Unbalanced MYC break-apart FISH patterns indicate the presence of a MYC rearrangement in HGBCL-DH-BCL2.

Fluorescence in situ hybridization (FISH) using break-apart probes is recommended for identifying high-grade B-cell lymphoma with MYC and BCL2 rearrangements (HGBCL-DH-BCL2). Unbalanced MYC break-apart patterns, where the red or green signal is lost, are commonly reported as an equivocal result by clinical laboratories. In a cohort of 297 HGBCL-DH-BCL2, 13% of tumors had unbalanced MYC break-apart patterns with loss of red (LR: 2%) or green (LG: 11%) signal. To determine the significance of these patterns, MYC rearrangements were characterized by sequencing in 130 HGBCL-DH-BCL2, including 3 LR and 14 LG tumors. A MYC rearrangement was identified for 71% of tumors with LR or LG patterns, with the majority involving immunoglobulin loci or other recurrent MYC rearrangement partners. The architecture of these rearrangements consistently preserved the rearranged MYC allele, with the MYC gene predicted to be on the derivative chromosome containing the signal that is still present in nearly all cases. MYC protein expression, MYC mRNA expression, and the proportion of tumors expressing the dark zone signature was not significantly different between balanced and unbalanced groups. These results support a recommendation that unbalanced MYC break-apart FISH patterns be reported as positive for MYC rearrangement in the context of diagnosing HGBCL-DH-BCL2.

Preclinical Development of a Romidepsin Nanoparticle Demonstrates Superior Tolerability and Efficacy in Models of Human T-Cell Lymphoma and Large Granular Lymphocyte Leukemia.

Histone deacetylase (HDAC) inhibitors are a widely recognized and valued treatment option for patients with relapsed or refractory peripheral T cell lymphomas (PTCL). Romidepsin is a relatively selective Class I HDAC inhibitor originally approved for patients with relapsed or refractory (R/R) cutaneous T cell lymphoma (CTCL) and subsequently R/R PTCL. Unfortunately, the FDA approval of romidepsin for R/R PTCL was withdrawn due to a negative Phase 4 post-marketing requirement (PMR), diminishing further the treatment options for patients with PTCL. Herein we describe the development of a first-in-class polymer nanoparticle of romidepsin (Nanoromidepsin) using an innovative amphiphilic di-block copolymer-based nanochemistry platform. Nanoromidepsin exhibited superior pharmacologic disposition, with improved tolerability and safety in murine models of T-cell lymphoma. Nanoromidepsin also exhibited superior anti-tumor efficacy in multiple models including in vitro T cell lymphoma (TCL) cell lines, ex vivo LGL leukemia primary patient samples, and murine TCL xenografts. Nanoromidepsin demonstrated greater accumulation in tumors and a statistically significant improvement in overall survival (OS) compared to romidepsin in murine xenograft models. These findings collectively justify the clinical development of Nanoromidepsin in patients with T-cell malignancies.

Integrated single cell analysis reveals co-evolution of malignant B cells and tumor micro-environment in transformed follicular lymphoma.

Histological transformation of follicular lymphoma (FL) to aggressive forms is associated with poor outcome. Phenotypic consequences of this evolution and its impact on the tumor microenvironment (TME) remain unknown. We perform single-cell whole genome sequencing (scWGS) and transcriptome sequencing (scWTS) of 11 paired pre/post-transformation patient samples and scWTS of additional samples from patients without transformation. Our analysis reveals evolutionary dynamics of transformation at single-cell resolution, highlighting a shifting TME landscape, with an emerging immune-cell exhaustion signature, co-evolving with the shifting malignant B phenotype in a regulatory ecosystem. Integration of scWGS and scWTS identifies malignant cell pathways upregulated during clonal tumor evolution. Using multi-color immunofluorescence, we transfer these findings to a TME-based transformation biomarker, subsequently validated in two independent pretreatment cohorts. Taken together, our results provide a comprehensive view of the combined genomic and phenotypic evolution of malignant cells during transformation and shifting crosstalk between malignant cells and the TME.

Current and upcoming treatment approaches to uncommon subtypes of PTCL (EATL/MEITL, SPTCL, HSTCL).

