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Diffuse large B-cell lymphoma (DLBCL) includes the activated B cell-like (ABC) and germinal center B cell-like (GCB) subtypes, which differ in cell of origin, genetics, and clinical response. By screening the subtype-specific activity of 211 drugs approved or in active clinical development for other diseases, we identified inhibitors of nicotinamide phosphoribosyl transferase (NAMPTi) as active in a subset of GCB-DLBCL in vitro and in vivo. We validated three chemically distinct NAMPTis for their on-target activity based on biochemical and genetic rescue approaches and found the ratio between NAMPT and PARP1 RNA levels was predictive of NAMPTi sensitivity across DLBCL subtypes. Notably, the NAMPT:PARP1 transcript ratio predicts higher antitumor activity in BCL2-translocated GCB-DLBCL. Accordingly, pharmacologic and genetic inhibition of BCL2 was potently synergistic with NAMPT blockade. These data support the inhibition of NAMPT as a therapeutically relevant strategy for BCL2-translocated DLBCLs. Significance: Targeted therapies have emerged for the ABC subtype of DLBCL, but not for the GCB subtype, despite the evidence of a significant subset of high-risk cases. We identify a drug that specifically targets a subset of GCB-DLBCL and provide preclinical evidence for BCL2 translocations as biomarkers for their identification., (©2024 American Association for Cancer Research.)

The myocyte enhancer factor 2B (MEF2B) transcription factor is frequently mutated in germinal center (GC)-derived B-cell lymphomas. Its ammino (N)-terminal mutations drive lymphomagenesis by escaping interaction with transcriptional repressors, while the function of carboxy (C)-terminal mutations remains to be elucidated. Here, we show that MEF2B C-tail is physiologically phosphorylated at specific residues and phosphorylation at serine (S)324 is impaired by lymphoma-associated mutations. Lack of phosphorylation at S324 enhances the interaction of MEF2B with the SWI/SNF chromatin remodeling complex, leading to higher transcriptional activity. In addition, these mutants show an increased protein stability due to impaired interaction with the CUL3/KLHL12 ubiquitin complex. Mice expressing a phosphorylation-deficient lymphoma-associated MEF2B mutant display GC enlargement and develop GC-derived lymphomas, when crossed with Bcl2 transgenic mice. These results unveil converging mechanisms of action for a diverse spectrum of MEF2B mutations, all leading to its dysregulation and GC B-cell lymphomagenesis., (© 2024. The Author(s).)

Interplay between chromatin-associated complexes and modifications critically contribute to the partitioning of epigenome into stable and functionally distinct domains. Yet there is a lack of systematic identification of chromatin crosstalk mechanisms, limiting our understanding of the dynamic transition between chromatin states during development and disease. Here we perform co-dependency mapping of genes using CRISPR-Cas9-mediated fitness screens in pan-cancer cell lines to quantify gene-gene functional relationships. We identify 145 co-dependency modules and further define the molecular context underlying the essentiality of these modules by incorporating mutational, epigenome, gene expression and drug sensitivity profiles of cell lines. These analyses assign new protein complex composition and function, and predict new functional interactions, including an unexpected co-dependency between two transcriptionally counteracting chromatin complexes - polycomb repressive complex 2 (PRC2) and MLL-MEN1 complex. We show that PRC2-mediated H3K27 tri-methylation regulates the genome-wide distribution of MLL1 and MEN1. In lymphoma cells with EZH2 gain-of-function mutations, the re-localization of MLL-MEN1 complex drives oncogenic gene expression and results in a hypersensitivity to pharmacologic inhibition of MEN1. Together, our findings provide a resource for discovery of trans-regulatory interactions as mechanisms of chromatin regulation and potential targets of synthetic lethality., (© 2023. The Author(s).)

The adaptive immune system has evolved to allow effective responses against a virtually unlimited number of invading pathogens. This process requires the transient formation of germinal centers (GC), a dynamic environment that ensures the generation and selection of B cells capable to produce antibodies with high antigen affinity, or to maintain the memory of that antigen for life. However, this comes at a cost, as the unique events accompanying the GC reaction pose a significant risk to the genome of B cells, which must endure elevated levels of replication stress, while proliferating at high rates and undergoing DNA breaks introduced by somatic hypermutation and class switch recombination. Indeed, the genetic/epigenetic disruption of programs implicated in normal GC biology has emerged as a hallmark of most B cell lymphomas. This improved understanding provides a conceptual framework for the identification of cellular pathways that could be exploited for precision medicine approaches., (© 2023 John Wiley & Sons Ltd.)

Heterozygous inactivating mutations of the KMT2D methyltransferase and the CREBBP acetyltransferase are among the most common genetic alterations in B cell lymphoma and co-occur in 40 to 60% of follicular lymphoma (FL) and 30% of EZB/C3 diffuse large B cell lymphoma (DLBCL) cases, suggesting they may be coselected. Here, we show that combined germinal center (GC)-specific haploinsufficiency of Crebbp and Kmt2d synergizes in vivo to promote the expansion of abnormally polarized GCs, a common preneoplastic event. These enzymes form a biochemical complex on select enhancers/superenhancers that are critical for the delivery of immune signals in the GC light zone and are only corrupted upon dual Crebbp / Kmt2d loss, both in mouse GC B cells and in human DLBCL. Moreover, CREBBP directly acetylates KMT2D in GC-derived B cells, and, consistently, its inactivation by FL/DLBCL-associated mutations abrogates its ability to catalyze KMT2D acetylation. Genetic and pharmacologic loss of CREBBP and the consequent decrease in KMT2D acetylation lead to reduced levels of H3K4me1, supporting a role for this posttranslational modification in modulating KMT2D activity. Our data identify a direct biochemical and functional interaction between CREBBP and KMT2D in the GC, with implications for their role as tumor suppressors in FL/DLBCL and for the development of precision medicine approaches targeting enhancer defects induced by their combined loss.

