Publications by authors named "Silvia Beà"

105 Publications

Genetic and Phenotypic Attributes of Splenic Marginal Zone Lymphoma.

Blood 2021 Oct 15. Epub 2021 Oct 15.

University of Milan & Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy.

Splenic marginal zone B-cell lymphoma (SMZL) is a heterogeneous clinico-biological entity. The clinical course is variable, multiple genes are mutated with no unifying mechanism, essential regulatory pathways and surrounding microenvironments are diverse. We sought to clarify the heterogeneity of SMZL by resolving different subgroups and their underlying genomic abnormalities, pathway signatures and microenvironment compositions to uncover biomarkers and therapeutic vulnerabilities. We studied 303 SMZL spleen samples collected through the IELSG46 multicenter, international study (NCT02945319) by using a multiplatform approach. We carried out genetic and phenotypic analyses, defined self-organized signatures, validated the findings in independent primary tumor meta-data and in genetically modified mouse models, and determined correlations with outcome data. We identified two prominent genetic clusters in SMZL, termed NNK (58% of cases, harboring NF-κB, NOTCH and KLF2 modules) and DMT (32% of cases, with DNA-damage response, MAPK and TLR modules). Genetic aberrations in multiple genes as well as cytogenetic and immunogenetic features distinguished NNK- from DMT-SMZLs. These genetic clusters not only have distinct underpinning biology, as judged by differences in gene-expression signatures, but also different outcome, with inferior survival in NNK-SMZLs. Digital cytometry and in situ profiling segregated two basic types of SMZL immune microenvironments termed immune-suppressive SMZL (50% of cases, associated with inflammatory cells and immune checkpoint activation) and immune-silent SMZL (50% of cases, associated with an immune-excluded phenotype) with distinct mutational and clinical connotations. In summary, we propose a nosology of SMZL that can implement its classification and also aid in the development of rationally targeted treatments.
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http://dx.doi.org/10.1182/blood.2021012386DOI Listing
October 2021

Clinico-biological features and outcome of patients with splenic marginal zone lymphoma with histological transformation.

Br J Haematol 2021 Sep 14. Epub 2021 Sep 14.

Hematology Department and Hematopathology Section, Pathology Department, Hospital Clínic of Barcelona, Barcelona, Spain.

We describe 36 patients with splenic marginal zone lymphoma (SMZL) with transformation (SMZL-T), including 15 from a series of 84 patients with SMZL diagnosed at the Hospital Clinic of Barcelona (HCB) and 21 diagnosed with SMZL-T in other centres. In the HCB cohort, the cumulative incidence of transformation at 5 years was 15%. Predictors for transformation were cytopenias, hypoalbuminaemia, complex karyotype (CK) and both the Intergruppo Italiano Linfomi (ILL) and simplified Haemoglobin, Platelet count, lactate dehydrogenase (LDH) and extrahilar Lymphadenopathy (HPLL)/ABC scores (P < 0·05). The only independent predictor for transformation in multivariate analysis was CK [hazard ratio (HR) 4·025, P = 0·05]. Patients with SMZL-T had a significantly higher risk of death than the remainder (HR 3·89, P < 0·001). Of the 36 patients with SMZL-T, one developed Hodgkin lymphoma and 35 a diffuse large B-cell lymphoma, 71% with a non-germinal centre phenotype. The main features were B symptoms, lymphadenopathy, and high serum LDH. CK was observed in 12/22 (55%) SMZL-T and fluorescence in situ hybridisation detected abnormalities of MYC proto-oncogene, basic helix-loop-helix transcription factor (MYC), B-cell leukaemia/lymphoma 2 (BCL2) and/or BCL6 in six of 14 (43%). In all, 21 patients received immunochemotherapy, six chemotherapy, one radiotherapy and three splenectomy. The complete response (CR) rate was 61% and the median survival from transformation was 4·92 years. Predictors for a worse survival in multivariate analysis were high-risk International Prognostic Index (HR 5·294, P = 0·016) and lack of CR (HR 2·67, P < 0·001).
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http://dx.doi.org/10.1111/bjh.17815DOI Listing
September 2021

Balanced and unbalanced translocations in a multicentric series of 2843 patients with chronic lymphocytic leukemia.

Genes Chromosomes Cancer 2021 Aug 19. Epub 2021 Aug 19.

Hematopathology Section, Department of Pathology, Hospital Clinic, Barcelona, Spain.