Rare subtypes of peripheral T-cell lymphoma (PTCL) including enteropathy-associated T-cell lymphoma (EATL), monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL), subcutaneous panniculitis-like T-cell lymphoma (SPTCL) and hepatosplenic T-cell lymphoma (HSTCL) are underrepresented in most registry and clinical studies. Most of the literature is obtained from small case series, single-institution retrospective studies and subgroup analyses of the largest studies with few recent and ongoing exceptions. While the pathogenesis and biology of these entities have yet to be fully elucidated, global efforts by the scientific community have started to shed some light on the most frequently deregulated pathways. In this review, we highlight the most pertinent clinical and pathologic features of rare subtypes of PTCL including EATL/MEITL, SPTCL and HSTCL. We also summarize the results of recent developments identifying potential targets for novel therapeutic strategies based on molecular studies. Finally, we highlight the underrepresentation of these rare subtypes in most clinical trials, making evidence-based therapeutic decisions extremely challenging.

Frontline therapy with bendamustine rituximab (BR) and rituximab cyclophosphamide vincristine prednisone (RCVP) confers similar long-term outcomes in patients with treatment naïve Waldenström macroglobulinemia in a real-world setting: a population-based analysis.

We report on outcomes of 111 patients with treatment naïve Waldenström macroglobulinemia (TN WM) treated with frontline bendamustine-rituximab (BR) (n = 57) or rituximab-cyclophosphamide-vincristine-prednisone (RCVP) (n = 54). Median follow-up was 60.7 months (range 1.9-231.6). Median progression-free survival (PFS) was 60.5 months (95% CI 47.6-73.4) for BR and 79.0 months (95% CI 31.3-126.8) for RCVP (p = .96). Median overall survival (OS) was not reached for BR and 153.4 months (95% CI 114.5-192.4) for RCVP (p = .37). While overall and major response rates did not differ between treatment groups, BR had numerically higher rate of very good partial response or better response (51% vs. 37%, p = .30) and complete response (26% vs. 13%, p = .13). RCVP confers comparable outcomes to BR in a real-world population of TN WM patients and remains an effective regimen, particularly when tolerance or frailty is an issue, or in resource-limited settings.

Kikuchi-Fujimoto Disease Following COVID-19 Vaccination: Experience at a Population-Based Referral Center.

OBJECTIVES: Multiple case reports describe Kikuchi-Fujimoto disease (KFD) following COVID-19 vaccination, but the true nature of this phenomenon is unknown. The purpose of this study was to further assess the relationship between KFD and COVID-19 vaccination at the population level. METHODS: Confirmed KFD cases from January 2018 to April 2022 were identified from provincial pathology archives and analyzed in the context of vaccination statistics from public health resources. RESULTS: Our statistical models provide evidence of a temporal association between KFD and both antecedent COVID-19 vaccine administration as well as age-stratified vaccination rates. Eight new cases of plausible COVID-19 vaccine-associated KFD are presented, collectively exhibiting clinical and pathologic features that overlap substantially with those of idiopathic KFD. CONCLUSIONS: Our findings indicate that KFD is observed in association with COVID-19 vaccination and suggest that mechanistic studies are warranted.

Classic Hodgkin lymphoma in young people.

Classic Hodgkin lymphoma (CHL) is a unique form of lymphoid cancer featuring a heterogeneous tumor microenvironment and a relative paucity of malignant Hodgkin and Reed-Sternberg (HRS) cells with characteristic phenotype. Younger individuals (children, adolescents and young adults) are affected as often as the elderly, producing a peculiar bimodal age-incidence profile that has generated immense interest in this disease and its origins. Decades of epidemiological investigations have documented the populations most susceptible and identified multiple risk factors that can be broadly categorized as either biological or environmental in nature. Most risk factors result in overt immunodeficiency or confer more subtle alterations to baseline health, physiology or immune function. Epstein Barr virus, however, is both a risk factor and well-established driver of lymphomagenesis in a significant subset of cases. Epigenetic changes, along with the accumulation of somatic driver mutations and cytogenetic abnormalities are required for the malignant transformation of germinal center-experienced HRS cell precursors. Chromosomal instability and the influence of endogenous mutational processes are critical in this regard, by impacting genes involved in key signaling pathways that promote the survival and proliferation of HRS cells and their escape from immune destruction. Here we review the principal features, known risk factors and lymphomagenic mechanisms relevant to newly diagnosed CHL, with an emphasis on those most applicable to young people.

Relapse Timing Is Associated With Distinct Evolutionary Dynamics in Diffuse Large B-Cell Lymphoma.

