Motive and Opportunity: MYC rearrangements in high-grade B-cell lymphoma with MYC and BCL2 rearrangements-an LLMPP study.

Rearrangements that place the oncogenes MYC, BCL2, or BCL6 adjacent to superenhancers are common in mature B-cell lymphomas. Lymphomas with diffuse large B-cell lymphoma (DLBCL) or high-grade morphology with both MYC and BCL2 rearrangements are classified as high-grade B-cell lymphoma with MYC and BCL2 rearrangements ("double hit": HGBCL-DH-BCL2) and are associated with aggressive disease and poor outcomes. Although it is established that MYC rearrangements involving immunoglobulin (IG) loci are associated with inferior outcomes relative to those involving other non-IG superenhancers, the frequency of, and mechanisms driving, IG vs non-IG MYC rearrangements have not been elucidated. Here we used custom targeted capture and/or whole genome sequencing to characterize oncogene rearrangements across 883 mature B-cell lymphomas including Burkitt lymphoma, follicular lymphoma, DLBCL, and HGBCL-DH-BCL2 tumors. We demonstrate that, while BCL2 rearrangement topology is consistent across entities, HGBCL-DH-BCL2 have distinct MYC rearrangement architecture relative to tumors with single MYC rearrangements or with both MYC and BCL6 rearrangements (HGBCL-DH-BCL6), including both a higher frequency of non-IG rearrangements and different architecture of MYC::IGH rearrangements. The distinct MYC rearrangement patterns in HGBCL-DH-BCL2 occur on the background of high levels of somatic hypermutation across MYC partner loci in HGBCL-DH-BCL2, creating more opportunity to form these rearrangements. Furthermore, because one IGH allele is already disrupted by the existing BCL2 rearrangement, the MYC rearrangement architecture in HGBCL-DH-BCL2 likely reflects selective pressure to preserve both BCL2 and B cell receptor expression. These data provide new mechanistic explanations for the distinct patterns of MYC rearrangements observed across different lymphoma entities.

Follicular lymphoma B cells exhibit heterogeneous transcriptional states with associated somatic alterations and tumor microenvironments.

Follicular lymphoma (FL) is an indolent non-Hodgkin lymphoma of germinal center origin, which presents with significant biologic and clinical heterogeneity. Using RNA-seq on B cells sorted from 87 FL biopsies, combined with machine-learning approaches, we identify 3 transcriptional states that divide the biological ontology of FL B cells into inflamed, proliferative, and chromatin-modifying states, with relationship to prior GC B cell phenotypes. When integrated with whole-exome sequencing and immune profiling, we find that each state was associated with a combination of mutations in chromatin modifiers, copy-number alterations to TNFAIP3, and T follicular helper cells (Tfh) cell interactions, or primarily by a microenvironment rich in activated T cells. Altogether, these data define FL B cell transcriptional states across a large cohort of patients, contribute to our understanding of FL heterogeneity at the tumor cell level, and provide a foundation for guiding therapeutic intervention.

Mediastinal large B cell lymphoma and surrounding gray areas: a report of the lymphoma workshop of the 20th meeting of the European Association for Haematopathology.

Session 3 of the 2021 European Association for Haematopathology/Society for Hematopathology Workshop focused on mediastinal large B cell lymphomas and surrounding gray areas. One half of the session was dedicated to primary mediastinal large B cell lymphoma (PMBL) and included cases with classic clinicopathologic features, as well as cases with either morphologic or immunophenotypic variation, and PMBL-like cases with primary extramediastinal disease. The role of additional immunophenotyping and/or molecular testing to aid in the diagnosis of PMBL was discussed. The second half of the session focused on mediastinal and non-mediastinal gray zone lymphomas (GZL) with features intermediate between diffuse large B cell lymphoma (DLBCL) and classic Hodgkin lymphoma (CHL). Several cases illustrating the current challenges in separating this entity from PMBL/DLBCL and CHL were presented. There was discussion regarding the clinical and genetic differences between mediastinal and non-mediastinal GZLs. Rare cases of PMBL and GZL associated with EBV or follicular lymphoma were reviewed. Finally, several cases included in the session highlighted composite or sequential CHL and PMBL/DLBCL and/or GZL, highlighting challenges in separating such cases from GZL.

Subclonal TP53 mutations are frequent and predict resistance to radioimmunotherapy in follicular lymphoma.

