed complex somatic mutations within cRSS motifs, which are rare V(D)J recombination products introduced by erroneous cleavage and error-prone repair (open-and-shut joints). Strikingly, 4 out of 6 BCR-ABL1/BCR-ABL1-like cases had mutations in the BTLA promoter-situated cRSS, frequently in combination with a RAG-mediated deletion of the other allele (Figure 1). Genomic screening in 142 B-ALL patients confirmed 8 additional cases with BTLA promoter mutations, predominantly (6 out of 8) belonging to the BCR-ABL1/BCR-ABL1-like subgroups. We provide strong evidence that aberrant RAG activity plays a pivotal role in the development of BCR-ABL1/BCR-ABL1-like adult ALL. We demonstrate that breakpoints are strongly enriched for RAG binding implying a predisposition for illegitimate V(D)J recombination. Importantly, we report on a novel mutational mechanism introducing mutations in cRSS motifs through open-and-shut joints, frequently resulting in the biallelic inactivation of BTLA. Proliferation and V(D)J recombination during pre-B cell development is orchestrated by the interplay of IL7R and pre-BCR signalling. Strikingly, most kinase-activating lesions constitutively activate these signalling cascades and could enact, in concert with BTLA inactivation, constant proliferation, pro-survival and V(D)J recombination-initiating signals with disastrous consequences. Disclosures No relevant conflicts of interest to declare.

scholarly journals B cell depletion impairs vaccination-induced CD8+ T cell responses in a type I interferon-dependent manner

2021 ◽  
pp. annrheumdis-2021-220435
Author(s):  
Theresa Graalmann ◽  
Katharina Borst ◽  
Himanshu Manchanda ◽  
Lea Vaas ◽  
Matthias Bruhn ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cell Expansion ◽  
De Novo ◽  
T Cell Responses ◽  
Type I ◽  
Cell Responses ◽  
T Cell Expansion

ObjectivesThe monoclonal anti-CD20 antibody rituximab is frequently applied in the treatment of lymphoma as well as autoimmune diseases and confers efficient depletion of recirculating B cells. Correspondingly, B cell-depleted patients barely mount de novo antibody responses during infections or vaccinations. Therefore, efficient immune responses of B cell-depleted patients largely depend on protective T cell responses.MethodsCD8+ T cell expansion was studied in rituximab-treated rheumatoid arthritis (RA) patients and B cell-deficient mice on vaccination/infection with different vaccines/pathogens.ResultsRituximab-treated RA patients vaccinated with Influvac showed reduced expansion of influenza-specific CD8+ T cells when compared with healthy controls. Moreover, B cell-deficient JHT mice infected with mouse-adapted Influenza or modified vaccinia virus Ankara showed less vigorous expansion of virus-specific CD8+ T cells than wild type mice. Of note, JHT mice do not have an intrinsic impairment of CD8+ T cell expansion, since infection with vaccinia virus induced similar T cell expansion in JHT and wild type mice. Direct type I interferon receptor signalling of B cells was necessary to induce several chemokines in B cells and to support T cell help by enhancing the expression of MHC-I.ConclusionsDepending on the stimulus, B cells can modulate CD8+ T cell responses. Thus, B cell depletion causes a deficiency of de novo antibody responses and affects the efficacy of cellular response including cytotoxic T cells. The choice of the appropriate vaccine to vaccinate B cell-depleted patients has to be re-evaluated in order to efficiently induce protective CD8+ T cell responses.

Association between the proliferative rate of neoplastic B cells, their maturation stage, and underlying cytogenetic abnormalities in B-cell chronic lymphoproliferative disorders: analysis of a series of 432 patients

Blood ◽  
2008 ◽  
Vol 111 (10) ◽  
pp. 5130-5141 ◽  
Author(s):  
Sandra Quijano ◽  
Antonio López ◽  
Ana Rasillo ◽  
Susana Barrena ◽  
Maria Luz Sánchez ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoproliferative Disorders ◽  
Lymphocytic Leukemia ◽  
Maturation Stage ◽  
Cytogenetic Abnormalities ◽  
M Phase ◽  
The Impact