PURPOSE: Diffuse large B-cell lymphoma (DLBCL) is cured in more than 60% of patients, but outcomes remain poor for patients experiencing disease progression or relapse (refractory or relapsed DLBCL [rrDLBCL]), particularly if these events occur early. Although previous studies examining cohorts of rrDLBCL have identified features that are enriched at relapse, few have directly compared serial biopsies to uncover biological and evolutionary dynamics driving rrDLBCL. Here, we sought to confirm the relationship between relapse timing and outcomes after second-line (immuno)chemotherapy and determine the evolutionary dynamics that underpin that relationship. PATIENTS AND METHODS: Outcomes were examined in a population-based cohort of 221 patients with DLBCL who experienced progression/relapse after frontline treatment and were treated with second-line (immuno)chemotherapy with an intention-to-treat with autologous stem-cell transplantation (ASCT). Serial DLBCL biopsies from a partially overlapping cohort of 129 patients underwent molecular characterization, including whole-genome or whole-exome sequencing in 73 patients. RESULTS: Outcomes to second-line therapy and ASCT are superior for late relapse (>2 years postdiagnosis) versus primary refractory (<9 months) or early relapse (9-24 months). Diagnostic and relapse biopsies were mostly concordant for cell-of-origin classification and genetics-based subgroup. Despite this concordance, the number of mutations exclusive to each biopsy increased with time since diagnosis, and late relapses shared few mutations with their diagnostic counterpart, demonstrating a branching evolution pattern. In patients with highly divergent tumors, many of the same genes acquired new mutations independently in each tumor, suggesting that the earliest mutations in a shared precursor cell constrain tumor evolution toward the same genetics-based subgroups at both diagnosis and relapse. CONCLUSION: These results suggest that late relapses commonly represent genetically distinct and chemotherapy-naïve disease and have implications for optimal patient management.

Relapse timing is associated with distinct evolutionary dynamics in DLBCL.

Diffuse large B-cell lymphoma (DLBCL) is cured in over 60% of patients, but outcomes are poor for patients with relapsed or refractory disease (rrDLBCL). Here, we performed whole genome/exome sequencing (WGS/WES) on tumors from 73 serially-biopsied patients with rrDLBCL. Based on the observation that outcomes to salvage therapy/autologous stem cell transplantation are related to time-to-relapse, we stratified patients into groups according to relapse timing to explore the relationship to genetic divergence and sensitivity to salvage immunochemotherapy. The degree of mutational divergence increased with time between biopsies, yet tumor pairs were mostly concordant for cell-of-origin, oncogene rearrangement status and genetics-based subgroup. In patients with highly divergent tumors, several genes acquired exclusive mutations independently in each tumor, which, along with concordance of genetics-based subgroups, suggests that the earliest mutations in a shared precursor cell constrain tumor evolution. These results suggest that late relapses commonly represent genetically distinct and chemotherapy-naïve disease.

Molecular determinants of clinical outcomes in a real-world diffuse large B-cell lymphoma population.

Molecular heterogeneity of diffuse large B-cell lymphoma (DLBCL) underlies the variable outcomes achieved with immunochemotherapy. However, outcomes of gene expression profiling (GEP)-defined molecular subgroups in a real-world DLBCL population remain unknown. Here we examined the prevalence and outcomes of molecular subgroups in an unselected population of 1149 patients with de novo DLBCL in British Columbia, Canada. Evaluable biopsies were profiled by fluorescence in situ hybridization (FISH), immunohistochemistry, and digital GEP to assign cell-of-origin and the so-called "double-hit signature" (DHITsig)-a signature originally described as being characteristic for high-grade B-cell lymphoma with MYC and BCL2 rearrangements (HGBCL-DH-BCL2). DHITsig was expressed in 21% of 431 germinal center B-cell-like (GCB)-DLBCL and all 55 Burkitt lymphomas examined. Reflecting this latter finding, DHITsig has been renamed the "dark zone signature" (DZsig). DZsigpos-DLBCL, non-DZsigpos GCB-DLBCL and activated B-cell-like (ABC)-DLBCL were associated with a 2 year overall survival of 57%, 89%, and 71%, respectively. 62% of DZsigpos tumors were negative for HGBCL-DH-BCL2 by FISH, but were associated with outcomes similar to HGBCL-DH-BCL2. A small group of HGBCL-DH-BCL2 that lacked DZsig expression had different molecular features compared with DZsig-expressing HGBCL-DH-BCL2 and were associated with favorable outcomes comparable to DLBCL, not otherwise specified. DZsigpos and ABC-DLBCL had a shorter diagnosis-to-treatment interval (DTI) than GCB-DLBCL, with this metric being associated with outcome. In conclusion, DZsig expression extends beyond HGBCL-DH-BCL2 and captures a poor-prognosis DLBCL subgroup with short DTI, including patients unidentifiable by routine FISH testing, that should be considered for treatment intensification or novel therapies in prospective trials.