Chromosomal translocations in chronic lymphocytic leukemia (CLL) are very rare, and therefore systematic analysis of large series of cases is needed to allow the identification of recurrent rearrangements, breakpoints involved, and target genes. The aims of the present study were to identify new translocations and their clinical impact and to establish their frequency in a large cohort of 2843 CLL patients. By conventional cytogenetics 250 translocations were identified in 215 (7.5%) patients, 186 (74%) were apparently balanced and 64 (26%) were unbalanced. All chromosomes were involved in translocations, except Y chromosome. The chromosomes more frequently translocated were in decreasing frequency chromosomes 14, 18, 13, 17, 1, 6, 2, 3, 8, and 11. Translocations were found in the karyotypes either as the unique chromosomal abnormality (27%), associated with another alteration (24%), or as a part of a complex karyotype (CK, 48%). A large proportion of rearranged breakpoints involved genes related to CLL such as IGH (14q32), RB1, MIR15A, MIR16-1 (13q14), BCL2 (18q21), IGL (22q11.2), TP53 (17p13), IRF4 (6p25-p23), ATM (11q22), and CDK6 (7q21). Overall, 76 novel CLL translocations were identified, including a recurrent t(8;11)(p21;q21-23). Whole-genome sequencing and/or copy-number microarray data of 24 cases with translocations confirmed all rearrangements, enabled refinement of 3 karyotypes and all breakpoints at gene level. The projected survival and time to first treatment significantly decreased linearly with the number of translocations. In summary, this study allowed to establish the frequency of translocations (7.5%) and to identify new translocations in a cohort of 2843 CLL patients.
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http://dx.doi.org/10.1002/gcc.22994DOI Listing
August 2021

SOX11, CD70 and Treg cells configure the tumor immune microenvironment of aggressive mantle cell lymphoma.

Blood 2021 Jun 29. Epub 2021 Jun 29.

IDIBAPS, Barcelona, Spain.

Mantle cell lymphoma (MCL) is a mature B-cell neoplasm with a heterogeneous clinical and biological behavior. SOX11 oncogenic expression contributes to the aggressiveness of these tumors by different mechanisms including tumor and stromal cell interactions. However, the precise composition of the immune cell microenvironment of MCL, its possible relationship to SOX11 expression, and how it may contribute to tumor behavior is not well known. Here, we performed an integrative transcriptome analysis of 730 immune-related genes combined with the immune cell phenotype analysis by immunohistochemistry in SOX11+ and SOX11- primary nodal MCL cases and non-neoplastic reactive lymph nodes (RLN). SOX11+ MCL had a significant lower T-cell intratumoral infiltration compared to negative cases. A reduced expression of MHCI/II-like and T-cell costimulation and signaling activation related transcripts was significantly associated with poor clinical outcome. Moreover, we identified CD70 as a SOX11 direct target gene, whose overexpression was induced in SOX11+ but not SOX11- tumor cells by CD40L in vitro. CD70 was overexpressed in primary SOX11+ MCL and it was associated with an immune unbalance of the tumor microenvironment characterized by increased number of effector Treg cell infiltration, higher proliferation, and aggressive clinical course. CD27 was expressed with moderate to strong intensity in 76% of cases. Overall, our results suggest that SOX11 expression in MCL is associated with an immunosuppressive microenvironment characterized by CD70 overexpression in tumor cells, increased Treg cell infiltration and downmodulation of antigen-processing and -presentation and T-cell activation that could promote MCL progression and represent a potential target for tailored therapies.
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http://dx.doi.org/10.1182/blood.2020010527DOI Listing
June 2021

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

Nat Cancer 2020 Nov 2;1(11):1066-1081. Epub 2020 Nov 2.

Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain.

We report a systematic analysis of the DNA methylation variability in 1,595 samples of normal cell subpopulations and 14 tumor subtypes spanning the entire human B-cell lineage. Differential methylation among tumor entities relates to differences in cellular origin and to epigenetic alterations, which allowed us to build an accurate machine learning-based diagnostic algorithm. We identify extensive patient-specific methylation variability in silenced chromatin associated with the proliferative history of normal and neoplastic B cells. Mitotic activity generally leaves both hyper- and hypomethylation imprints, but some B-cell neoplasms preferentially gain or lose DNA methylation. Subsequently, we construct a DNA methylation-based mitotic clock called epiCMIT, whose lapse magnitude represents a strong independent prognostic variable in B-cell tumors and is associated with particular driver genetic alterations. Our findings reveal DNA methylation as a holistic tracer of B-cell tumor developmental history, with implications in the differential diagnosis and prediction of clinical outcome.
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http://dx.doi.org/10.1038/s43018-020-00131-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8168619PMC
November 2020

Chromosome banding analysis and genomic microarrays are both useful but not equivalent methods for genomic complexity risk stratification in chronic lymphocytic leukemia patients.

Haematologica 2021 Mar 11. Epub 2021 Mar 11.

Molecular Cytogenetics Laboratory, Pathology Department, Hospital del Mar, Barcelona, Spain; Translational Research on Hematological Neoplasms Group, Cancer Research Program, Institut Hospital del Mar d'Investigacions Mèdiques (IMIM), Barcelona, Spain.

Genome complexity has been associated with poor outcome in patients with chronic lymphocytic leukemia (CLL). Previous cooperative studies established five abnormalities as the cut-off that best predicts an adverse evolution by chromosome banding analysis (CBA) and genomic microarrays (GM). However, data comparing risk stratification by both methods are scarce. Herein, we assessed a cohort of 340 untreated CLL patients highly enriched in cases with complex karyotype (CK, 46.5%) with parallel CBA and GM studies. Abnormalities found by both techniques were compared. Prognostic stratification in three risk groups based on genomic complexity [0-2, 3-4 and ≥5 abnormalities] was also analyzed. No significant differences in the percentage of patients classified into each category were detected, but only a moderate agreement was observed between methods when focusing in individual cases (κ=0.507; p.
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http://dx.doi.org/10.3324/haematol.2020.274456DOI Listing
March 2021

Dynamics of genome architecture and chromatin function during human B cell differentiation and neoplastic transformation.