With the introduction of large-scale molecular profiling methods and high-throughput sequencing technologies, the genomic features of most lymphoid neoplasms have been characterized at an unprecedented scale. Although the principles for the classification and diagnosis of these disorders, founded on a multidimensional definition of disease entities, have been consolidated over the past 25 years, novel genomic data have markedly enhanced our understanding of lymphomagenesis and enriched the description of disease entities at the molecular level. Yet, the current diagnosis of lymphoid tumors is largely based on morphological assessment and immunophenotyping, with only few entities being defined by genomic criteria. This paper, which accompanies the International Consensus Classification of mature lymphoid neoplasms, will address how established assays and newly developed technologies for molecular testing already complement clinical diagnoses and provide a novel lens on disease classification. More specifically, their contributions to diagnosis refinement, risk stratification, and therapy prediction will be considered for the main categories of lymphoid neoplasms. The potential of whole-genome sequencing, circulating tumor DNA analyses, single-cell analyses, and epigenetic profiling will be discussed because these will likely become important future tools for implementing precision medicine approaches in clinical decision making for patients with lymphoid malignancies., (© 2022 by The American Society of Hematology.)

Most B cell lymphomas arise from the oncogenic transformation of B cells that have undergone the germinal center (GC) reaction of the T cell-dependent immune response, where high-affinity memory B cells and plasma cells are generated. The high proliferation of GC B cells coupled with occasional errors in the DNA-modifying processes of somatic hypermutation and class switch recombination put the cell at a risk to obtain transforming genetic aberrations, which may activate proto-oncogenes or inactivate tumour suppressor genes. Several subtypes of GC lymphomas harbor genetic mutations leading to constitutive, aberrant activation of the nuclear factor-κB (NF-κB) signaling pathway. In normal B cells, NF-κB has crucial biological roles in development and physiology. GC lymphomas highjack these activities to promote tumour-cell growth and survival. It has become increasingly clear that the separate canonical and non-canonical routes of the NF-κB pathway and the five downstream NF-κB transcription factors have distinct functions in the successive stages of GC B-cell development. These findings may have direct implications for understanding how aberrant NF-κB activation promotes the genesis of various GC lymphomas corresponding to the developmentally distinct GC B-cell subsets. The knowledge arising from these studies may be explored for the development of precision medicine approaches aimed at more effective treatments of the corresponding tumours with specific NF-κB inhibitors, thus reducing systemic toxicity. We here provide an overview on the patterns of genetic NF-κB mutations encountered in the various GC lymphomas and discuss the consequences of aberrant NF-κB activation in those malignancies as related to the biology of NF-κB in their putative normal cellular counterparts.

Diffuse large B cell lymphoma (DLBCL) is the most common B cell non-Hodgkin lymphoma and remains incurable in around 40% of patients. Efforts to sequence the coding genome identified several genes and pathways that are altered in this disease, including potential therapeutic targets 1-5 . However, the non-coding genome of DLBCL remains largely unexplored. Here we show that active super-enhancers are highly and specifically hypermutated in 92% of samples from individuals with DLBCL, display signatures of activation-induced cytidine deaminase activity, and are linked to genes that encode B cell developmental regulators and oncogenes. As evidence of oncogenic relevance, we show that the hypermutated super-enhancers linked to the BCL6, BCL2 and CXCR4 proto-oncogenes prevent the binding and transcriptional downregulation of the corresponding target gene by transcriptional repressors, including BLIMP1 (targeting BCL6) and the steroid receptor NR3C1 (targeting BCL2 and CXCR4). Genetic correction of selected mutations restored repressor DNA binding, downregulated target gene expression and led to the counter-selection of cells containing corrected alleles, indicating an oncogenic dependency on the super-enhancer mutations. This pervasive super-enhancer mutational mechanism reveals a major set of genetic lesions deregulating gene expression, which expands the involvement of known oncogenes in DLBCL pathogenesis and identifies new deregulated gene targets of therapeutic relevance., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

In response to T-cell-dependent antigens, mature B cells in the secondary lymphoid organs are stimulated to form germinal centers (GCs), which are histological structures deputed to antibody affinity maturation, a process associated with immunoglobulin gene editing by somatic hypermutation (SHM) and class switch recombination (CSR). GC B cells are heterogeneous and transition across multiple stages before being eliminated by apoptosis or committing to post-GC differentiation as memory B cells or plasma cells. In order to explore the dynamics of SHM and CSR during the GC reaction, we identified GC subpopulations by single-cell (sc) transcriptomics and analyzed the load of immunoglobulin variable (V) region mutations as well as the isotype class distribution in each subpopulation. The results showed that the large majority of GC B cells display a quantitatively similar mutational load in the V regions and analogous IGH isotype class distribution, except for the precursors of memory B cells (PreM) and plasma cells (PBL). PreM showed a bimodal pattern with about half of the cells displaying high V region germline identity and enrichment for unswitched IGH, while the rest of the cells carried a mutational load similar to the bulk of GC B cells and showed a switched isotype. PBL displayed a bias toward expression of IGHG and higher V region germline identity compared to the bulk of GC B cells. Genes implicated in SHM and CSR were significantly induced in specific GC subpopulations, consistent with the occurrence of SHM in dark zone cells and suggesting that CSR can occur within the GC., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Corinaldesi, Holmes, Shen, Grunstein, Pasqualucci, Dalla-Favera and Basso.)

In this issue of Blood Cancer Discovery , Schaffer and colleagues uncover a novel epigenetic mechanism of resistance to the Bruton tyrosine kinase inhibitor ibrutinib in activated B-cell-like diffuse large B-cell lymphoma (ABC-DLBCL), whereby tumor cells rewire the B-cell receptor (BCR)-driven NF-κB signaling cascade through the small GTPase RAC2. This circuit can be efficiently targeted by RAC1/2 small-molecule inhibitors, suggesting a promising new therapeutic approach to overcome acquired ibrutinib resistance in ABC-DLBCL and possibly other B-cell malignancies relying on active BCR signaling. See related article by Shaffer et al., p. 630 ., (©2021 American Association for Cancer Research.)

CtIP is a DNA end resection factor widely implicated in alternative end-joining (A-EJ)-mediated translocations in cell-based reporter systems. To address the physiological role of CtIP, an essential gene, in translocation-mediated lymphomagenesis, we introduced the T855A mutation at murine CtIP to nonhomologous end-joining and Tp53 double-deficient mice that routinely succumbed to lymphomas carrying A-EJ-mediated IgH-Myc translocations. T855 of CtIP is phosphorylated by ATM or ATR kinases upon DNA damage to promote end resection. Here, we reported that the T855A mutation of CtIP compromised the neonatal development of Xrcc4 -/- Tp53 -/- mice and the IgH-Myc translocation-driven lymphomagenesis in DNA-PKcs -/- Tp53 -/- mice. Mechanistically, the T855A mutation limits DNA end resection length without affecting hairpin opening, translocation frequency, or fork stability. Meanwhile, after radiation, CtIP-T855A mutant cells showed a consistent decreased Chk1 phosphorylation and defects in the G2/M cell cycle checkpoint. Consistent with the role of T855A mutation in lymphomagenesis beyond translocation, the CtIP-T855A mutation also delays splenomegaly in λ-Myc mice. Collectively, our study revealed a role of CtIP-T855 phosphorylation in lymphomagenesis beyond A-EJ-mediated chromosomal translocation., Competing Interests: The authors declare no competing interest.