Increased IgG4+ plasma cells are common in excised lymph nodes from children and adolescents without IgG4-related disease.

Lymphadenopathy is a common finding in patients with IgG4-related disease (IgG4-RD) and often associated with increased IgG4+ plasma cells in this setting. The histologic features of so-called IgG4-related lymphadenopathy (IgG4-LAD) have seldom been investigated in children and adolescents, and step-wise progression to extranodal IgG4-RD has not been described. This study was performed to further evaluate the frequency, pathologic features, and clinical significance of IgG4-LAD-like histologic changes in the pediatric setting. We analyzed 37 benign lymph nodes collected semi-consecutively from children aged 0-18 years at our institution for both absolute and relative IgG4+ plasma cell abundance and recurrent histomorphologic patterns associated with IgG4-LAD. The combination of IgG4+/IgG+ plasma cell ratio >40% and IgG4+ plasma cell count ≥50 were considered as IgG4-LAD-like per expert consensus guidelines. Seven cases (19%) met both diagnostic criteria. The dominant histomorphologic patterns were follicular hyperplasia (n = 6), interfollicular expansion (n = 3), and progressive transformation of germinal centers (n = 3). Extranodal manifestations of IgG4-RD were not identified in this cohort (38 months average follow-up). Instead, clinical and laboratory findings indicated that lymph node enlargement in most patients could likely be attributed to alternative processes including antecedent dentistry, concurrent infection, and incipient Crohn's disease. Our findings suggest that the histologic features of IgG4-LAD are likely much more common in children and adolescents than previously recognized, often existing in complex with common reactive lymphadenopathies. The diagnostic value of routine immunohistochemical assessment for IgG4+ plasma cells in benign lymph nodes from pediatric patients without established extranodal IgG4-RD and/or other supportive clinical and laboratory data is therefore uncertain.

Single-cell profiling reveals a memory B cell-like subtype of follicular lymphoma with increased transformation risk.

Follicular lymphoma (FL) is an indolent cancer of mature B-cells but with ongoing risk of transformation to more aggressive histology over time. Recurrent mutations associated with transformation have been identified; however, prognostic features that can be discerned at diagnosis could be clinically useful. We present here comprehensive profiling of both tumor and immune compartments in 155 diagnostic FL biopsies at single-cell resolution by mass cytometry. This revealed a diversity of phenotypes but included two recurrent patterns, one which closely resembles germinal center B-cells (GCB) and another which appears more related to memory B-cells (MB). GCB-type tumors are enriched for EZH2, TNFRSF14, and MEF2B mutations, while MB-type tumors contain increased follicular helper T-cells. MB-type and intratumoral phenotypic diversity are independently associated with increased risk of transformation, supporting biological relevance of these features. Notably, a reduced 26-marker panel retains sufficient information to allow phenotypic profiling of future cohorts by conventional flow cytometry.

The outcome of older adults with classic Hodgkin lymphoma in British Columbia.

Outcomes in older adults with classic Hodgkin lymphoma (cHL) have traditionally been poor, in part, related to poor tolerance to standard chemotherapy. Herein, we evaluated the survival of patients with cHL aged ≥60 years in British Columbia in a population-based analysis. From 1961 to 2019, 744 patients with newly diagnosed cHL were identified. With a median follow-up of 9 years, 5-year disease-specific survival (DSS) and overall survival (OS) have improved by decade comparison (both P < .001), remaining stable in the past 20 years (DSS, P = .35; OS, P = .26). In the modern management era (2000-present), 361 of 401 patients (90%) received active therapy for cHL and had a 5-year OS of 60%. For those who received curative-intent therapy (n = 327), the 5-year progression-free survival (PFS), OS, and DSS were 60%, 65%, and 76%, respectively, and estimates were superior in those who were 60 to 69 years of age (72%, 77%, and 83%, respectively) compared with those who were 70 to 79 years of age (54%, 57%, and 70%, respectively) and ≥80 years of age (28%, 39%, and 63%, respectively) (P < .05 for all). Overall, pulmonary toxicity occurred in 58 of 279 patients (21%) treated with bleomycin, with 22 of 58 (38%) occurring after cycles 1 or 2, accounting for 8 of 20 (40%) treatment-related deaths. Outcomes in older adults with cHL have improved in recent decades; however, they remain poor for those aged ≥70 years, even in the modern treatment era. Furthermore, treatment-related toxicity remains a significant concern and use of bleomycin should be avoided in most patients.