Nat Commun 2021 01 28;12(1):651. Epub 2021 Jan 28.

Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain.

To investigate the three-dimensional (3D) genome architecture across normal B cell differentiation and in neoplastic cells from different subtypes of chronic lymphocytic leukemia and mantle cell lymphoma patients, here we integrate in situ Hi-C and nine additional omics layers. Beyond conventional active (A) and inactive (B) compartments, we uncover a highly-dynamic intermediate compartment enriched in poised and polycomb-repressed chromatin. During B cell development, 28% of the compartments change, mostly involving a widespread chromatin activation from naive to germinal center B cells and a reversal to the naive state upon further maturation into memory B cells. B cell neoplasms are characterized by both entity and subtype-specific alterations in 3D genome organization, including large chromatin blocks spanning key disease-specific genes. This study indicates that 3D genome interactions are extensively modulated during normal B cell differentiation and that the genome of B cell neoplasias acquires a tumor-specific 3D genome architecture.
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http://dx.doi.org/10.1038/s41467-020-20849-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7844026PMC
January 2021

Monomorphic Epitheliotropic Intestinal T-Cell Lymphoma in Asia Frequently Shows Alterations.

Cancers (Basel) 2020 Nov 27;12(12). Epub 2020 Nov 27.

Department of Pathology, School of Medicine, Tokai University, 143 Shimokasuya, Kanagawa, Isehara 259-1193, Japan.

Monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL) is a rare primary T-cell lymphoma of the digestive tract derived from intraepithelial lymphocytes and characterized by an aggressive clinical course. In this study, nine cases of Japanese MEITL were analyzed by targeted Next Generation Sequencing (NGS) and immunohistochemistry and were integrated with previously reported whole-genome copy number microarray-based assay data. The highlight of our findings is that all cases showed alterations of the tumor suppressor gene by mutations and/or loss of the corresponding 3p21 locus. We also demonstrated that all cases showed mutations in one or more genes of JAK/STAT pathway. Therefore, the combination of epigenetic deregulation and cell signaling activation represent major oncogenic events in the pathogenesis of MEITL in Asian MEITL, similar to Western MEITL.
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http://dx.doi.org/10.3390/cancers12123539DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7759862PMC
November 2020

Mutational Landscape and Tumor Burden Assessed by Cell-free DNA in Diffuse Large B-Cell Lymphoma in a Population-Based Study.

Clin Cancer Res 2021 01 29;27(2):513-521. Epub 2020 Oct 29.

Department of Hematology, Hospital Clínic de Barcelona, Barcelona, Spain.

Purpose: We analyzed the utility of cell-free DNA (cfDNA) in a prospective population-based cohort to determine the mutational profile, assess tumor burden, and estimate its impact in response rate and outcome in patients with diffuse large B-cell lymphoma (DLBCL).

Experimental Design: A total of 100 patients were diagnosed with DLBCL during the study period. Mutational status of 112 genes was studied in cfDNA by targeted next-generation sequencing. Paired formalin-fixed, paraffin-embedded samples and volumetric PET/CT were assessed when available.

Results: Appropriate cfDNA to perform the analyses was obtained in 79 of 100 cases. At least one mutation could be detected in 69 of 79 cases (87%). The sensitivity of cfDNA to detect the mutations was 68% (95% confidence interval, 56.2-78.7). The mutational landscape found in cfDNA samples was highly consistent with that shown in the tissue and allowed genetic classification in 43% of the cases. A higher amount of circulating tumor DNA (ctDNA) significantly correlated with clinical parameters related to tumor burden (elevated lactate dehydrogenase and β2-microglobulin serum levels, advanced stage, and high-risk International Prognostic Index) and total metabolic tumor volume assessed by PET/CT. In patients treated with curative intent, high ctDNA levels (>2.5 log hGE/mL) were associated with lower complete response (65% vs. 96%; < 0.004), shorter progression-free survival (65% vs. 85%; = 0.038), and overall survival (73% vs. 100%; = 0.007) at 2 years, although it did not maintain prognostic value in multivariate analyses.

Conclusions: In a population-based prospective DLBCL series, cfDNA resulted as an alternative source to estimate tumor burden and to determine the tumor mutational profile and genetic classification, which have prognostic implications and may contribute to a future tailored treatment.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-2558DOI Listing
January 2021

A Cyclin D1-Dependent Transcriptional Program Predicts Clinical Outcome in Mantle Cell Lymphoma.

Clin Cancer Res 2021 01 12;27(1):213-225. Epub 2020 Oct 12.

Lymphoid Neoplasm Program, August Pi i Sunyer Biomedical Research Institute (IDIBAPS), Barcelona, Spain.