Although TP53 is commonly mutated in transformed follicular lymphoma, mutations are reported in <5% of pretreatment follicular lymphoma (FL) specimens. We assayed archival follicular B-cell non-Hodgkin lymphoma specimens from a completed clinical trial, Southwest Oncology Group S0016, a phase 3 randomized intergroup trial of CHOP (cyclophosphamide, hydroxydaunorubicin, oncovin, and prednisone) chemotherapy plus R-CHOP (rituximab-CHOP) compared with CHOP chemotherapy plus 131-iodine tositumomab (radioimmunotherapy [RIT]-CHOP). Subclonal TP53 mutations (median allele frequency 0.02) were found in 25% of diagnostic FL specimens and in 27% of a separate validation cohort. In the R-CHOP arm, pathogenic TP53 mutations were not associated with progression-free survival (PFS) (10-year PFS 43% vs 44%). In contrast, among patients with no detectable pathogenic TP53 mutation, RIT-CHOP was associated with a longer PFS than with R-CHOP (10-year PFS 67% vs 44%; hazard ratio = 0.49; P = .008). No relationship was detected between PFS and the extent of activation-induced cytidine deaminase (AICDA)-mediated heterogeneity. In summary, subclonal TP53 mutations are common in FL and are a distinct phenomenon from AICDA-mediated genetic heterogeneity. The absence of a detectable subclonal mutation in TP53 defined a population that particularly benefited from RIT.

Molecular classification and identification of an aggressive signature in low-grade B-cell lymphomas.

Non-follicular low-grade B-cell lymphomas (LGBCL) are biologically diverse entities that share clinical and histologic features that make definitive pathologic categorization challenging. While most patients with LGBCL have an indolent course, some experience aggressive disease, highlighting additional heterogeneity across these subtypes. To investigate the potential for shared biology across subtypes, we performed RNA sequencing and applied machine learning approaches that identified five clusters of patients that grouped independently of subtype. One cluster was characterized by inferior outcome, upregulation of cell cycle genes, and increased tumor immune cell content. Integration of whole exome sequencing identified novel LGBCL mutations and enrichment of TNFAIP3 and BCL2 alterations in the poor survival cluster. Building on this, we further refined a transcriptomic signature associated with early clinical failure in two independent cohorts. Taken together, this study identifies unique clusters of LGBCL defined by novel gene expression signatures and immune profiles associated with outcome across diagnostic subtypes.

Transformation of FL into DLBCL with a PMBL gene expression signature.

We investigated the clinicopathologic features of 5 follicular lymphomas (FLs) that transformed (tFL) morphologically to diffuse large B-cell lymphomas (DLBCLs) and had a primary mediastinal large B-cell lymphoma (PMBL)-like gene expression profile (tFL-PMBLsig-pos). None of the tFL-PMBLsig-pos cases arose in the mediastinum, all cases tested had a germinal center B-cell phenotype, 20% were CD30+, 60% CD23+, 80% MAL+, 20% CD200+, and 0% CD273/PDL2+. Whole-exome sequencing detected alterations in genes associated with both FL/DLBCL (CREBBP, KMT2C, KMT2D, ARID1A, HIST1 members, and TNFRSF14) and PMBL (JAK-STAT pathway genes, B2M, and CD58). Copy number (CN) analysis detected gains/amplification of REL and STAT6 in 60%, gains of SOCS1 in 40%, and gains of chromosome 16, including IL4R, in 40% of the cases. CN gains/amplification of BCL6 and MYC and loss of TNFRSF14 and TNFAIP3 were identified in 20% of the cases. Three of 5 cases lacked a BCL2 rearrangement. Despite having some features that are less common in DLBCL (MAL and CD23 expression and JAK-STAT activation), these tFL-PMBLsig-pos cases lack the most characteristic CN alteration seen in PMBL (9p24.1 gain/amplification). This cohort expands the biologic heterogeneity of tFL, illustrating a subset with gene expression and some genetic features reminiscent of PMBL, with potential treatment implications that include the use of novel targeted therapies.

Longitudinal expression profiling identifies a poor risk subset of patients with ABC-type diffuse large B-cell lymphoma.