Over the last decades, the revolution in DNA sequencing has changed the way we understand the genetics and biology of B-cell lymphomas by uncovering a large number of recurrently mutated genes, whose aberrant function is likely to play an important role in the initiation and/or maintenance of these cancers. Dissecting how the involved genes contribute to the physiology and pathology of germinal center (GC) B cells -the origin of most B-cell lymphomas- will be key to advance our ability to diagnose and treat these patients. Genetically engineered mouse models (GEMM) that faithfully recapitulate lymphoma-associated genetic alterations offer a valuable platform to investigate the pathogenic roles of candidate oncogenes and tumor suppressors in vivo , and to pre-clinically develop new therapeutic principles in the context of an intact tumor immune microenvironment. In this review, we provide a summary of state-of-the art GEMMs obtained by accurately modelling the most common genetic alterations found in human GC B cell malignancies, with a focus on Burkitt lymphoma, follicular lymphoma, and diffuse large B-cell lymphoma, and we discuss how lessons learned from these models can help guide the design of novel therapeutic approaches for this disease., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Meyer, Koul and Pasqualucci.)

The transcription factor forkhead box O1 (FOXO1), which instructs the dark zone program to direct germinal center (GC) polarity, is typically inactivated by phosphatidylinositol 3-kinase (PI3K) signals. Here, we investigated how FOXO1 mutations targeting this regulatory axis in GC-derived B cell non-Hodgkin lymphomas (B-NHLs) contribute to lymphomagenesis. Examination of primary B-NHL tissues revealed that FOXO1 mutations and PI3K pathway activity were not directly correlated. Human B cell lines bearing FOXO1 mutations exhibited hyperactivation of PI3K and Stress-activated protein kinase (SAPK)/Jun amino-terminal kinase (JNK) signaling, and increased cell survival under stress conditions as a result of alterations in FOXO1 transcriptional affinities and activation of transcriptional programs characteristic of GC-positive selection. When modeled in mice, FOXO1 mutations conferred competitive advantage to B cells in response to key T-dependent immune signals, disrupting GC homeostasis. FOXO1 mutant transcriptional signatures were prevalent in human B-NHL and predicted poor clinical outcomes. Thus, rather than enforcing FOXO1 constitutive activity, FOXO1 mutations enable co-option of GC-positive selection programs during the pathogenesis of GC-derived lymphomas., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Fifty percent of diffuse large B cell lymphoma (DLBCL) cases lack cell-surface expression of the class I major histocompatibility complex (MHC-I), thus escaping recognition by cytotoxic T cells. Here we show that, across B cell lymphomas, loss of MHC-I, but not MHC-II, is preferentially restricted to DLBCL. To identify the involved mechanisms, we performed whole exome and targeted HLA deep-sequencing in 74 DLBCL samples, and found somatic inactivation of B2M and the HLA-I loci in 80% (34 of 42) of MHC-I NEG tumors. Furthermore, 70% (22 of 32) of MHC-I POS DLBCLs harbored monoallelic HLA-I genetic alterations (MHC-I POS/mono ), indicating allele-specific inactivation. MHC-I NEG and MHC-I POS/mono cases harbored significantly higher mutational burden and inferred neoantigen load, suggesting potential coselection of HLA-I loss and sustained neoantigen production. Notably, the analysis of >500,000 individuals across different cancer types revealed common germline HLA-I homozygosity, preferentially in DLBCL. In mice, germinal-center B cells lacking HLA-I expression did not progress to lymphoma and were counterselected in the context of oncogene-driven lymphomagenesis, suggesting that additional events are needed to license immune evasion. These results suggest a multistep process of HLA-I loss in DLBCL development including both germline and somatic events, and have direct implications for the pathogenesis and immunotherapeutic targeting of this disease., Competing Interests: Competing interest statement: R.R. is founder of Genotwin and a member of the scientific advisory board of AimedBio. R.D.-F. is a member of the scientific advisory board of Akemara and NeoGenomics, and a consultant of Astra Zeneca. The work reported in this paper has no relation to the current activities in these companies., (Copyright © 2021 the Author(s). Published by PNAS.)

Over the past two decades, genomic analyses of several B-cell lymphoma entities have identified a large number of genes that are recurrently mutated, suggesting that their aberrant function promotes lymphomagenesis. For many of those genes, the specific role in normal B-cell development is unknown; moreover, whether and how their deregulated activity contributes to lymphoma initiation and/or maintenance is often difficult to determine. Genetically engineered mouse models that faithfully mimic lymphoma-associated genetic alterations represent valuable tools for elucidating the pathogenic roles of candidate oncogenes and tumor suppressors in vivo, as well as for the preclinical testing of novel therapeutic principles in an intact microenvironment. Here we summarize what has been learned about the mechanisms of oncogenic transformation from accurately modeling the most common and well-characterized genetic alterations identified in mature B-cell malignancies. This information is expected to guide the design of improved molecular diagnostics and mechanism-based therapeutic approaches for these diseases., (Copyright © 2021 Cold Spring Harbor Laboratory Press; all rights reserved.)

Based on gene expression profiles, diffuse large B cell lymphoma (DLBCL) is sub-divided into germinal center B cell-like (GCB) and activated B cell-like (ABC) DLBCL. Two of the most common genomic aberrations in ABC-DLBCL are mutations in MYD88 , as well as BCL2 copy number gains. Here, we employ immune phenotyping, RNA-Seq and whole exome sequencing to characterize a Myd88 and Bcl2 -driven mouse model of ABC-DLBCL. We show that this model resembles features of human ABC-DLBCL. We further demonstrate an actionable dependence of our murine ABC-DLBCL model on BCL2. This BCL2 dependence was also detectable in human ABC-DLBCL cell lines. Moreover, human ABC-DLBCLs displayed increased PD-L1 expression, compared to GCB-DLBCL. In vivo experiments in our ABC-DLBCL model showed that combined venetoclax and RMP1-14 significantly increased the overall survival of lymphoma bearing animals, indicating that this combination may be a viable option for selected human ABC-DLBCL cases harboring MYD88 and BCL2 aberrations.