Purpose: Mantle cell lymphoma (MCL) is characterized by the t(11;14)(q13;q32) translocation leading to cyclin D1 overexpression. Cyclin D1 is a major cell-cycle regulator and also regulates transcription, but the impact of cyclin D1-mediated transcriptional dysregulation on MCL pathogenesis remains poorly understood. The aim of this study was to define a cyclin D1-dependent gene expression program and analyze its prognostic value.

Experimental Design: We integrated genome-wide expression analysis of cyclin D1-silenced and overexpressing cells with cyclin D1 chromatin-binding profiles to identify a cyclin D1-dependent transcriptional program in MCL cells. We analyzed this gene program in two MCL series of peripheral blood samples ( = 53) and lymphoid tissues ( = 106) to determine its biological and clinical relevance. We then obtained a simplified signature of this program and evaluated a third series of peripheral blood MCL samples ( = 81) by NanoString gene expression profiling to validate our findings.

Results: We identified a cyclin D1-dependent transcriptional program composed of 295 genes that were mainly involved in cell-cycle control. The cyclin D1-dependent gene program was overexpressed in MCL tumors directly proportional to cyclin D1 levels. High expression of this program conferred an adverse prognosis with significant shorter overall survival of the patients. These observations were validated in an independent cohort of patients using a simplified 37-gene cyclin D1 signature. The cyclin D1-dependent transcriptional program was also present in multiple myeloma and breast tumors with cyclin D1 overexpression.

Conclusions: We identified a cyclin D1-dependent transcriptional program that is overexpressed in MCL and predicts clinical outcome.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-2868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8051616PMC
January 2021

PI3Kδ inhibition reshapes follicular lymphoma-immune microenvironment cross talk and unleashes the activity of venetoclax.

Blood Adv 2020 09;4(17):4217-4231

Department of Hematology-Oncology, Institut d'Investigacions Biomèdiques August Pi I Sunyer (IDIBAPS), Barcelona, Spain.

Despite idelalisib approval in relapsed follicular lymphoma (FL), a complete characterization of the immunomodulatory consequences of phosphatidylinositol 3-kinase δ (PI3Kδ) inhibition, biomarkers of response, and potential combinatorial therapies in FL remain to be established. Using ex vivo cocultures of FL patient biopsies and follicular dendritic cells (FDCs) to mimic the germinal center (n = 42), we uncovered that PI3Kδ inhibition interferes with FDC-induced genes related to angiogenesis, extracellular matrix formation, and transendothelial migration in a subset of FL samples, defining an 18-gene signature fingerprint of idelalisib sensitivity. A common hallmark of idelalisib found in all FL cases was its interference with the CD40/CD40L pathway and induced proliferation, together with the downregulation of proteins crucial for B-T-cell synapses, leading to an inefficient cross talk between FL cells and the supportive T-follicular helper cells (TFH). Moreover, idelalisib downmodulates the chemokine CCL22, hampering the recruitment of TFH and immunosupressive T-regulatory cells to the FL niche, leading to a less supportive and tolerogenic immune microenvironment. Finally, using BH3 profiling, we uncovered that FL-FDC and FL-macrophage cocultures augment tumor addiction to BCL-XL and MCL-1 or BFL-1, respectively, limiting the cytotoxic activity of the BCL-2 inhibitor venetoclax. Idelalisib restored FL dependence on BCL-2 and venetoclax activity. In summary, idelalisib exhibits a patient-dependent activity toward angiogenesis and lymphoma dissemination. In all FL cases, idelalisib exerts a general reshaping of the FL immune microenvironment and restores dependence on BCL-2, predisposing FL to cell death, providing a mechanistic rationale for investigating the combination of PI3Kδ inhibitors and venetoclax in clinical trials.
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http://dx.doi.org/10.1182/bloodadvances.2020001584DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7479943PMC
September 2020

Molecular Pathogenesis of Mantle Cell Lymphoma.

Hematol Oncol Clin North Am 2020 10 22;34(5):795-807. Epub 2020 Jul 22.

Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Rosselló, 149-153, Barcelona 08036, Spain; Centro de Investigación Biomédica en Red de Cáncer, Madrid, Spain; Hematopathology Unit, Hospital Clínic of Barcelona, University of Barcelona, Villarroel 170, Barcelona 08036, Spain. Electronic address:

Mantle cell lymphoma (MCL) is a mature B-cell neoplasm with heterogeneous clinical behavior molecularly characterized by the constitutive overexpression of cyclin D1 and deregulation of different signaling pathways. SOX11 expression determines an aggressive phenotype associated with accumulation of many chromosomal alterations and somatic gene mutations. A subset of patients with the SOX11-negative leukemic non-nodal MCL subtype follows an initial indolent clinical evolution and may not require treatment at diagnosis, although eventually may progress to an aggressive disease. We discuss the genetic and molecular alterations with impact on the cancer hallmarks that characterize the lymphomagenesis of the 2 MCL subtypes.
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http://dx.doi.org/10.1016/j.hoc.2020.05.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7473344PMC
October 2020

IgCaller for reconstructing immunoglobulin gene rearrangements and oncogenic translocations from whole-genome sequencing in lymphoid neoplasms.