Outcome of limited-stage nodular lymphocyte-predominant Hodgkin lymphoma and the impact of a PET-adapted approach.

Radiotherapy (RT) is typically incorporated into the treatment of limited-stage nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL), although it remains unknown whether chemotherapy alone may be suitable in select patients. We evaluated outcomes of limited-stage NLPHL at BC Cancer on the basis of era-specific guidelines: routine RT era, 1995 to 2005 (n = 36), combined modality with 2 cycles of doxorubicin, bleomycin, vinblastine, dacarbazine (ABVD) chemotherapy followed by RT or RT alone; positron emission tomography (PET) era, after 2005 (n = 63), ABVD alone (4 cycles) if the PET scan after the second cycle of ABVD (PET2) is negative, or treatment is changed to RT if PET2 is positive. Median age of patients was 38 years (range, 16-82 years), 73% were male, and 43% had stage II. With a median follow-up of 10.5 years for all patients, 5-year progression-free survival (PFS) was 91% [corrected] and was 97% for overall survival (OS), with no difference by treatment era (PFS, P = .15; [corrected] OS, P = .35). For the 49 patients who had a PET2 scan, 86% were PET negative and 14% were PET positive by Deauville criteria with 5-year PFS rates of 92% and 80% (P = .87) [corrected], respectively. This is the largest study of a PET-adapted approach in NLPHL and supports that ABVD alone may be a viable option in select patients with a negative PET2 scan, with consideration of acute and long-term toxicities.

The Incidence and Treatment Response of Double Expression of MYC and BCL2 in Patients with Diffuse Large B-Cell Lymphoma: A Systematic Review and Meta-Analysis.

MYC/BCL2 protein co-expression (i.e., double expressor) has been shown to be a negative predictor of outcome in diffuse large B-cell lymphoma (DLBCL). We aimed to establish the incidence of double expressor status in patients with de novo DLBCL and identify the predictive value of this biomarker on treatment response through systematic review and meta-analysis. PubMed and Embase were searched for studies published through December 2019 that reported proportions of double expressor DLBCL. The pooled proportions of MYC and BCL2 expression, both alone and in combination, were computed using the inverse variance method for calculating weights and by the DerSimonian-Laird method. The pooled odds ratios (ORs) of complete remission (CR) rate were calculated, and meta-regression analysis was conducted to explore heterogeneity. Forty-one studies (7054 patients) were included. The pooled incidence of double expressor status in DLBCL was 23% (95% confidence interval [CI], 20-26%), with an adjusted estimate of 31% (95% CI, 27-36%). Neither MYC/BCL2 protein cutoff values, race, mean, or median age of included patients, or overall study quality was a significant factor of heterogeneity (p ≥ 0.20). Cases without double expressor status demonstrated a higher probability of CR to rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone treatment (OR, 2.69; 95% CI, 1.55-4.67). Our results reaffirm the predictive power of this important biomarker.

Prevalence and Predictors of Bacterial Contamination in Excisional Lymph Node Biopsies: Implications for Diagnosis and Management.

