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Burkitt lymphoma (BL) is an aggressive, MYC-driven lymphoma comprising 3 distinct clinical subtypes: sporadic BLs that occur worldwide, endemic BLs that occur predominantly in sub-Saharan Africa, and immunodeficiency-associated BLs that occur primarily in the setting of HIV. In this study, we comprehensively delineated the genomic basis of BL through whole-genome sequencing (WGS) of 101 tumors representing all 3 subtypes of BL to identify 72 driver genes. These data were additionally informed by CRISPR screens in BL cell lines to functionally annotate the role of oncogenic drivers. Nearly every driver gene was found to have both coding and non-coding mutations, highlighting the importance of WGS for identifying driver events. Our data implicate coding and non-coding mutations in IGLL5, BACH2, SIN3A, and DNMT1. Epstein-Barr virus (EBV) infection was associated with higher mutation load, with type 1 EBV showing a higher mutational burden than type 2 EBV. Although sporadic and immunodeficiency-associated BLs had similar genetic profiles, endemic BLs manifested more frequent mutations in BCL7A and BCL6 and fewer genetic alterations in DNMT1, SNTB2, and CTCF. Silencing mutations in ID3 were a common feature of all 3 subtypes of BL. In vitro, mass spectrometry–based proteomics demonstrated that the ID3 protein binds primarily to TCF3 and TCF4. In vivo knockout of ID3 potentiated the effects of MYC, leading to rapid tumorigenesis and tumor phenotypes consistent with those observed in the human disease.

Introduction:Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) is a rare B-cell malignancy with a paucity of malignant cells embedded in a diverse tumor microenvironment (TME). The histological composition of the TME is known to influence outcomes, with nodular B-cell rich TME (classical histology, Fan patterns A&B) associated with indolent clinical course, while T-cell infiltration or diffuse growth (variant histology, Fan patterns C-F) may resemble aggressive T/cell-histiocyte rich large B-cell lymphoma (THRLBCL) to which NLPHL can transform. Molecular features of NLPHL and THRLBCL remain to be discovered. To gain further insights into the biology of these lymphomas, we recruited NLPHL and THRLBCL cases as part of the Atlas of Blood Cancer Genomes (ABCG) initiative, a consortium consisting of 26 institutions.

Introduction:

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

GNA13 is the most frequently mutated gene in germinal center (GC)-derived B-cell lymphomas, including nearly a quarter of Burkitt lymphoma and GC-derived diffuse large B-cell lymphoma. These mutations occur in a pattern consistent with loss of function. We have modeled the GNA13-deficient state exclusively in GC B cells by crossing the Gna13 conditional knockout mouse strain with the GC-specific AID-Cre transgenic strain. AID-Cre+ GNA13-deficient mice demonstrate disordered GC architecture and dark zone/light zone distribution in vivo, and demonstrate altered migration behavior, decreased levels of filamentous actin, and attenuated RhoA activity in vitro. We also found that GNA13-deficient mice have increased numbers of GC B cells that display impaired caspase-mediated cell death and increased frequency of somatic hypermutation in the immunoglobulin VH locus. Lastly, GNA13 deficiency, combined with conditional MYC transgene expression in mouse GC B cells, promotes lymphomagenesis. Thus, GNA13 loss is associated with GC B-cell persistence, in which impaired apoptosis and ongoing somatic hypermutation may lead to an increased risk of lymphoma development.

In this study, we define the genetic landscape of mantle cell lymphoma (MCL) through exome sequencing of 56 cases of MCL. We identified recurrent mutations in ATM, CCND1, MLL2, and TP53. We further identified a number of novel genes recurrently mutated in patients with MCL including RB1, WHSC1, POT1, and SMARCA4. We noted that MCLs have a distinct mutational profile compared with lymphomas from other B-cell stages. The ENCODE project has defined the chromatin structure of many cell types. However, a similar characterization of primary human mature B cells has been lacking. We defined, for the first time, the chromatin structure of primary human naïve, germinal center, and memory B cells through chromatin immunoprecipitation and sequencing for H3K4me1, H3K4me3, H3Ac, H3K36me3, H3K27me3, and PolII. We found that somatic mutations that occur more frequently in either MCLs or Burkitt lymphomas were associated with open chromatin in their respective B cells of origin, naïve B cells, and germinal center B cells. Our work thus elucidates the landscape of gene-coding mutations in MCL and the critical interplay between epigenetic alterations associated with B-cell differentiation and the acquisition of somatic mutations in cancer.