Nat Commun 2020 07 7;11(1):3390. Epub 2020 Jul 7.

Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain.

Immunoglobulin (Ig) gene rearrangements and oncogenic translocations are routinely assessed during the characterization of B cell neoplasms and stratification of patients with distinct clinical and biological features, with the assessment done using Sanger sequencing, targeted next-generation sequencing, or fluorescence in situ hybridization (FISH). Currently, a complete Ig characterization cannot be extracted from whole-genome sequencing (WGS) data due to the inherent complexity of the Ig loci. Here, we introduce IgCaller, an algorithm designed to fully characterize Ig gene rearrangements and oncogenic translocations from short-read WGS data. Using a cohort of 404 patients comprising different subtypes of B cell neoplasms, we demonstrate that IgCaller identifies both heavy and light chain rearrangements to provide additional information on their functionality, somatic mutational status, class switch recombination, and oncogenic Ig translocations. Our data thus support IgCaller to be a reliable alternative to Sanger sequencing and FISH for studying the genetic properties of the Ig loci.
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http://dx.doi.org/10.1038/s41467-020-17095-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7341758PMC
July 2020

Genomic and epigenomic insights into the origin, pathogenesis, and clinical behavior of mantle cell lymphoma subtypes.

Blood 2020 09;136(12):1419-1432

Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain.

Mantle cell lymphoma (MCL) is a mature B-cell neoplasm initially driven by CCND1 rearrangement with 2 molecular subtypes, conventional MCL (cMCL) and leukemic non-nodal MCL (nnMCL), that differ in their clinicobiological behavior. To identify the genetic and epigenetic alterations determining this diversity, we used whole-genome (n = 61) and exome (n = 21) sequencing (74% cMCL, 26% nnMCL) combined with transcriptome and DNA methylation profiles in the context of 5 MCL reference epigenomes. We identified that open and active chromatin at the major translocation cluster locus might facilitate the t(11;14)(q13;32), which modifies the 3-dimensional structure of the involved regions. This translocation is mainly acquired in precursor B cells mediated by recombination-activating genes in both MCL subtypes, whereas in 8% of cases the translocation occurs in mature B cells mediated by activation-induced cytidine deaminase. We identified novel recurrent MCL drivers, including CDKN1B, SAMHD1, BCOR, SYNE1, HNRNPH1, SMARCB1, and DAZAP1. Complex structural alterations emerge as a relevant early oncogenic mechanism in MCL, targeting key driver genes. Breakage-fusion-bridge cycles and translocations activated oncogenes (BMI1, MIR17HG, TERT, MYC, and MYCN), generating gene amplifications and remodeling regulatory regions. cMCL carried significant higher numbers of structural variants, copy number alterations, and driver changes than nnMCL, with exclusive alterations of ATM in cMCL, whereas TP53 and TERT alterations were slightly enriched in nnMCL. Several drivers had prognostic impact, but only TP53 and MYC aberrations added value independently of genomic complexity. An increasing genomic complexity, together with the presence of breakage-fusion-bridge cycles and high DNA methylation changes related to the proliferative cell history, defines patients with different clinical evolution.
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http://dx.doi.org/10.1182/blood.2020005289DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7498364PMC
September 2020

FMOD expression in whole blood aids in distinguishing between chronic lymphocytic leukemia and other leukemic lymphoproliferative disorders. A pilot study.

Cytometry B Clin Cytom 2020 09 12;98(5):421-428. Epub 2020 Jun 12.

Hematology Laboratory, ICO-Hospital Germans Trias i Pujol, Institut de Recerca Josep Carreras, Universitat Autònoma de Barcelona, Badalona, Spain.

Background: Within the hematopoietic compartment, fibromodulin (FMOD) is almost exclusively expressed in chronic lymphocytic leukemia (CLL) lymphocytes. We set out to determine whether FMOD could be of help in diagnosing borderline lymphoproliferative disorders (LPD).

Methods: We established 3 flow cytometry-defined groups (CLL [n = 65], borderline LPD [n = 28], broadly defined as those with CLLflow score between 35 and -20 or discordant CD43 and CLLflow, and non-CLL LPD [n = 40]). FMOD expression levels were determined by standard RT-PCR in whole-blood samples. Patients were included regardless of lymphocyte count but with tumor burden ≥40%.

Results: FMOD expression levels distinguished between CLL (median 98.5, interquartile range [IQR] 37.8-195.1) and non-CLL LPD (median 0.012, IQR 0.003-0.033) with a sensitivity and specificity of 1. Most borderline LPDs were CD5/CD23/CD200-positive with no loss of B-cell antigens and negative or partial expression of CD43. 16/22 patients with available cytogenetic analysis showed trisomy 12. In 25/28 (89%) of these patients, FMOD expression levels fell between CLL and non-CLL (median 3.58, IQR 1.06-6.21).