Routine tissue handling exposes lymph node specimens to microbial contamination that can confound microbiological culture results and interfere with diagnosis. The scope and impact of this problem remain poorly understood. We combined over 13 years of lymph node pathology, culture data, and patient records to define the prevalence, predisposing factors, microbiology, and clinical management of false-positive lymph node cultures at a large academic medical center. Nearly one third (31.9%) of 216 cultured lymph nodes yielded bacterial growth. Approximately 90% of positive bacterial cultures grew 1 of 2 common skin-resident taxa-coagulase-negative Staphylococcus and Cutibacterium acnes-with well-documented predispositions for contamination in other clinical settings. Lymph nodes excised from axillary, cervical, and inguinal regions yielded higher positive culture rates than nodes excised from the mediastinum, suggesting proximity to the skin surface may increase contamination risk. Accordingly, cultures from thoracoscopic pulmonary resections displayed contamination rates over 5-fold lower than those from percutaneously accessed lymph nodes. Lymph nodal tissue allocated for culture in the operating room yielded unexpectedly high contamination rates, significantly higher than cultures sent from the frozen section processing area. A significant minority of contamination events were noted in the clinical record and prompted antibiotic therapy on multiple occasions. Collectively, our results illuminate the risk factors contributing to bacterial contamination and argue that routine lymph node bacterial cultures provide minimal clinical benefit for adult patients. This widespread bacterial contamination also warrants cautious implementation of increasingly sensitive molecular microbiology tools for excised tissues.

Characterization of DLBCL with a PMBL gene expression signature.

Primary mediastinal large B-cell lymphoma (PMBL) is a type of aggressive B-cell lymphoma that typically affects young adults, characterized by presence of a bulky anterior mediastinal mass. Lymphomas with gene expression features of PMBL have been described in nonmediastinal sites, raising questions about how these tumors should be classified. Here, we investigated whether these nonmediastinal lymphomas are indeed PMBLs or instead represent a distinct group within diffuse large B-cell lymphoma (DLBCL). From a cohort of 325 de novo DLBCL cases, we identified tumors from patients without evidence of anterior mediastinal involvement that expressed a PMBL expression signature (nm-PMBLsig+; n = 16; 5%). A majority of these tumors expressed MAL and CD23, proteins typically observed in bona fide PMBL (bf-PMBL). Evaluation of clinical features of nm-PMBLsig+ cases revealed close associations with DLBCL, and a majority displayed a germinal center B cell-like cell of origin (GCB). In contrast to patients with bf-PMBL, patients with nm-PMBLsig+ presented at an older age and did not show pleural disease, and bone/bone marrow involvement was observed in 3 cases. However, although clinically distinct from bf-PMBL, nm-PMBLsig+ tumors resembled bf-PMBL at the molecular level, with upregulation of immune response, JAK-STAT, and NF-κB signatures. Mutational analysis revealed frequent somatic gene mutations in SOCS1, IL4R, ITPKB, and STAT6, as well as CD83 and BIRC3, with the latter genes significantly more frequently affected than in GCB DLBCL or bf-PMBL. Our data establish nm-PMBLsig+ lymphomas as a group within DLBCL with distinct phenotypic and genetic features. These findings may have implications for gene expression- and mutation-based subtyping of aggressive B-cell lymphomas and related targeted therapies.

Notch activation is pervasive in SMZL and uncommon in DLBCL: implications for Notch signaling in B-cell tumors.

Notch receptors participate in a signaling pathway in which ligand-induced proteolysis frees the Notch intracellular domain (NICD), allowing it to translocate to the nucleus, form a transcription complex, and induce target gene expression. Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), splenic marginal zone B-cell lymphoma (SMZL), and distinct subsets of diffuse large B-cell lymphoma (DLBCL) are strongly associated with mutations in the 3' end of NOTCH1 or NOTCH2 that disrupt a proline, glutamic acid, serine, and threonine (PEST) degron domain and stabilize NICD1 and NICD2. By contrast, mutations leading to constitutive Notch activation are rare in primary B-cell neoplasms, suggesting that Notch activation is confined to ligand-rich tumor microenvironments, or that cryptic strong gain-of-function mutations have been missed in prior analyses. To test these ideas, we used immunohistochemical stains to screen a broad range of B-cell tumors for Notch activation. Our analyses reveal that among small B-cell neoplasms, NICD2 is primarily detected in SMZL and is a common feature of both NOTCH2 wild-type and NOTCH2-mutated SMZLs, similar to prior findings with NOTCH1 in CLL/SLL. The greatest NOTCH2 activation was observed in NOTCH2-mutated SMZLs, particularly within splenic marginal zones. By contrast, little evidence of NOTCH2 activation was observed in DLBCL, even in NOTCH2-mutated tumors, suggesting that selective pressure for NOTCH2 activation is mainly confined to low-grade B-cell neoplasms, whereas DLBCLs with NOTCH1 mutations frequently showed evidence of ongoing NOTCH1 activation. These observations have important implications for the pathogenic role of Notch and its therapeutic targeting in B-cell lymphomas.