Follicular lymphoma (FL) is an indolent yet challenging disease. Despite a generally favorable response to immunochemotherapy regimens, a fraction of patients does not respond or relapses early with unfavorable prognosis. For the vast majority of those who initially respond, relapses will repeatedly occur with increasing refractoriness to available treatments. Addressing the clinical challenges in FL warrants deep understanding of the nature of treatment-resistant FL cells seeding relapses, and of the biological basis of early disease progression. Great progress has been made in the last decade in the description and interrogation of the (epi)genomic landscape of FL cells, of their major dependency to the tumor microenvironment (TME), and of the stepwise lymphomagenesis process, from healthy to subclinical disease and to overt FL. A new picture is emerging, in which an ever-evolving tumor-TME duo sparks a complex and multilayered clonal and functional heterogeneity, blurring the discovery of prognostic biomarkers, patient stratification and reliable designs of risk-adapted treatments. Novel technological approaches allowing to decipher both tumor and TME heterogeneity at the single-cell level are beginning to unravel unsuspected cell dynamics and plasticity of FL cells. The upcoming drawing of a comprehensive functional picture of FL within its ecosystem holds great promise to address the unmet medical needs of this complex lymphoma., (Copyright © 2021 Elsevier Inc. All rights reserved.)

In response to T cell-dependent antigens, mature B cells are stimulated to form germinal centers (GCs), the sites of B cell affinity maturation and the cell of origin (COO) of most B cell lymphomas. To explore the dynamics of GC B cell development beyond the known dark zone and light zone compartments, we performed single-cell (sc) transcriptomic analysis on human GC B cells and identified multiple functionally linked subpopulations, including the distinct precursors of memory B cells and plasma cells. The gene expression signatures associated with these GC subpopulations were effective in providing a sc-COO for ∼80% of diffuse large B cell lymphomas (DLBCLs) and identified novel prognostic subgroups of DLBCL., Competing Interests: Disclosures: N. Compagno is currently employed at Novartis. No other disclosures were reported., (© 2020 Holmes et al.)

Plasmablastic lymphoma (PBL) is an aggressive B-cell non-Hodgkin lymphoma associated with immunodeficiency in the context of Human Immunodeficiency Virus (HIV) infection or iatrogenic immunosuppression. While a rare disease in general, the incidence is dramatically increased in regions of the world with high HIV prevalence. The molecular pathogenesis of this disease is poorly characterized. Here, we defined the genomic features of PBL in a cohort of 110 patients from South Africa (15 by whole exome sequencing and 95 by deep targeted sequencing). We identified recurrent mutations in genes of the JAK-STAT signaling pathway, including STAT3 (42%), JAK1 (14%) and SOCS1 (10%), leading to its constitutive activation. Moreover, 24% of cases harbored gain-of-function mutations in RAS family members ( NRAS and KRAS ). Comparative analysis with other B-cell malignancies uncovered PBL-specific somatic mutations and transcriptional programs. We also found recurrent copy number gains encompassing the CD44 gene (37%), which encodes for a cell surface receptor involved in lymphocyte activation and homing, and was found expressed at high levels in all tested cases, independent of genetic alterations. These findings have implications for the understanding of the pathogenesis of this disease and the development of personalized medicine approaches., Competing Interests: A conflict of interest disclosure statement: R.R. is a consultant for AimedBio in a project unrelated to the current manuscript.

Although mutations in the gene encoding the RNA splicing factor SF3B1 are frequent in multiple cancers, their functional effects and therapeutic dependencies are poorly understood. Here, we characterize 98 tumors and 12 isogenic cell lines harboring SF3B1 hotspot mutations, identifying hundreds of cryptic 3' splice sites common and specific to different cancer types. Regulatory network analysis revealed that the most common SF3B1 mutation activates MYC via effects conserved across human and mouse cells. SF3B1 mutations promote decay of transcripts encoding the protein phosphatase 2A (PP2A) subunit PPP2R5A, increasing MYC S62 and BCL2 S70 phosphorylation which, in turn, promotes MYC protein stability and impair apoptosis, respectively. Genetic PPP2R5A restoration or pharmacologic PP2A activation impaired SF3B1-mutant tumorigenesis, elucidating a therapeutic approach to aberrant splicing by mutant SF3B1. SIGNIFICANCE: Here, we identify that mutations in SF3B1 , the most commonly mutated splicing factor gene across cancers, alter splicing of a specific subunit of the PP2A serine/threonine phosphatase complex to confer post-translational MYC and BCL2 activation, which is therapeutically intervenable using an FDA-approved drug. See related commentary by O'Connor and Narla, p. 765 . This article is highlighted in the In This Issue feature, p. 747 ., (©2020 American Association for Cancer Research.)

Inactivating mutations of the CREBBP and EP300 acetyltransferases are among the most common genetic alterations in diffuse large B cell lymphoma (DLBCL) and follicular lymphoma (FL). Here, we examined the relationship between these two enzymes in germinal center (GC) B cells, the normal counterpart of FL and DLBCL, and in lymphomagenesis by using conditional GC-directed deletion mouse models targeting Crebbp or Ep300. We found that CREBBP and EP300 modulate common as well as distinct transcriptional programs implicated in separate anatomic and functional GC compartments. Consistently, deletion of Ep300 but not Crebbp impaired the fitness of GC B cells in vivo. Combined loss of Crebbp and Ep300 completely abrogated GC formation, suggesting that these proteins partially compensate for each other through common transcriptional targets. This synthetic lethal interaction was retained in CREBBP-mutant DLBCL cells and could be pharmacologically targeted with selective small molecule inhibitors of CREBBP and EP300 function. These data provide proof-of-principle for the clinical development of EP300-specific inhibitors in FL and DLBCL., (Copyright © 2019 Elsevier Inc. All rights reserved.)

To repurpose compounds for diffuse large B cell lymphoma (DLBCL), we screened a library of drugs and other targeted compounds approved by the US Food and Drug Administration on 9 cell lines and validated the results on a panel of 32 genetically characterized DLBCL cell lines. Dasatinib, a multikinase inhibitor, was effective against 50% of DLBCL cell lines, as well as against in vivo xenografts. Dasatinib was more broadly active than the Bruton kinase inhibitor ibrutinib and overcame ibrutinib resistance. Tumors exhibiting dasatinib resistance were commonly characterized by activation of the PI3K pathway and loss of PTEN expression as a specific biomarker. PI3K suppression by mTORC2 inhibition synergized with dasatinib and abolished resistance in vitro and in vivo. These results provide a proof of concept for the repurposing approach in DLBCL, and point to dasatinib as an attractive strategy for further clinical development in lymphomas., Competing Interests: Conflict of interest statement: G.I. and reviewer Margaret Shipp are coauthors on a 2016 paper.