Despite the effectiveness of immuno-chemotherapy, 40% of patients with diffuse large B-cell lymphoma (DLBCL) experience relapse or refractory disease. Longitudinal studies have previously focused on the mutational landscape of relapse but fell short of providing a consistent relapse-specific genetic signature. In our study, we have focused attention on the changes in GEP accompanying DLBCL relapse using archival paired diagnostic/relapse specimens from 38 de novo patients with DLBCL. COO remained stable from diagnosis to relapse in 80% of patients, with only a single patient showing COO switching from activated B-cell-like (ABC) to germinal center B-cell-like (GCB). Analysis of the transcriptomic changes that occur following relapse suggest ABC and GCB relapses are mediated via different mechanisms. We developed a 30-gene discriminator for ABC-DLBCLs derived from relapse-associated genes that defined clinically distinct high- and low-risk subgroups in ABC-DLBCLs at diagnosis in datasets comprising both population-based and clinical trial cohorts. This signature also identified a population of <60-year-old patients with superior PFS and OS treated with ibrutinib-R-CHOP as part of the PHOENIX trial. Altogether this new signature adds to the existing toolkit of putative genetic predictors now available in DLBCL that can be readily assessed as part of prospective clinical trials.

"Quadruple-hit" primary testicular diffuse large B-cell lymphoma with MYD88 L265P mutation, IGH::MYC, and IRF4- and BCL6-rearrangements.

Classification of DLBCL relies on clinical, immunohistochemical, and genetic information. We report a case of primary testicular diffuse large B-cell lymphoma (PT-DLBCL) with a hitherto unreported constellation of pathologic findings to illustrate the challenges of DLBCL classification. A standard hematopathology workup was followed by gene expression profiling (GEP) to determine the DLBCL cell of origin (COO). A 75-year-old man presented with a unilateral testicular mass that had developed over the course of 1 month. Pathologic examination demonstrated involvement by DLBCL. Clinical staging revealed no systemic disease. Genetic testing showed an MYD88 mutation, as well as IGH::MYC and IRF4- and BCL6-rearrangements. Gene expression profiling demonstrated an activated B-cell expression profile. This case highlights the genetic complexity of DLBCL arising in the testis and questions the clinical significance of the identified genetic abnormalities.

Transcriptional profiles define drug refractory disease in myeloma.

Identifying biomarkers associated with disease progression and drug resistance are important for personalized care. We investigated the expression of 121 curated genes, related to immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs) responsiveness. We analyzed 28 human multiple myeloma (MM) cell lines with known drug sensitivities and 130 primary MM patient samples collected at different disease stages, including newly diagnosed (ND), on therapy (OT), and relapsed and refractory (RR, collected within 12 months before the patients' death) timepoints. Our findings led to the identification of a subset of genes linked to clinical drug resistance, poor survival, and disease progression following combination treatment containing IMIDs and/or PIs. Finally, we built a seven-gene model (MM-IMiD and PI sensitivity-7 genes [IP-7]) using digital gene expression profiling data that significantly separates ND patients from IMiD- and PI-refractory RR patients. Using this model, we retrospectively analyzed RNA sequcencing (RNAseq) data from the Mulltiple Myeloma Research Foundation (MMRF) CoMMpass (n = 578) and Mayo Clinic MM patient registry (n = 487) to divide patients into probabilities of responder and nonresponder, which subsequently correlated with overall survival, disease stage, and number of prior treatments. Our findings suggest that this model may be useful in predicting acquired resistance to treatments containing IMiDs and/or PIs.

Gene Expression Signatures for the Accurate Diagnosis of Peripheral T-Cell Lymphoma Entities in the Routine Clinical Practice.