Abstract Limited knowledge exists about the impact of specific genetic abnormalities on the proliferation of neoplastic B cells from chronic lymphoproliferative disorders (B-CLPDs). Here we analyze the impact of cytogenetic abnormalities on the proliferation of neoplastic B cells in 432 B-CLPD patients, grouped according to diagnosis and site of sampling, versus their normal counterparts. Overall, proliferation of neoplastic B cells highly varied among the different B-CLPD subtypes, the greatest numbers of proliferating cells being identified in diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma (BL). Compared with normal B cells, neoplastic B-CLPD cells showed significantly increased S + G2/M-phase values in mantle cell lymphoma (MCL), B-chronic lymphocytic leukemia (B-CLL), BL, and some DLBCL cases. Conversely, decreased proliferation was observed in follicular lymphoma, lymphoplasmacytic lymphoma/Waldenström macroglobulinemia (LPL/WM), and some DLBCL patients; hairy cell leukemia, splenic marginal zone, and MALT-lymphoma patients showed S + G2/M phase values similar to normal mature B lymphocytes from LN. Interestingly, in B-CLL and MCL significantly higher percentages of S + G2/M cells were detected in BM versus PB and in LN versus BM and PB samples, respectively. In turn, presence of 14q32.3 gene rearrangements and DNA aneuploidy, was associated with a higher percentage of S + G2/M-phase cells among LPL/WM and B-CLL cases, respectively.

scholarly journals B cell activation and plasma cell differentiation are inhibited by de novo DNA methylation

2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Benjamin G. Barwick ◽  
Christopher D. Scharer ◽  
Ryan J. Martinez ◽  
Madeline J. Price ◽  
Alexander N. Wein ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Cell Differentiation ◽  
B Cell ◽  
Plasma Cell ◽  
Cell Activation ◽  
De Novo ◽  
B Cell Activation ◽  
Plasma Cell Differentiation

scholarly journals Epstein–Barr virus-encoded latent membrane protein 1 and B-cell growth transformation induces lipogenesis through fatty acid synthase.

2020 ◽  
Author(s):  
Michael Hulse ◽  
Sarah M Johnson ◽  
Sarah Boyle ◽  
Lisa Beatrice Caruso ◽  
Italo Tempera
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
B Cells ◽  
Cell Growth ◽  
B Cell ◽  
Fatty Acid Synthase ◽  
De Novo ◽  
Ectopic Expression ◽  
Barr Virus ◽  
Growth Transformation

Latent membrane protein 1 (LMP1) is the major transforming protein of Epstein-Barr virus (EBV) and is critical for EBV-induced B-cell transformation in vitro. Several B-cell malignancies are associated with latent LMP1-positive EBV infection, including Hodgkin’s and diffuse large B-cell lymphomas. We have previously reported that promotion of B cell proliferation by LMP1 coincided with an induction of aerobic glycolysis. To further examine LMP1-induced metabolic reprogramming in B cells, we ectopically expressed LMP1 in an EBV-negative Burkitt’s lymphoma (BL) cell line preceding a targeted metabolic analysis. This analysis revealed that the most significant LMP1-induced metabolic changes were to fatty acids. Significant changes to fatty acid levels were also found in primary B cells following EBV-mediated B-cell growth transformation. Ectopic expression of LMP1 and EBV-mediated B-cell growth transformation induced fatty acid synthase (FASN) and increased lipid droplet formation. FASN is a crucial lipogenic enzyme responsible for de novo biogenesis of fatty acids in transformed cells. Furthermore, inhibition of lipogenesis caused preferential killing of LMP1-expressing B cells and significantly hindered EBV immortalization of primary B-cells. Finally, our investigation also found that USP2a, a ubiquitin-specific protease, is significantly increased in LMP1-positive BL cells and mediates FASN stability. Our findings demonstrate that ectopic expression of LMP1 and EBV-mediated B-cell growth transformation leads to induction of FASN, fatty acids and lipid droplet formation, possibly pointing to a reliance on lipogenesis. Therefore, the use of lipogenesis inhibitors could potentially be used in the treatment of LMP1+ EBV associated malignancies by targeting a LMP1-specific dependency on lipogenesis. Importance Despite many attempts to develop novel therapies, EBV-specific therapies currently remain largely investigational and EBV-associated malignancies are often associated with a worse prognosis. Therefore, there is a clear demand for EBV-specific therapies for both prevention and treatment of viral-associated malignancies. Non-cancerous cells preferentially obtain fatty acids from dietary sources whereas cancer cells will often produce fatty acids themselves by de novo lipogenesis, often becoming dependent on the pathway for cell survival and proliferation. LMP1 and EBV-mediated B-cell growth transformation leads to induction of FASN, a key enzyme responsible for the catalysis of endogenous fatty acids. Preferential killing of LMP1-expressing B cells following inhibition of FASN suggests that targeting LMP-induced lipogenesis could be an effective strategy in treating LMP1-positive EBV-associated malignancies. Importantly, targeting unique metabolic perturbations induced by EBV could be a way to explicitly target EBV-positive malignancies and distinguish their treatment from EBV-negative counterparts.

scholarly journals CD1c but neither CD1a nor CD1b molecules are expressed on normal, activated, and malignant human B cells: identification of a new B-cell subset