Despite considerable advances in the treatment of lymphoma, the prognosis of patients with relapsed and/or refractory disease continues to be poor; thus, a continued need exists for the development of novel approaches and therapies. Epigenetic dysregulation might drive and/or promote tumorigenesis in various types of malignancies and is prevalent in both B cell and T cell lymphomas. Over the past decade, a large number of epigenetic-modifying agents have been developed and introduced into the clinical management of patients with haematological malignancies. In this Review, we provide a concise overview of the most promising epigenetic therapies for the treatment of lymphomas, including inhibitors of histone deacetylases (HDACs), DNA methyltransferases (DNMTs), enhancer of zeste homologue 2 (EZH2), bromodomain and extra-terminal domain proteins (BETs), protein arginine N-methyltransferases (PRMTs) and isocitrate dehydrogenases (IDHs), and highlight the most promising future directions of research in this area.

B cell lymphomas comprise a heterogeneous group of genetically, biologically, and clinically distinct neoplasms that, in most cases, originate from the clonal expansion of B cells in the germinal center (GC). In recent years, the advent of novel genomics technologies has revolutionized our understanding of the molecular pathogenesis of lymphoid malignancies as a multistep process that requires the progressive accumulation of multiple genetic and epigenetic alterations. A common theme that emerged from these studies is the ability of lymphoma cells to co-opt the same biological programs and signal transduction networks that operate during the normal GC reaction, and misuse them for their own survival advantage. This review summarizes recent progress in the understanding of the genetic and epigenetic mechanisms that drive the malignant transformation of GC B cells. These insights provide a conceptual framework for the identification of cellular pathways that may be explored for precision medicine approaches., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Kabuki syndrome is a rare autosomal dominant condition characterized by facial features, various organs malformations, postnatal growth deficiency and intellectual disability. The discovery of frequent germline mutations in the histone methyltransferase KMT2D and the demethylase KDM6A revealed a causative role for histone modifiers in this disease. However, the role of missense mutations has remained unexplored. Here, we expanded the mutation spectrum of KMT2D and KDM6A in KS by identifying 37 new KMT2D sequence variants. Moreover, we functionally dissected 14 KMT2D missense variants, by investigating their impact on the protein enzymatic activity and the binding to members of the WRAD complex. We demonstrate impaired H3K4 methyltransferase activity in 9 of the 14 mutant alleles and show that this reduced activity is due in part to disruption of protein complex formation. These findings have relevant implications for diagnostic and counseling purposes in this disease.

The gene encoding the MEF2B transcription factor is mutated in germinal center (GC)-derived B cell lymphomas, but its role in GC development and lymphomagenesis is unknown. We demonstrate that Mef2b deletion reduces GC formation in mice and identify MEF2B transcriptional targets in GC, with roles in cell proliferation, apoptosis, GC confinement, and differentiation. The most common lymphoma-associated MEF2B mutant (MEF2B D83V ) is hypomorphic, yet escapes binding and negative regulation by components of the HUCA complex and class IIa HDACs. Mef2b D83V expression in mice leads to GC enlargement and lymphoma development, a phenotype that becomes fully penetrant in combination with BCL2 de-regulation, an event associated with human MEF2B mutations. These results identify MEF2B as a critical GC regulator and a driver oncogene in lymphomagenesis., (Copyright © 2018 Elsevier Inc. All rights reserved.)

One cause of cancer mortality is tumor evolution to therapy-resistant disease. First line therapy often targets the dominant clone, and drug resistance can emerge from preexisting clones that gain fitness through therapy-induced natural selection. Such mutations may be identified using targeted sequencing assays by analysis of noise in high-depth data. Here, we develop a comprehensive, unbiased model for sequencing error background. We find that noise in sufficiently deep DNA sequencing data can be approximated by aggregating negative binomial distributions. Mutations with frequencies above noise may have prognostic value. We evaluate our model with simulated exponentially expanded populations as well as data from cell line and patient sample dilution experiments, demonstrating its utility in prognosticating tumor progression. Our results may have the potential to identify significant mutations that can cause recurrence. These results are relevant in the pre-treatment clinical setting to determine appropriate therapy and prepare for potential recurrence pretreatment.

Dissecting the pathogenesis of classical Hodgkin lymphoma (cHL), a common cancer in young adults, remains challenging because of the rarity of tumor cells in involved tissues (usually <5%). Here, we analyzed the coding genome of cHL by microdissecting tumor and normal cells from 34 patient biopsies for a total of ∼50 000 singly isolated lymphoma cells. We uncovered several recurrently mutated genes, namely, STAT6 (32% of cases), GNA13 (24%), XPO1 (18%), and ITPKB (16%), and document the functional role of mutant STAT6 in sustaining tumor cell viability. Mutations of STAT6 genetically and functionally cooperated with disruption of SOCS1 , a JAK-STAT pathway inhibitor, to promote cHL growth. Overall, 87% of cases showed dysregulation of the JAK-STAT pathway by genetic alterations in multiple genes (also including STAT3 , STAT5B , JAK1 , JAK2 , and PTPN1 ), attesting to the pivotal role of this pathway in cHL pathogenesis and highlighting its potential as a new therapeutic target in this disease., (© 2018 by The American Society of Hematology.)

Diffuse large B-cell lymphoma (DLBCL), the most frequent subtype of lymphoid malignancy, remains a significant clinical challenge, as ∼30% of patients are not cured. Over the past decade, remarkable progress has been made in the understanding of the pathogenesis of this disease, spurred by the implementation of powerful genomic technologies that enabled the definition of its genetic and epigenetic landscape. These studies have uncovered a multitude of genomic alterations that contribute to the initiation and maintenance of the tumor clone by disrupting biological functions known to be critical for the normal biology of its cells of origin, germinal center B cells. The identified alterations involve epigenetic remodeling, block of differentiation, escape from immune surveillance, and the constitutive activation of several signal transduction pathways. This wealth of new information offers unique opportunities for the development of improved diagnostic and prognostic tools that could help guide the clinical management of DLBCL patients. Furthermore, a number of the mutated genes identified are potentially actionable targets that are currently being explored for the development of novel therapeutic strategies. This review summarizes current knowledge of the most common genetic alterations associated with DLBCL in relation to their functional impact on the malignant transformation process, and discusses their clinical implications for mechanism-based therapeutics., (© 2018 by The American Society of Hematology.)