PURPOSE: Peripheral T-cell lymphoma (PTCL) includes heterogeneous clinicopathologic entities with numerous diagnostic and treatment challenges. We previously defined robust transcriptomic signatures that distinguish common PTCL entities and identified two novel biologic and prognostic PTCL-not otherwise specified subtypes (PTCL-TBX21 and PTCL-GATA3). We aimed to consolidate a gene expression-based subclassification using formalin-fixed, paraffin-embedded (FFPE) tissues to improve the accuracy and precision in PTCL diagnosis. MATERIALS AND METHODS: We assembled a well-characterized PTCL training cohort (n = 105) with gene expression profiling data to derive a diagnostic signature using fresh-frozen tissue on the HG-U133plus2.0 platform (Affymetrix, Inc, Santa Clara, CA) subsequently validated using matched FFPE tissues in a digital gene expression profiling platform (nCounter, NanoString Technologies, Inc, Seattle, WA). Statistical filtering approaches were applied to refine the transcriptomic signatures and then validated in another PTCL cohort (n = 140) with rigorous pathology review and ancillary assays. RESULTS: In the training cohort, the refined transcriptomic classifier in FFPE tissues showed high sensitivity (> 80%), specificity (> 95%), and accuracy (> 94%) for PTCL subclassification compared with the fresh-frozen-derived diagnostic model and showed high reproducibility between three independent laboratories. In the validation cohort, the transcriptional classifier matched the pathology diagnosis rendered by three expert hematopathologists in 85% (n = 119) of the cases, showed borderline association with the molecular signatures in 6% (n = 8), and disagreed in 8% (n = 11). The classifier improved the pathology diagnosis in two cases, validated by clinical findings. Of the 11 cases with disagreements, four had a molecular classification that may provide an improvement over pathology diagnosis on the basis of overall transcriptomic and morphological features. The molecular subclassification provided a comprehensive molecular characterization of PTCL subtypes, including viral etiologic factors and translocation partners. CONCLUSION: We developed a novel transcriptomic approach for PTCL subclassification that facilitates translation into clinical practice with higher precision and uniformity than conventional pathology diagnosis.

Causes of death in low-grade B-cell lymphomas in the rituximab era: a prospective cohort study.

Low-grade B-cell lymphomas other than follicular and small lymphocytic lymphoma (LGBCL) account for 10% of all B-cell non-Hodgkin lymphomas. Despite improvements in survival outcomes for these patients, little is known about cause of death (COD) in the rituximab era. For a better understanding, we studied 822 newly diagnosed patients with marginal zone, lymphoplasmacytic, and unclassifiable low-grade B-cell lymphoma prospectively enrolled in the University of Iowa/Mayo Clinic Specialized Program of Research Excellence Molecular Epidemiology Resource from 2002 to 2015. COD was assigned based on medical record review using a standard protocol. At a median follow-up of 107 months, 219 (27%) patients had died. The incidence of lymphoma-related deaths when pooling across subtypes was lower than non-lymphoma-related deaths (10-year incidence, 8.0%; 95% confidence interval [CI]: 6.2-10.4 vs 13.6%; 95% CI: 11.2-16.6). The incidence of lymphoma-related deaths varied by subtype, ranging from 3.7% at 10 years in extranodal marginal zone lymphoma to 19.3% in lymphoplasmacytic lymphoma/Waldenström macroglobulinemia. Patients with early progression or retreatment events, defined using event-free survival at 24 months from diagnosis, had significantly higher likelihood of lymphoma-related death compared with patients without early events (10-year estimate: 19.1% vs 5.1%, respectively; P < .001), whereas the rates for non-lymphoma-related death were comparable in patients with or without early events (10-year estimates: 11.0% vs 15.3%, respectively). In conclusion, the most common COD in LGBCLs in the first decade after diagnosis was for causes other than lymphoma. Progression or retreatment within the first 2 years of diagnosis was a strong predictor for risk of lymphoma-related death.

DNMT3A mutations define a unique biological and prognostic subgroup associated with cytotoxic T cells in PTCL-NOS.