Introduction

Koff: Burroughs Wellcome Fund: Research Funding; V Foundation: Research Funding; Lymphoma Research Foundation: Research Funding; American Association for Cancer Research: Research Funding. Leppä:Roche: Honoraria, Research Funding; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bayer: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen-Cilag: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees. Gang:ROCHE: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Hsi:Abbvie: Research Funding; Eli Lilly: Research Funding; Cleveland Clinic&Abbvie Biotherapeutics Inc: Patents & Royalties: US8,603,477 B2; Jazz: Consultancy. Flowers:AbbVie: Consultancy, Research Funding; Denovo Biopharma: Consultancy; BeiGene: Consultancy, Research Funding; Burroughs Wellcome Fund: Research Funding; Eastern Cooperative Oncology Group: Research Funding; National Cancer Institute: Research Funding; V Foundation: Research Funding; Optimum Rx: Consultancy; Millenium/Takeda: Research Funding; TG Therapeutics: Research Funding; Gilead: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Karyopharm: Consultancy; AstraZeneca: Consultancy; Pharmacyclics/Janssen: Consultancy, Research Funding; Spectrum: Consultancy; Bayer: Consultancy; Acerta: Research Funding; Genentech, Inc./F. Hoffmann-La Roche Ltd: Consultancy, Research Funding. Neff:Enzyvant: Consultancy; EUSA Pharma: Honoraria, Membership on an entity's Board of Directors or advisory committees. Fedoriw:Alexion Pharmaceuticals: Other: Consultant and Speaker. Reddy:Genentech: Research Funding; BMS: Consultancy, Research Funding; Celgene: Consultancy; KITE Pharma: Consultancy; Abbvie: Consultancy. Mason:Sysmex: Honoraria. Behdad:Loxo-Bayer: Membership on an entity's Board of Directors or advisory committees; Thermo Fisher: Membership on an entity's Board of Directors or advisory committees; Pfizer: Other: Speaker. Burton:Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel; Celgene: Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees. Dave:Data Driven Bioscience: Equity Ownership.

No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

Flowers: National Cancer Institute: Research Funding; Millennium/Takeda: Research Funding; Eastern Cooperative Oncology Group: Research Funding; OptumRx: Consultancy; Denovo Biopharma: Consultancy; Genentech/Roche: Research Funding; V Foundation: Research Funding; Abbvie: Consultancy, Research Funding; Acerta: Research Funding; Celgene: Research Funding; Pharmacyclics/ Janssen: Consultancy; Spectrum: Consultancy; Burroughs Wellcome Fund: Research Funding; Bayer: Consultancy; Karyopharm: Consultancy; Gilead: Research Funding; Genentech/Roche: Consultancy; Pharmacyclics: Research Funding; Janssen Pharmaceutical: Research Funding; Abbvie: Research Funding; BeiGene: Research Funding; TG Therapeutics: Research Funding; Gilead: Consultancy. Lossos:Affimed: Research Funding. Bernal-Mizrachi:Takeda Pharmaceutical Company: Research Funding; Kodikaz Therapeutic Solutions: Consultancy, Equity Ownership.

Introduction:Plasmablastic lymphoma (PL) is an aggressive variant of lymphoma, with strong association with HIV. Despite significant improvements in the survival of other lymphomas, PL has a short overall survival (14 months). The association with Epstein-Barr virus (EBV) infection and MYC chromosomal translocations are defining features of PL. However, the genetic causes and the role of specific mutation in PL are largely unknown. This limitation hinders the design of therapeutic approaches aimed to improve PL survival. Therefore, we performed a comprehensive analysis of the genetic landscape in PL.

Diffuse large B cell lymphoma (DLBCL) is the most common hematologic malignancy and is characterized by a striking degree of clinical and molecular heterogeneity. Gene expression profiling has long been used to define subtypes and understand the heterogeneity within DLBCL. Microarray-based gene expression studies have identified cell of origin subgroups (Alizadeh et al., 2000), as well as gene expression signatures derived from host inflammatory response (Monti et al., 2005), stromal tissues (Lenz et al., 2008) as well as other biological processes. However, the connections between these gene expression signatures and genetic alterations is largely unknown. In this study, we sought to define connections between gene expression signatures and genetic alterations to better understand the biology of DLBCL.

No relevant conflicts of interest to declare.