Blood ◽  
1988 ◽  
Vol 72 (1) ◽  
pp. 241-247 ◽  
Author(s):  
D Delia ◽  
G Cattoretti ◽  
N Polli ◽  
E Fontanella ◽  
A Aiello ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
De Novo ◽  
Cell Subset ◽  
Ultrastructural Level ◽  
Cytoplasmic Staining ◽  
Clear Cut ◽  
Hodgkin's Lymphomas

Abstract The CD1 cluster of monoclonal antibodies (MoAbs) CD1a, CD1b, and CD1c, identifies molecules that are differentially expressed on hematopoietic and nonhematopoietic tissues. Our earlier finding that the mantle zone (MZ) but not the germinal center (GC) of normal lymph nodes (LN) is CD1c+, CD1a-, and CD1b- prompted us to further investigate the expression of these molecules on normal, activated, and malignant B cells. We report that blood and spleen contain CD1c+ B cells that account for 49% +/- 20.4% (mean +/- SD) and 50.9% +/- 4.4% of the total B cell population, respectively. CD1a- and CD1b-specific MoAbs are unreactive with both B and T cells; these latter are CD1c- as well. When CD1c+ and CD1c- B cells are activated in vitro, the CD1c molecule is upregulated in the former subset and induced de novo in the latter. Conversely, activated blood T cells remain CD1c-. Neither CD1a nor CD1b molecules are detected on activated T and B lymphocytes. At ultrastructural level, the CD1c+ B cells exhibit distinctive features, namely, condensed chromatin with or without a nucleolus and a unique cluster of cytoplasmic vesicles and organelles; the number of nucleolated cells is higher in the spleen (95%) than in the tonsil (40%) or blood (5%). These findings further confirm the similarity between blood and MZ B cells. The CD1c expression assessed on 27 B-cell chronic lymphocytic leukemias (B-CLL) and 46 B non-Hodgkin's lymphomas (B-NHL) was detected on 41% and 32% of cases, respectively; the latter comprised four follicular and 11 diffuse histotypes. The Burkitt's lymphomas were CD1c-negative. The B-cell neoplasms were all CD1a- and, except for four with a weak cytoplasmic staining, all CD1b- as well. The clear-cut CD1c distribution in normal LN (MZ+, GC-) contrasted with the evidence that some B-NHL cells of GC origin (eg, follicular with predominantly small cleaved cells) were CD1c+. Overall, the finding that CD1c expression is restricted to a fraction of B cells present in lymphoid organs and in peripheral blood indicates that CD1c is a powerful marker for the identification and dissection of B-cell subsets whose functional properties can now be evaluated.

scholarly journals Specific Targeting of Lymphoma Cells Using Semisynthetic Anti-Idiotype Shark Antibodies

2020 ◽  
Vol 11 ◽  
Author(s):  
Arturo Macarrón Palacios ◽  
Julius Grzeschik ◽  
Lukas Deweid ◽  
Simon Krah ◽  
Stefan Zielonka ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lymphoma ◽  
Specific Binding ◽  
B Cell Lymphoma ◽  
Therapeutic Window ◽  
Patient Specific ◽  
Soluble Antigen ◽  
Single Digit ◽  
Lymphoma Therapy

The B-cell receptor (BCR) is a key player of the adaptive immune system. It is a unique part of immunoglobulin (Ig) molecules expressed on the surface of B cells. In case of many B-cell lymphomas, the tumor cells express a tumor-specific and functionally active BCR, also known as idiotype. Utilizing the idiotype as target for lymphoma therapy has emerged to be demanding since the idiotype differs from patient to patient. Previous studies have shown that shark-derived antibody domains (vNARs) isolated from a semi-synthetic CDR3-randomized library allow for the rapid generation of anti-idiotype binders. In this study, we evaluated the potential of generating patient-specific binders against the idiotype of lymphomas. To this end, the BCRs of three different lymphoma cell lines SUP-B8, Daudi, and IM-9 were identified, the variable domains were reformatted and the resulting monoclonal antibodies produced. The SUP-B8 BCR served as antigen in fluorescence-activated cell sorting (FACS)-based screening of the yeast-displayed vNAR libraries which resulted after three rounds of screening in the enrichment of antigen-binding vNARs. Five vNARs were expressed as Fc fusion proteins and consequently analyzed for their binding to soluble antigen using biolayer interferometry (BLI) revealing binding constants in the lower single-digit nanomolar range. These variants showed specific binding to the parental SUP-B8 cell line confirming a similar folding of the recombinantly expressed proteins compared with the native cell surface-presented BCR. First initial experiments to utilize the generated vNAR-Fc variants for BCR-clustering to induce apoptosis or ADCC/ADCP did not result in a significant decrease of cell viability. Here, we report an alternative approach for a personalized B-cell lymphoma therapy based on the construction of vNAR-Fc antibody-drug conjugates to enable specific killing of malignant B cells, which may widen the therapeutic window for B-cell lymphoma therapy.

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