Peripheral T-cell lymphomas (PTCLs) are heterogenous T-cell neoplasms often associated with epigenetic dysregulation. We investigated de novo DNA methyltransferase 3A (DNMT3A) mutations in common PTCL entities, including angioimmunoblastic T-cell lymphoma and novel molecular subtypes identified within PTCL-not otherwise specified (PTCL-NOS) designated as PTCL-GATA3 and PTCL-TBX21. DNMT3A-mutated PTCL-TBX21 cases showed inferior overall survival (OS), with DNMT3A-mutated residues skewed toward the methyltransferase domain and dimerization motif (S881-R887). Transcriptional profiling demonstrated significant enrichment of activated CD8+ T-cell cytotoxic gene signatures in the DNMT3A-mutant PTCL-TBX21 cases, which was further validated using immunohistochemistry. Genomewide methylation analysis of DNMT3A-mutant vs wild-type (WT) PTCL-TBX21 cases demonstrated hypomethylation in target genes regulating interferon-γ (IFN-γ), T-cell receptor signaling, and EOMES (eomesodermin), a master transcriptional regulator of cytotoxic effector cells. Similar findings were observed in a murine model of PTCL with Dnmt3a loss (in vivo) and further validated in vitro by ectopic expression of DNMT3A mutants (DNMT3A-R882, -Q886, and -V716, vs WT) in CD8+ T-cell line, resulting in T-cell activation and EOMES upregulation. Furthermore, stable, ectopic expression of the DNMT3A mutants in primary CD3+ T-cell cultures resulted in the preferential outgrowth of CD8+ T cells with DNMT3AR882H mutation. Single-cell RNA sequencing(RNA-seq) analysis of CD3+ T cells revealed differential CD8+ T-cell subset polarization, mirroring findings in DNMT3A-mutated PTCL-TBX21 and validating the cytotoxic and T-cell memory transcriptional programs associated with the DNMT3AR882H mutation. Our findings indicate that DNMT3A mutations define a cytotoxic subset in PTCL-TBX21 with prognostic significance and thus may further refine pathological heterogeneity in PTCL-NOS and suggest alternative treatment strategies for this subset.

Bendamustine or high-dose cytarabine-based induction with rituximab in transplant-eligible mantle cell lymphoma.

The objective of this study was to explore differences in outcomes between first-line rituximab plus bendamustine (R-B) and R-CHOP/R-DHAP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone, dexamethasone, cytarabine, cisplatin) in transplant-eligible patients with mantle cell lymphoma (MCL). A population-based cohort of 97 patients aged 18 to 65 years with stage II-IV MCL, consecutively treated with R-B was retrospectively identified at BC Cancer. Baseline characteristics, response rates, and outcomes were compared with the cohort of 232 patients with MCL randomized to the R-CHOP/R-DHAP arm of the MCL Younger trial. The primary endpoint was the hazard ratio (HR) of the progression-free survival (PFS) comparison between both groups, adjusted for MCL International Prognostic Index (MIPI), Ki67 index, and blastoid/ pleomorphic morphology. Ann Arbor stage, lactate dehydrogenase, MIPI, blastoid morphology, and MCL35 assignments were similar between both groups. The overall response rate (ORR) to R-B was 90% (54% complete response [CR]); 77% of patients proceeded to autologous stem cell transplantation (ASCT) and 78% received maintenance rituximab (MR). The ORR to R-CHOP/R-DHAP was 94% (54% CR); 78% proceeded to ASCT and 2% received MR. There were no differences in PFS in unadjusted (HR, 0.87; 95% confidence interval [CI], 0.53-1.41; P = .56) or adjusted (HR, 0.79; 95% CI, 0.45-1.37; P = .40) comparisons. There were no clear differences in secondary endpoints in unadjusted or adjusted analyses. This retrospective adjusted comparison of 2 independent cohorts of younger patients with MCL suggests that R-B with ASCT and maintenance rituximab is a feasible and effective first-line treatment, with outcomes comparable to R-CHOP/R-DHAP with ASCT.

Diffuse large B-cell lymphomas in adults with aberrant coexpression of CD10, BCL6, and MUM1 are enriched in IRF4 rearrangements.

Diffuse large B-cell lymphoma (DLBCL) with aberrant coexpression of CD10+BCL6+MUM1+ (DLBCL-AE), classified as germinal center B cell (GCB) type by the Hans algorithm (HA), was genetically characterized. To capture the complexity of DLBCL-AE, we used an integrated approach that included gene expression profiling (GEP), fluorescence in situ hybridization, targeted gene sequencing, and copy number (CN) arrays. According to GEP, 32/54 (59%) cases were classified as GCB-DLBCL, 16/54 (30%) as activated B-cell (ABC) DLBCL, and 6/54 (11%) as unclassifiable. The discrepancy between HA and GEP was 41%. Three genetic subgroups were identified. Group 1 included 13/50 (26%) cases without translocations and mainly showing and ABC/MCD molecular profile. Group 2 comprised 11/50 (22%) cases with IRF4 alterations (DLBCL-IRF4), frequent mutations in IRF4 (82%) and NF-κB pathway genes (MYD88, CARD11, and CD79B), and losses of 17p13.2. Five cases each were classified as GCB- or ABC-type. Group 3 included 26/50 (52%) cases with 1 or several translocations in BCL2/BCL6/MYC/IGH, and GCB/EZB molecular profile predominated. Two cases in this latter group showed complex BCL2/BCL6/IRF4 translocations. DLBCL-IRF4 in adults showed a similar copy number profile and shared recurrent CARD11 and CD79B mutations when compared with LBCL-IRF4 in the pediatric population. However, adult cases showed higher genetic complexity, higher mutational load with frequent MYD88 and KMT2D mutations, and more ABC GEP. IRF4 mutations were identified only in IRF4-rearranged cases, indicating its potential use in the diagnostic setting. In conclusion, DLBCL-AE is genetically heterogeneous and enriched in cases with IRF4 alterations. DLBCL-IRF4 in adults has many similarities to the pediatric counterpart.