Discussion: This study could suggest that borderline LPDs may constitute a distinct group laying in the biological spectrum of chronic leukemic LPDs. Future studies will have to confirm these results with other biological data. Quantification of FMOD can potentially be of help in the diagnosis of phenotypically complex LPDs.
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http://dx.doi.org/10.1002/cyto.b.21890DOI Listing
September 2020

Reproducibility of histologic prognostic parameters for mantle cell lymphoma: cytology, Ki67, p53 and SOX11.

Virchows Arch 2020 Aug 23;477(2):259-267. Epub 2020 Jan 23.

Hematopathology Section, Institute of Pathology and Lymph Node Registry, Universitätsklinikum Schleswig-Holstein, Campus Kiel, Kiel, Germany.

Mantle cell lymphoma (MCL) shows a clinical aggressiveness that varies from patient to patient. Despite major advances in outcomes with current immunochemotherapy, the future development of therapies requires risk stratification to tailor therapy intensity. Within the group of reference pathologists for the ongoing trials of the European MCL Network, we performed a round robin test on a tissue microarray to evaluate the reproducibility in assessing the biomarkers of outcome in MCL. Cytological subtype, Ki67-index and expression of p53 and SOX11 were evaluated on 20 diagnostic tumour samples by eight participating labs independently. We demonstrate that the assessment of the proliferation index by counting the Ki67 positive cells as well as assessment of SOX11 and p53 expression status is reproducible between labs. For the most established prognostic biomarker, Ki67, the intra-class correlation coefficient was very good when assessed as a continuous parameter (0.87). The agreement was lower when the values were analysed in a dichotomized way applying the commonly used cutoff of 30% (kappa = 0.65, complete concordance of all labs in 13/20 (65%)). Cases with discrepant results between labs in the dichotomized analysis showed mean values close to the cutoff of 30%. Centralised scoring and digital image analysis revealed results in line with the scores from individual labs. All cases in our cohort were additionally assessed for gene expression signatures and of TP53 gene alterations. Given the good reproducibility when guidelines of assessment are applied, the biomarker studied in this inter-laboratory test presents potential candidates to be enhanced for risk-stratification in the future clinical trials.
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http://dx.doi.org/10.1007/s00428-020-02750-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415076PMC
August 2020

Notch1 signaling in NOTCH1-mutated mantle cell lymphoma depends on Delta-Like ligand 4 and is a potential target for specific antibody therapy.

J Exp Clin Cancer Res 2019 Nov 1;38(1):446. Epub 2019 Nov 1.

Experimental Therapeutics in Lymphoid Malignancies Group, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain.

Background: NOTCH1 gene mutations in mantle cell lymphoma (MCL) have been described in about 5-10% of cases and are associated with significantly shorter survival rates. The present study aimed to investigate the biological impact of this mutation in MCL and its potential as a therapeutic target.

Methods: Activation of Notch1 signaling upon ligand-stimulation and inhibitory effects of the monoclonal anti-Notch1 antibody OMP-52M51 in NOTCH1-mutated and -unmutated MCL cells were assessed by Western Blot and gene expression profiling. Effects of OMP-52M51 treatment on tumor cell migration and tumor angiogenesis were evaluated with chemotaxis and HUVEC tube formation assays. The expression of Delta-like ligand 4 (DLL4) in MCL lymph nodes was analyzed by immunofluorescence staining and confocal microscopy. A MCL mouse model was used to assess the activity of OMP-52M51 in vivo.

Results: Notch1 expression can be effectively stimulated in NOTCH1-mutated Mino cells by DLL4, whereas in the NOTCH1-unmutated cell line JeKo-1, less effect was observed upon any ligand-stimulation. DLL4 was expressed by histiocytes in both, NOTCH1-mutated and -unmutated MCL lymph nodes. Treatment of NOTCH1-mutated MCL cells with the monoclonal anti-Notch1 antibody OMP-52M51 effectively prevented DLL4-dependent activation of Notch1 and suppressed the induction of numerous direct Notch target genes involved in lymphoid biology, lymphomagenesis and disease progression. Importantly, in lymph nodes from primary MCL cases with NOTCH1/2 mutations, we detected an upregulation of the same gene sets as observed in DLL4-stimulated Mino cells. Furthermore, DLL4 stimulation of NOTCH1-mutated Mino cells enhanced tumor cell migration and angiogenesis, which could be abolished by treatment with OMP-52M51. Importantly, the effects observed were specific for NOTCH1-mutated cells as they did not occur in the NOTCH1-wt cell line JeKo-1. Finally, we confirmed the potential activity of OMP-52M51 to inhibit DLL4-induced Notch1-Signaling in vivo in a xenograft mouse model of MCL.

Conclusion: DLL4 effectively stimulates Notch1 signaling in NOTCH1-mutated MCL and is expressed by the microenvironment in MCL lymph nodes. Our results indicate that specific inhibition of the Notch1-ligand-receptor interaction might provide a therapeutic alternative for a subset of MCL patients.
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http://dx.doi.org/10.1186/s13046-019-1458-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6825347PMC
November 2019

Insight into genetic predisposition to chronic lymphocytic leukemia from integrative epigenomics.