The 13th International Conference on Malignant Lymphoma held in Lugano in June 2015 was preceded by a closed workshop (organized in collaboration with the American Association for Cancer Research and the European School of Oncology) with the aim of developing an up-to-date understanding of the biology of follicular lymphoma and the clinical implications of new findings in the field. Discussed topics included the mutational spectrum at diagnosis, the clinical correlates of genetic and epigenetic alterations, the mechanisms of clonal evolution and histological transformation, the cross talk between tumor cells and microenvironment, and the development of novel treatments. This report represents a summary of the workshop., (Copyright © 2017 John Wiley & Sons, Ltd.)

Similar to resting mature B cells, where the B-cell antigen receptor (BCR) controls cellular survival, surface BCR expression is conserved in most mature B-cell lymphomas. The identification of activating BCR mutations and the growth disadvantage upon BCR knockdown of cells of certain lymphoma entities has led to the view that BCR signalling is required for tumour cell survival. Consequently, the BCR signalling machinery has become an established target in the therapy of B-cell malignancies. Here we study the effects of BCR ablation on MYC-driven mouse B-cell lymphomas and compare them with observations in human Burkitt lymphoma. Whereas BCR ablation does not, per se, significantly affect lymphoma growth, BCR-negative (BCR - ) tumour cells rapidly disappear in the presence of their BCR-expressing (BCR + ) counterparts in vitro and in vivo. This requires neither cellular contact nor factors released by BCR + tumour cells. Instead, BCR loss induces the rewiring of central carbon metabolism, increasing the sensitivity of receptor-less lymphoma cells to nutrient restriction. The BCR attenuates glycogen synthase kinase 3 beta (GSK3β) activity to support MYC-controlled gene expression. BCR - tumour cells exhibit increased GSK3β activity and are rescued from their competitive growth disadvantage by GSK3β inhibition. BCR - lymphoma variants that restore competitive fitness normalize GSK3β activity after constitutive activation of the MAPK pathway, commonly through Ras mutations. Similarly, in Burkitt lymphoma, activating RAS mutations may propagate immunoglobulin-crippled tumour cells, which usually represent a minority of the tumour bulk. Thus, while BCR expression enhances lymphoma cell fitness, BCR-targeted therapies may profit from combinations with drugs targeting BCR - tumour cells.

PRDM1/BLIMP-1, a master regulator of plasma-cell differentiation, is frequently inactivated in activated B-cell-like (ABC) diffuse large B-cell lymphoma (DLBCL) patients. Little is known about its genetic aberrations and relevant clinical implications. A large series of patients with de novo DLBCL was effectively evaluated for PRDM1/BLIMP-1 deletion, mutation, and protein expression. BLIMP-1 expression was frequently associated with the ABC phenotype and plasmablastic morphologic subtype of DLBCL, yet 63% of the ABC-DLBCL patients were negative for BLIMP-1 protein expression. In these patients, loss of BLIMP-1 was associated with Myc overexpression and decreased expression of p53 pathway molecules. In addition, homozygous PRDM1 deletions and PRDM1 mutations within exons 1 and 2, which encode for domains crucial for transcriptional repression, were found to show a poor prognostic impact in patients with ABC-DLBCL but not in those with germinal center B-cell-like DLBCL (GCB-DLBCL). Gene expression profiling revealed that loss of PRDM1/BLIMP-1 expression correlated with a decreased plasma-cell differentiation signature and upregulation of genes involved in B-cell receptor signaling and tumor-cell proliferation. In conclusion, these results provide novel clinical and biological insight into the tumor-suppressive role of PRDM1/BLIMP-1 in ABC-DLBCL patients and suggest that loss of PRDM1/BLIMP-1 function contributes to the overall poor prognosis of ABC-DLBCL patients.

Inactivating mutations of the CREBBP acetyltransferase are highly frequent in diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL), the two most common germinal center (GC)-derived cancers. However, the role of CREBBP inactivation in lymphomagenesis remains unclear. Here, we show that CREBBP regulates enhancer/super-enhancer networks with central roles in GC/post-GC cell fate decisions, including genes involved in signal transduction by the B-cell receptor and CD40 receptor, transcriptional control of GC and plasma cell development, and antigen presentation. Consistently, Crebbp -deficient B cells exhibit enhanced response to mitogenic stimuli and perturbed plasma cell differentiation. Although GC-specific loss of Crebbp was insufficient to initiate malignant transformation, compound Crebbp -haploinsufficient/BCL2-transgenic mice, mimicking the genetics of FL and DLBCL, develop clonal lymphomas recapitulating the features of the human diseases. These findings establish CREBBP as a haploinsufficient tumor-suppressor gene in GC B cells and provide insights into the mechanisms by which its loss contributes to lymphomagenesis. Significance: Loss-of-function mutations of CREBBP are common and early lesions in FL and DLBCL, suggesting a prominent role in lymphoma initiation. Our studies identify the cellular program by which reduced CREBBP dosage facilitates malignant transformation, and have direct implications for targeted lymphoma therapy based on drugs affecting CREBBP-mediated chromatin acetylation. Cancer Discov; 7(3); 322-37. ©2017 AACR. This article is highlighted in the In This Issue feature, p. 235 ., (©2017 American Association for Cancer Research.)

Nodal marginal zone lymphoma (NMZL) is a rare, indolent B-cell tumor that is distinguished from splenic marginal zone lymphoma (SMZL) by the different pattern of dissemination. NMZL still lacks distinct markers and remains orphan of specific cancer gene lesions. By combining whole-exome sequencing, targeted sequencing of tumor-related genes, whole-transcriptome sequencing, and high-resolution single nucleotide polymorphism array analysis, we aimed at disclosing the pathways that are molecularly deregulated in NMZL and we compare the molecular profile of NMZL with that of SMZL. These analyses identified a distinctive pattern of nonsilent somatic lesions in NMZL. In 35 NMZL patients, 41 genes were found recurrently affected in ≥3 (9%) cases, including highly prevalent molecular lesions of MLL2 (also known as KMT2D; 34%), PTPRD (20%), NOTCH2 (20%), and KLF2 (17%). Mutations of PTPRD, a receptor-type protein tyrosine phosphatase regulating cell growth, were enriched in NMZL across mature B-cell tumors, functionally caused the loss of the phosphatase activity of PTPRD, and were associated with cell-cycle transcriptional program deregulation and increased proliferation index in NMZL. Although NMZL shared with SMZL a common mutation profile, NMZL harbored PTPRD lesions that were otherwise absent in SMZL. Collectively, these findings provide new insights into the genetics of NMZL, identify PTPRD lesions as a novel marker for this lymphoma across mature B-cell tumors, and support the distinction of NMZL as an independent clinicopathologic entity within the current lymphoma classification., (© 2016 by The American Society of Hematology.)