Genome-Wide miRNA Expression Profiling of Molecular Subgroups of Peripheral T-cell Lymphoma.

PURPOSE: Peripheral T-cell lymphoma (PTCL) is a heterogeneous group of non-Hodgkin lymphomas with aggressive clinical behavior. We performed comprehensive miRNA profiling in PTCLs and corresponding normal CD4+ Th1/2 and TFH-like polarized subsets to elucidate the role of miRNAs in T-cell lymphomagenesis. EXPERIMENTAL DESIGN: We used nCounter (NanoString Inc) for miRNA profiling and validated using Taqman qRT-PCR (Applied Biosystems, Inc). Normal CD4+ T cells were polarized into effector Th subsets using signature cytokines, and miRNA significance was revealed using functional experiments. RESULTS: Effector Th subsets showed distinct miRNA expression with corresponding transcription factor expression (e.g., BCL6/miR-19b, -106, -30d, -26b, in IL21-polarized; GATA3/miR-155, miR-337 in Th2-polarized; and TBX21/miR-181a, -331-3p in Th1-polarized cells). Integration of miRNA signatures suggested activation of TCR and PI3K signaling in IL21-polarized cells, ERK signaling in Th1-polarized cells, and AKT-mTOR signaling in Th2-polarized cells, validated at protein level. In neoplastic counterparts, distinctive miRNAs were identified and confirmed in an independent cohort. Integrative miRNA-mRNA analysis identified a decrease in target transcript abundance leading to deregulation of sphingolipid and Wnt signaling and epigenetic dysregulation in angioimmunoblastic T-cell lymphoma (AITL), while ERK, MAPK, and cell cycle were identified in PTCL subsets, and decreased target transcript abundance was validated in an independent cohort. Elevated expression of miRNAs (miR-126-3p, miR-145-5p) in AITL was associated with poor clinical outcome. In silico and experimental validation suggest two targets (miR-126→ SIPR2 and miR-145 → ROCK1) resulting in reduced RhoA-GTPase activity and T-B-cell interaction. CONCLUSIONS: Unique miRNAs and deregulated oncogenic pathways are associated with PTCL subtypes. Upregulated miRNA-126-3p and miR-145-5p expression regulate RhoA-GTPase and inhibit T-cell migration, crucial for AITL pathobiology.

EBV-positive HIV-associated diffuse large B cell lymphomas are characterized by JAK/STAT (STAT3) pathway mutations and unique clinicopathologic features.