Leppä: Roche: Consultancy, Honoraria, Other: Travel expenses, Research Funding; Takeda Pharmaceuticals: Consultancy, Honoraria, Other: Travel expenses; Janssen: Consultancy, Research Funding; CTI Bio Pharma: Consultancy; Mundipharma: Research Funding; Bayer: Other: Travel expenses, Research Funding. Flowers:ECOG: Research Funding; Gilead: Consultancy, Research Funding; Millenium/Takeda: Research Funding; Pharmacyclics, LLC, an AbbVie Company: Research Funding; TG Therapeutics: Research Funding; Mayo Clinic: Research Funding; NIH: Research Funding; Infinity: Research Funding; AbbVie: Research Funding; Genentech: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Acerta: Research Funding. Hsi:Seattle Genetics: Honoraria, Speakers Bureau; HTG Molecular Diagnostics: Consultancy, Honoraria; Eli Lilly: Research Funding; Cellerant Therapeutics: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding. Evens:Takeda: Other: Advisory board. Reddy:GILEAD: Membership on an entity's Board of Directors or advisory committees; INFINITY: Membership on an entity's Board of Directors or advisory committees; celgene: Membership on an entity's Board of Directors or advisory committees; KITE: Membership on an entity's Board of Directors or advisory committees.

No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

Burkitt lymphoma (BL) is an aggressive cancer, characterized by the deregulation of the c-myc (MYC) gene. BL is a major model disease for molecular studies in many other malignancies. Treatment of BL requires long courses of intensive chemotherapy which extracts a disproportionate toll on patients.

Mann: Quiagen: Research Funding.

No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

Over 90% of Ph-positive chronic myelogenous leukemia (“typical CML”) patients have breakpoints in the M-bcr, which typically result in b2a2 (e13a2) and/or b3a2 (e14a2) fusion mRNAs, both of which are translated into the p210 BCR-ABL protein. CML patients with the p190 BCR-ABL (m-bcr) or p230 BCR-ABL (μ-bcr) fusion genes have been reported. Atypical BCR breakpoints outside these cluster regions are extremely rare. For instance, only 8 cases have been described of e6a2 fusion CML. Very little is known about the clinical or biological characteristics of this subtype of CML, including the role of collaborating gene mutations in the development of disease. In this study, we defined the gene mutations that occurred in a rare e6a2 CML case and compared the observed gene mutations to those in “typical” chronic phase (CP)-CML cases. To our knowledge, this is the first comparison of the genetic mutations occurring in typical CML and in this rare atypical form of CML.We identified the index e6a2 CML patient, and eight additional typical CML patients for whom we had bone marrow aspirate, peripheral blood and paired normal tissue. We performed whole-exome sequencing for all of these samples using the Agilent solution-based system of exon capture, which uses RNA baits to target all protein coding genes (CCDS database), as well as ∼700 human miRNAs from miRBase (v13). In all, we generated over 3 GB of sequencing data using high throughput sequencing on the Illumina platform.We identified 15 candidate cancer genes that were somatically mutated in our e6a2 CML patient. Commonly implicated biological processes comprising these genes included transcription (STAT5A, TET2, GTF2F1), cellular differentiation (TP73), and signal transduction (GPR116). Interestingly, the majority of these mutations also occurred in typical CML, albeit at lower frequency. Thus, genes mutated common to our atypical case and typical CMLs included STAT5A, TET2, GTF2F1, ABL1 and CYP2A6. Thus, while atypical e6a2 BCR-ABL fusion CML cases are extremely rare, they appear to share many aspects of the biology with typical CMLs.This study represents an in-depth analysis of a rare e6a2 CML in combination with one of the first analyses of gene mutations that occur in typical CML. Our data provide a significant first step to identifying genes that play a role in the pathogenesis along with BCR-ABL that perhaps contribute to drug resistance, and ultimately impact overall survival.No relevant conflicts of interest to declare.

Hepatosplenic T-cell lymphoma (HSTL) is a rare form of lymphoma, comprising less than 1% of the cases. However, HSTL extracts a highly disproportionate toll on patients with a median age of diagnosis of 35 years and an expected median survival of less than two years. The vast majority of HSTL patients eventually succumb to their disease. The genetic basis of the disease is largely unknown. Although abnormalities of chromosome 7, including isochromosome 7q occur commonly in the disease, the role of specific genes and genetic mutations to the disease remains essentially unknown.In this study, we sought to define the genetic features of HSTL through the whole genome sequencing and exome sequencing of 32 HSTL tumors and germline DNA (where available) from the same patients. Exome enrichment of DNA was carried out using the Agilent solution-based system of exon capture, which uses RNA baits to target all protein coding genes as well as ∼700 human microRNAs. Both whole genome and exome sequencing were performed using the Illumina platform.We identified 28 candidate cancer genes that were recurrently mutated in HSTL. Commonly implicated biological processes comprising these genes included signal transduction (e.g. PIK3CD, KRAS) and chromatin modification (e.g. TET1, SETD2 and MLL3), accounting for 16% and 23% of the total genetic events, respectively. Nearly all of these genes have been implicated in HSTL for the first time and provide new insights into the pathogenesis of the disease and potential targets for therapy. Whole genome sequencing confirmed isochromosome 7q as the most common recurrent chromosomal abnormality in HSTL and additional structural genetic alterations in chromosome 7.Our study provides the most comprehensive genetic portrait of HSTL to date, and is a significant step in defining the genetic causes of this disease.No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