Nat Commun 2019 08 9;10(1):3615. Epub 2019 Aug 9.

Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036, Barcelona, Spain.

Genome-wide association studies have provided evidence for inherited genetic predisposition to chronic lymphocytic leukemia (CLL). To gain insight into the mechanisms underlying CLL risk we analyze chromatin accessibility, active regulatory elements marked by H3K27ac, and DNA methylation at 42 risk loci in up to 486 primary CLLs. We identify that risk loci are significantly enriched for active chromatin in CLL with evidence of being CLL-specific or differentially regulated in normal B-cell development. We then use in situ promoter capture Hi-C, in conjunction with gene expression data to reveal likely target genes of the risk loci. Candidate target genes are enriched for pathways related to B-cell development such as MYC and BCL2 signalling. At 14 loci the analysis highlights 63 variants as the probable functional basis of CLL risk. By integrating genetic and epigenetic information our analysis reveals novel insights into the relationship between inherited predisposition and the regulatory chromatin landscape of CLL.
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http://dx.doi.org/10.1038/s41467-019-11582-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6689100PMC
August 2019

Increased tumour angiogenesis in SOX11-positive mantle cell lymphoma.

Histopathology 2019 Nov 19;75(5):704-714. Epub 2019 Aug 19.

Haematopathology Unit, Pathology Department, Hospital Clinic, University of Barcelona, Barcelona, Spain.

Aims: Mantle cell lymphoma (MCL) is a heterogeneous disease with an aggressive behaviour in most cases, which is associated with expression of sex determining region-Y-box11 (SOX11). Experimental studies have shown that SOX11 expression is associated with an angiogenic switch characterised by increased expression of angiogenic-related signatures and vascularisation of murine tumours. However, the relationship between angiogenesis and SOX11 expression in primary tumours is not well understood. Therefore, the aim of this study was to evaluate the development of microvascular angiogenesis in primary MCL in relation to SOX11 expression and its potential prognostic value.

Methods And Results: Fifty-six patients diagnosed with MCL, 38 SOX11-positive and 18 SOX11-negative, were studied. The relative intratumoral microvascular area (MVA) and microvessel density (MVD) (number of intratumoral microvessels/μm ) were measured on CD34-stained slides using a computerised image analysis system. SOX11-positive MCL showed a significant higher microvascular development than negative tumours (median MVA = 14.5 × 10 versus 5.0 × 10 P < 0.001; median MVD = 18.6/μm versus 14.2/μm , P = 0.021). Analysing the MVA and MVD as continuous variables, a high MVD was associated with shorter overall survival (P = 0.004), and a similar tendency was observed for high MVA (P = 0.064). The microvascular development was not related to the Ki-67 proliferative index or 17p/TP53, 9p or 11q alterations.

Conclusions: These findings suggest that SOX11 promotes an angiogenic phenotype in primary MCL, which may contribute to the more aggressive behaviour of these tumours.
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http://dx.doi.org/10.1111/his.13935DOI Listing
November 2019

and hijack immunoglobulin light-chain enhancers in cyclin D1 mantle cell lymphoma.

Blood 2019 02 11;133(9):940-951. Epub 2018 Dec 11.

Department of Clinical Pathology and.

Mantle cell lymphoma (MCL) is characterized by the t(11;14)(q13;q32) translocation resulting in overexpression of cyclin D1. However, a small subset of cyclin D1 MCL has been recognized, and approximately one-half of them harbor translocations while the primary event in cyclin D1/D2 MCL remains elusive. To identify other potential mechanisms driving MCL pathogenesis, we investigated 56 cyclin D1/SOX11 MCL by fluorescence in situ hybridization (FISH), whole-genome/exome sequencing, and gene-expression and copy-number arrays. FISH with break-apart probes identified rearrangements in 39 cases (70%) but not rearrangements. We analyzed 3 of these negative cases by whole-genome/exome sequencing and identified IGK (n = 2) and IGL (n = 1) enhancer hijackings near that were associated with cyclin D3 overexpression. By specific FISH probes, including the IGK enhancer region, we detected 10 additional cryptic IGK juxtapositions to (6 cases) and (4 cases) in MCL that overexpressed, respectively, these cyclins. A minor subset of 4 cyclin D1 MCL cases lacked cyclin D rearrangements and showed upregulation of and These cases had blastoid morphology, high genomic complexity, and and deletions. Both genomic and gene-expression profiles of cyclin D1 MCL cases were indistinguishable from cyclin D1 MCL. In conclusion, virtually all cyclin D1 MCLs carry rearrangements with immunoglobulin genes, including a novel IGK/L enhancer hijacking mechanism. A subset of cyclin D1/D2/D3 MCL with aggressive features has cyclin E dysregulation. Specific FISH probes may allow the molecular identification and diagnosis of cyclin D1 MCL.
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http://dx.doi.org/10.1182/blood-2018-07-862151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6396173PMC
February 2019

Cyclin D1 overexpression induces global transcriptional downregulation in lymphoid neoplasms.