Diffuse large B cell lymphoma (DLBCL) is the most common form of B cell non-Hodgkin lymphoma worldwide and comprises a heterogeneous group of malignancies that originate from the malignant transformation of germinal center (GC) B cells. Over the past decade, significant improvement has been achieved in our understanding of the molecular pathogenesis underlying this disease, thanks in part to the implementation of powerful genomic technologies allowing genome-wide structural and functional analyses. These studies revealed the presence of multiple oncogenic alterations dysregulating signal transduction pathways that are normally required for the normal biology of the cells from which these tumors are derived. Among the pathways identified as recurrent targets of genetic lesions in DLBCL, NF-κB has emerged as a central player in the development and maintenance of this disease, particularly in the less curable, activated B cell (ABC)- like subtype. These lesions reveal vulnerabilities of the lymphoma cells that can be exploited for the design of more rationale therapeutic approaches. The purpose of this review is to summarize recent progresses in understanding the role of NF-κB deregulation in the pathogenesis of DLBCL, with emphasis on the genetic basis underlying its aberrant activation, in relationship to the normal biology of B lymphocytes, and the modelling of these lesions in the mouse., (Copyright © 2016. Published by Elsevier Ltd.)

Diffuse large B-cell lymphoma (DLBCL) is one of the most common and aggressive types of B-cell lymphoma. Deregulation of proto-oncogene expression after a translocation, most notably to the immunoglobulin heavy-chain locus (IGH), is one of the hallmarks of DLBCL. Using whole-genome sequencing analysis, we have identified the PD-L1/PD-L2 locus as a recurrent translocation partner for IGH in DLBCL. PIM1 and TP63 were also identified as novel translocation partners for PD-L1/PD-L2 Fluorescence in situ hybridization was furthermore used to rapidly screen an expanded DLBCL cohort. Collectively, a subset of samples was found to be affected by gains (12%), amplifications (3%), and translocations (4%) of the PD-L1/PD-L2 locus. RNA sequencing data coupled with immunohistochemistry revealed that these cytogenetic alterations correlated with increased expression of PD-L1 but not of PD-L2 Moreover, cytogenetic alterations affecting the PD-L1/PD-L2 locus were more frequently observed in the non-germinal center B cell-like (non-GCB) subtype of DLBCL. These findings demonstrate the genetic basis of PD-L1 overexpression in DLBCL and suggest that treatments targeting the PD-1-PD-L1/PD-L2 axis might benefit DLBCL patients, especially those belonging to the more aggressive non-GCB subtype., (© 2016 by The American Society of Hematology.)

Mutations in the gene encoding the KMT2D (or MLL2) methyltransferase are highly recurrent and occur early during tumorigenesis in diffuse large B cell lymphoma (DLBCL) and follicular lymphoma (FL). However, the functional consequences of these mutations and their role in lymphomagenesis are unknown. Here we show that FL- and DLBCL-associated KMT2D mutations impair KMT2D enzymatic activity, leading to diminished global H3K4 methylation in germinal-center (GC) B cells and DLBCL cells. Conditional deletion of Kmt2d early during B cell development, but not after initiation of the GC reaction, results in an increase in GC B cells and enhances B cell proliferation in mice. Moreover, genetic ablation of Kmt2d in mice overexpressing Bcl2 increases the incidence of GC-derived lymphomas resembling human tumors. These findings suggest that KMT2D acts as a tumor suppressor gene whose early loss facilitates lymphomagenesis by remodeling the epigenetic landscape of the cancer precursor cells. Eradication of KMT2D-deficient cells may thus represent a rational therapeutic approach for targeting early tumorigenic events.

Diffuse large B cell lymphoma (DLBCL) is a complex disease comprising diverse subtypes and genetic profiles. Possibly because of the prevalence of genetic alterations activating canonical NF-κB activity, a role for oncogenic lesions that activate the alternative NF-κB pathway in DLBCL has remained elusive. Here, we show that deletion/mutation of TRAF3, a negative regulator of the alternative NF-κB pathway, occurs in ∼15% of DLBCLs and that it often coexists with BCL6 translocation, which prevents terminal B cell differentiation. Accordingly, in a mouse model constitutive activation of the alternative NF-κB pathway cooperates with BCL6 deregulation in DLBCL development. This work demonstrates a key oncogenic role for the alternative NF-κB pathway in DLBCL development., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Diffuse large B-cell lymphoma (DLBCL), the most common lymphoid malignancy in the western world, is an aggressive disease that remains incurable in approximately 30% of patients. Over the past decade, the rapid expansion of sequencing technologies allowing the genome-wide assessment of genomic and transcriptional changes has revolutionized our understanding of the genetic basis of DLBCL by providing a comprehensive and unbiased view of the genes/pathways that are disrupted by genetic alterations in this disease, and may contribute to tumor initiation and expansion. These studies uncovered the existence of several previously unappreciated alterations in key cellular pathways that may also influence treatment outcome. Indeed, a number of newly identified genetic lesions are currently being explored as markers for improved diagnosis and risk stratification, or are entering clinical trials as promising therapeutic targets. This review focuses on recent advances in the genomic characterization of DLBCL and discusses how information gained from these efforts has provided new insights into its biology, uncovering potential targets of prognostic and therapeutic relevance., (Copyright © 2015 Elsevier Inc. All rights reserved.)