Wnt signaling is a highly conserved pathway with diverse roles in proliferation, self-renewal and differentiation in multiple tissue types. Aberrant Wnt signaling and deletions/mutations of Wnt regulatory genes have been found in a variety of malignancies including lymphoma. The factors which govern this pathway and the precise role of Wnt signaling in mediating tumor proliferation are unknown. We previously identified recurrent mutations in Wnt pathway genes including APC and WIF1 in diffuse large B cell lymphoma (DLBCL) using exome sequencing and thus sought to investigate if targeting of Wnt with molecular agents would be a viable treatment approach in NHL.Alamar blue proliferation assays were used to determine IC50 values for 2 small molecule inhibitors of Wnt signaling: ICG001 (a selective inhibitor of TCF/LEF1 mediated transcription) and salinomycin (an ionophore which has been shown to target Wnt) in a panel of 65 cell lines comprising subtypes of lymphoma (including 7 ABC DLBCL, 16 GCB DLBCL, 10 Burkitt lymphoma, 5 mantle cell lymphoma and 5 primary effusion lymphoma cell lines) and leukemia. Both molecules inhibited proliferation of cell lines in the screening panel at physiologically achievable concentrations (IC50 range for salinomycin 0.11–6.23 μM, IC50 range for ICG001 0.96–28.9 μM). IC50s values for salinomycin and ICG001 correlated well (Pearson r=0.56) suggesting a common mechanism of action between the 2 agents. Furthermore, we found lower IC50 values in primary effusion lymphoma (PEL) cell lines (0.35 μM vs. 1.37 μM for other cell line histologies, p=0.01 for salinomycin and 3.25 μM vs. 6.22 μM, p=0.13 for ICG001), which is a lymphoma subtype associated with aberrant Wnt signaling. We did not note significant differences in cytoxicity among other subsets. Wnt activity was confirmed using TOPFLASH luciferase assays and correlated with sensitivity to ICG001 and salinomycin.We then used gene expression profiles derived from Affymetrix Gene 1.0 arrays from each cell line and a Cox proportional hazards model to identify genes associated with sensitivity or resistance to both agents. A gene list consisting of 367 genes associated with either sensitivity or resistance at p<0.05 for both agents was derived from this analysis. Hierarchical clustering was used to define 2 groups of cell lines based upon these genes and analyzed with gene set enrichment (GSEA). GSEA comparisons of sensitive and resistance groups derived from this method revealed that genes associated with mitochondrial pathways were highly associated with response to both ICG001 and salinomycin (FDR=0.036, p=0.03).Specifically, expression of genes involved in glycolysis and mitochondrial respiration including FBG1, CS, UBQRB5, OXA1L, LDHA and ATP5B was higher in ICG001 and salinomycin-sensitive cell lines. Conversely, we noted higher IC50s in cell lines with higher expression of mitochondrial genes having opposing functions, including PC and CPT1A. This overall suggests that tumor cells utilizing glycolytic and oxidative pathways are sensitive to Wnt inhibition. Remarkably, the GCB DLBCL cell line Pfeiffer also was noted to contain an inactivating mutation in APC and had the lowest noted IC50 among DLBCL cell lines.The Wnt pathway appears an important oncologic driver in at least a subset of NHLs and we found ICG001 and salinomycin have significant in vitro activity in NHLs. These agents may be most useful in subsets of tumors with high mitochondrial mass and/or oxidative phosphorylation. This work serves as a starting point to characterize the importance of the Wnt pathway in NHL and identifies inhibition of this pathway as a therapeutic opportunity in lymphomas.No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

Abstract 234

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Presents the poem "saving face," by Cassandra Love. First Line: before i had a brother; Last Line: to wash my face.

Presents the poem "street walking," by Cassandra Love. First Line: street walking; Last Line: 'slut.'

Presents the poem "She, Alone, Learned to Love," by Cassandra Love. First Line: she spends intimate nights; Last Line: sweet dreams.