J Clin Invest 2018 08 20;128(9):4132-4147. Epub 2018 Aug 20.

Lymphoid Neoplasm Program and.

Cyclin D1 is an oncogene frequently overexpressed in human cancers that has a dual function as cell cycle and transcriptional regulator, although the latter is widely unexplored. Here, we investigated the transcriptional role of cyclin D1 in lymphoid tumor cells with cyclin D1 oncogenic overexpression. Cyclin D1 showed widespread binding to the promoters of most actively transcribed genes, and the promoter occupancy positively correlated with the transcriptional output of targeted genes. Despite this association, the overexpression of cyclin D1 in lymphoid cells led to a global transcriptional downmodulation that was proportional to cyclin D1 levels. This cyclin D1-dependent global transcriptional downregulation was associated with a reduced nascent transcription and an accumulation of promoter-proximal paused RNA polymerase II (Pol II) that colocalized with cyclin D1. Concordantly, cyclin D1 overexpression promoted an increase in the Poll II pausing index. This transcriptional impairment seems to be mediated by the interaction of cyclin D1 with the transcription machinery. In addition, cyclin D1 overexpression sensitized cells to transcription inhibitors, revealing a synthetic lethality interaction that was also observed in primary mantle cell lymphoma cases. This finding of global transcriptional dysregulation expands the known functions of oncogenic cyclin D1 and suggests the therapeutic potential of targeting the transcriptional machinery in cyclin D1-overexpressing tumors.
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http://dx.doi.org/10.1172/JCI96520DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6118591PMC
August 2018

The reference epigenome and regulatory chromatin landscape of chronic lymphocytic leukemia.

Nat Med 2018 06 21;24(6):868-880. Epub 2018 May 21.

European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Wellcome Trust Genome Campus, Hinxton, UK.

Chronic lymphocytic leukemia (CLL) is a frequent hematological neoplasm in which underlying epigenetic alterations are only partially understood. Here, we analyze the reference epigenome of seven primary CLLs and the regulatory chromatin landscape of 107 primary cases in the context of normal B cell differentiation. We identify that the CLL chromatin landscape is largely influenced by distinct dynamics during normal B cell maturation. Beyond this, we define extensive catalogues of regulatory elements de novo reprogrammed in CLL as a whole and in its major clinico-biological subtypes classified by IGHV somatic hypermutation levels. We uncover that IGHV-unmutated CLLs harbor more active and open chromatin than IGHV-mutated cases. Furthermore, we show that de novo active regions in CLL are enriched for NFAT, FOX and TCF/LEF transcription factor family binding sites. Although most genetic alterations are not associated with consistent epigenetic profiles, CLLs with MYD88 mutations and trisomy 12 show distinct chromatin configurations. Furthermore, we observe that non-coding mutations in IGHV-mutated CLLs are enriched in H3K27ac-associated regulatory elements outside accessible chromatin. Overall, this study provides an integrative portrait of the CLL epigenome, identifies extensive networks of altered regulatory elements and sheds light on the relationship between the genetic and epigenetic architecture of the disease.
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http://dx.doi.org/10.1038/s41591-018-0028-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6363101PMC
June 2018

A gene signature that distinguishes conventional and leukemic nonnodal mantle cell lymphoma helps predict outcome.

Blood 2018 07 16;132(4):413-422. Epub 2018 May 16.

Institute for Biomedical Research August Pi i Sunyer, Barcelona, Spain.

Mantle cell lymphoma (MCL) is an aggressive B-cell malignancy, but some patients have a very indolent evolution. This heterogeneous course is related, in part, to the different biological characteristics of conventional MCL (cMCL) and the distinct subgroup of leukemic nonnodal MCL (nnMCL). Robust criteria to distinguish these MCL subtypes and additional biological parameters that influence their evolution are not well defined. We describe a novel molecular assay that reliably distinguishes cMCL and nnMCL using blood samples. We trained a 16-gene assay (L-MCL16 assay) on the NanoString platform using 19 purified leukemic samples. The locked assay was applied to an independent cohort of 70 MCL patients with leukemic presentation. The assay assigned 37% of cases to nnMCL and 56% to cMCL. nnMCL and cMCL differed in nodal presentation, lactate dehydrogenase, immunoglobulin heavy chain gene mutational status, management options, genomic complexity, and / deletions, but the proportion with 17p/ aberrations was similar in both subgroups. Sequential samples showed that assay prediction was stable over time. nnMCL had a better overall survival (OS) than cMCL (3-year OS 92% vs 69%; = .006) from the time of diagnosis and longer time to first treatment. Genomic complexity and / aberrations predicted for shorter OS in the entire series and cMCL, whereas only genomic complexity was associated with shorter time to first treatment and OS in nnMCL. In conclusion, the newly developed assay robustly recognizes the 2 molecular subtypes of MCL in leukemic samples. Its combination with genetic alterations improves the prognostic evaluation and may provide useful biological information for management decisions.
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http://dx.doi.org/10.1182/blood-2018-03-838136DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6071558PMC
July 2018
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