BCL2 is a target of somatic hypermutation in t(14;18) positive and also in a small fraction of t(14;18) negative diffuse large B-cell lymphoma (DLBCL), suggesting an aberrant role of somatic hypermutation (ASHM). To elucidate the prevalence of BCL2 mutations in lymphomas other than DLBCL, we Sanger-sequenced the hypermutable region of the BCL2 gene in a panel of 69 mature B-cell lymphomas, including Richter's syndrome DLBCL, marginal-zone lymphomas, post-transplant lymphoproliferative disorders, HIV-associated and common-variable immunodeficiency-associated DLBCL, all known to harbour ASHM-dependent mutations in other genes, as well as 16 t(14,18) negative and 21 t(14;18) positive follicular lymphomas (FLs). We also investigated the pattern of BCL2 mutations in longitudinal samples from 10 FL patients relapsing to FL or transforming to DLBCL (tFL). By direct sequencing, we found clonally represented BCL2 mutations in 2/16 (13%) of t(14;18) negative FLs, 2/16 (13%) HIV-DLBCLs, 1/9 (11%) of Richter's syndrome DLBCL, 1/17 (6%) of post-transplant lymphoproliferative disorders and 1/2 (50%) common-variable immunodeficiency-associated DLBCL. The proportion of mutated cases was significantly lower than in FLs carrying the t(14;18) translocation (15/21, 71%). However, the absence of t(14;18) by FISH or PCR and the molecular features of the mutations strongly suggest that BCL2 represents an additional target of ASHM in these entities. Analysis of the BCL2 mutation pattern in clonally related FL/FL and FL/tFL samples revealed two distinct scenarios of genomic evolution: (i) direct evolution from the antecedent FL clone, with few novel clonal mutations acquired by the tFL major clone, and (ii) evolution from a common mutated long-lived progenitor cell, which subsequently acquired distinct mutations in the FL and in the relapsed or transformed counterpart., (Copyright © 2014 John Wiley & Sons, Ltd.)

A balance between quiescence and proliferation is critical for proper maintenance of the hematopoietic stem cell (HSC) pool. Although a lot is known about hematopoiesis, molecular mechanisms that control HSC quiescence remain largely unknown. The ubiquitin-editing enzyme A20 functions as a central regulator of inflammation and adaptive immunity. Here, we show that a deficiency of A20 in the hematopoietic system causes anemia, lymphopenia, and postnatal lethality. Lack of A20 in HSCs results in diminished pool size, impaired radioprotection, defective repopulation, and loss of quiescence. A20-deficient HSCs display increased IFN-γ signaling, caused by augmented NF-κB activation. Strikingly, deletion of both IFN-γ and A20 in hematopoietic cells results in partial rescue of the HSC phenotype. We anticipate that our experiments will facilitate the understanding of mechanisms through which A20-mediated inflammatory signals control HSC quiescence and functions., (© 2015 Nakagawa et al.)

Cancer is a clonal evolutionary process, caused by successive accumulation of genetic alterations providing milestones of tumor initiation, progression, dissemination, and/or resistance to certain therapeutic regimes. To unravel these milestones we propose a framework, tumor evolutionary directed graphs (TEDG), which is able to characterize the history of genetic alterations by integrating longitudinal and cross-sectional genomic data. We applied TEDG to a chronic lymphocytic leukemia (CLL) cohort of 70 patients spanning 12 years and show that: (a) the evolution of CLL follows a time-ordered process represented as a global flow in TEDG that proceeds from initiating events to late events; (b) there are two distinct and mutually exclusive evolutionary paths of CLL evolution; (c) higher fitness clones are present in later stages of the disease, indicating a progressive clonal replacement with more aggressive clones. Our results suggest that TEDG may constitute an effective framework to recapitulate the evolutionary history of tumors.

Fludarabine refractoriness (FR) represents an unsolved clinical problem of chronic lymphocytic leukemia (CLL) management. Although next-generation sequencing studies have led to the identification of a number of genes frequently mutated in FR-CLL, a comprehensive evaluation of the FR-CLL genome has not been reported. Toward this end, we studied 10 FR-CLLs by combining whole-exome sequencing and copy number aberration (CNA) analysis, which showed an average of 16.3 somatic mutations and 4 CNAs per sample. Screening of recurrently mutated genes in 48 additional FR-CLLs revealed that ~70% of FR-CLLs carry ≥1 mutation in genes previously associated with CLL clinical course, including TP53 (27.5%), NOTCH1 (24.1%), SF3B1 (18.9%), and BIRC3 (15.5%). In addition, this analysis showed that 10.3% of FR-CLL cases display mutations of the FAT1 gene, which encodes for a cadherin-like protein that negatively regulates Wnt signaling, consistent with a tumor suppressor role. The frequency of FAT1-mutated cases was significantly higher in FR-CLL than in unselected CLLs at diagnosis (10.3% vs 1.1%, P = .004), suggesting a role in the development of a high-risk phenotype. These findings have general implications for the mechanisms leading to FR and point to Wnt signaling as a potential therapeutic target in FR-CLL.

Follicular lymphoma (FL) is an indolent disease, but 30%-40% of cases undergo histologic transformation to an aggressive malignancy, typically represented by diffuse large B cell lymphoma (DLBCL). The pathogenesis of this process remains largely unknown. Using whole-exome sequencing and copy-number analysis, we show here that the dominant clone of FL and transformed FL (tFL) arise by divergent evolution from a common mutated precursor through the acquisition of distinct genetic events. Mutations in epigenetic modifiers and antiapoptotic genes are introduced early in the common precursor, whereas tFL is specifically associated with alterations deregulating cell-cycle progression and DNA damage responses (CDKN2A/B, MYC, and TP53) as well as aberrant somatic hypermutation. The genomic profile of tFL shares similarities with that of germinal center B cell-type de novo DLBCL but also displays unique combinations of altered genes with diagnostic and therapeutic implications., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Richter syndrome (RS) derives from the rare transformation of chronic lymphocytic leukemia (CLL) into an aggressive lymphoma, most commonly of the diffuse large B cell lymphoma (DLBCL) type. The molecular pathogenesis of RS is only partially understood. By combining whole-exome sequencing and copy-number analysis of 9 CLL-RS pairs and of an extended panel of 43 RS cases, we show that this aggressive disease typically arises from the predominant CLL clone by acquiring an average of ∼20 genetic lesions/case. RS lesions are heterogeneous in terms of load and spectrum among patients, and include those involved in CLL progression and chemorefractoriness (TP53 disruption and NOTCH1 activation) as well as some not previously implicated in CLL or RS pathogenesis. In particular, disruption of the CDKN2A/B cell cycle regulator is associated with ∼30% of RS cases. Finally, we report that the genomic landscape of RS is significantly different from that of de novo DLBCL, suggesting that they represent distinct disease entities. These results provide insights into RS pathogenesis, and identify dysregulated pathways of potential diagnostic and therapeutic relevance.