Even in the era of highly active combination antiretroviral therapy (cART), patients with HIV have a disproportionate risk of developing aggressive lymphomas that are frequently Epstein-Barr virus (EBV)-related. Here, we investigate HIV-associated diffuse large B-cell lymphoma (HIV-DLBCL) and compare EBV-positive and EBV-negative cases. HIV-DLBCL were identified from two academic medical centres and characterised by immunohistochemistry, EBV status, fluorescence in situ hybridisation, cell of origin determination by gene expression profiling, and targeted deep sequencing using a custom mutation panel of 334 genes. We also applied the Lymphgen tool to determine the genetic subtype of each case. Thirty HIV-DLBCL were identified, with a median patient age of 46 years and male predominance (5:1). Thirteen cases (48%) were EBV-positive and 14 (52%) EBV-negative. Nine of the 16 tested cases (56%) had MYC rearrangement, three (19%) had BCL6 (two of which were double hit MYC/BCL6) and none had BCL2 rearrangements. Using the Lymphgen tool, half of the cases (15) were classified as other. All HIV-DLBCL showed mutational abnormalities, the most frequent being TP53 (37%), MYC (30%), STAT3 (27%), HIST1H1E (23%), EP300 (20%), TET2 (20%), SOCS1 (17%) and SGK1 (17%). EBV-negative cases were mostly of germinal centre B-cell (GCB) origin (62%), showed more frequent mutations per case (a median of 13·5/case) and significant enrichment of TP53 (57% vs. 15%; P = 0·046), SGK1 (36% vs. 0%; P = 0·04), EP300 (43% vs. 0%; P = 0·02) and histone-modifying gene (e.g. HIST1H1E, HIST1H1D, 79% vs. 31%; P = 0·02) mutations. EBV-positive cases were mostly of non-GCB origin (70%), with fewer mutations per case (median 8/case; P = 0·007), and these tumours were enriched for STAT3 mutations (P = 0·10). EBV-positive cases had a higher frequency of MYC mutations but the difference was not significant (36% vs. 15%; P = 0·38). EBV-association was more frequent in HIV-DLBCLs, arising in patients with lower CD4 counts at diagnosis (median 46·5 vs. 101, P = 0·018). In the era of cART, approximately half of HIV-DLBCL are EBV-related. HIV-DLBCL are enriched for MYC rearrangements, MYC mutations and generally lack BCL2 rearrangements, regardless of EBV status. Among HIV-DLBCL, tumours that are EBV-negative and EBV-positive appear to have important differences, the latter arising in context of lower CD4 count, showing frequent non-GCB origin, lower mutation burden and recurrent STAT3 mutations.

MAPK and JAK-STAT pathways dysregulation in plasmablastic lymphoma.

Plasmablastic lymphoma (PBL) is an aggressive B-cell lymphoma with an immunoblastic/large-cell morphology and terminal B-cell differentiation. The differential diagnosis from Burkitt lymphoma, plasma cell myeloma and some variants of diffuse large B-cell lymphoma may be challenging because of the overlapping morphological, genetic and immunophenotypic features. Furthermore, the genomic landscape in PBL is not well known. To characterize the genetic and molecular heterogeneity of these tumors, we investigated 34 cases of PBL using an integrated approach, including fluorescence in situ hybridization, targeted sequencing of 94 B-cell lymphoma-related genes, and copy-number arrays. PBL were characterized by high genetic complexity including MYC translocations (87%), gains of 1q21.1-q44, trisomy 7, 8q23.2- q24.21, 11p13-p11.2, 11q14.2-q25, 12p and 19p13.3-p13.13, losses of 1p33, 1p31.1-p22.3, 13q and 17p13.3-p11.2, and recurrent mutations of STAT3 (37%), NRAS and TP53 (33%), MYC and EP300 (19%) and CARD11, SOCS1 and TET2 (11%). Pathway enrichment analysis suggested a cooperative action between MYC alterations and MAPK (49%) and JAK-STAT (40%) signaling pathways. Of note, Epstein-Barr virus (EBV)-negative PBL cases had higher mutational and copy-number load and more frequent TP53, CARD11 and MYC mutations, whereas EBV-positive PBL tended to have more mutations affecting the JAK-STAT pathway. In conclusion, these findings further unravel the distinctive molecular heterogeneity of PBL identifying novel molecular targets and the different genetic profile of these tumors in relation to EBV infection.

Mastocytosis.

OBJECTIVES: The 2019 Workshop of the Society for Hematopathology/European Association for Haematopathology received and reviewed cases covering the spectrum of mastocytosis and related diseases, including morphologic mimics, focusing on recent updates and relevant findings for pathologists. METHODS: The workshop panel reviewed 99 cases of cutaneous and systemic mastocytosis (SM) and SM and associated hematologic neoplasms (SM-AHN). RESULTS: Despite a common theme of KIT mutation (particularly D816V), mastocytosis is a heterogeneous neoplasm with a wide variety of presentations. This spectrum, including rare subtypes and extramedullary organ involvement, is discussed and illustrated by representative cases. CONCLUSIONS: In the age of targeted treatment aimed at KIT, the accurate diagnosis and classification of mastocytosis has major implications for therapy and further interventions. Understanding the clinical, pathologic, and genetic findings of mastocytosis is crucial for selecting the proper tests to perform and subsequent arrival at a correct diagnosis in this rare disease.