What is new in the 5th edition of the World Health Organization classification of mature B and T/NK cell tumors and stromal neoplasms?

The classification of tumors is essential in the diagnosis and clinical management of patients with malignant neoplasms. The World Health Organization (WHO) provides a globally applicable classification scheme of neoplasms and it was updated several times. In this review, we briefly outline the cornerstones of the upcoming 5th edition of the World Health Organization Classification of Haematolymphoid Tumours on lymphoid neoplasms. As is adopted throughout the 5th edition of the WHO classification of tumors of all organ systems, entities are listed by a hierarchical system. For the first time, tumor-like lesions have been included in the classification, and modifications of nomenclature for some entities, revisions of diagnostic criteria or subtypes, deletion of certain entities, and introduction of new entities are presented along with mesenchymal lesions specific to the stroma of lymph nodes and the spleen. In addition to specific outlines on constitutional and somatic genetic changes associated with given entities, a separate chapter on germline predisposition syndromes related to hematologic neoplasms has been added.

The fifth edition of the WHO classification of mature B-cell neoplasms: open questions for research.

The fifth edition of the World Health Organization Classification of Haematolymphoid Tumours (WHO-HAEM5) is the product of an evidence-based evolution of the revised fourth edition with wide multidisciplinary consultation. Nonetheless, while every classification incorporates scientific advances and aims to improve upon the prior version, medical knowledge remains incomplete and individual neoplasms may not be easily subclassified in a given scheme. Thus, optimal classification requires ongoing study, and there are certain aspects of some entities and subtypes that require further refinements. In this review, we highlight a selection of these challenging areas to prompt more research investigations. These include (1) a 'placeholder term' of splenic B-cell lymphoma/leukaemia with prominent nucleoli (SBLPN) to accommodate many of the splenic lymphomas previously classified as hairy cell leukaemia variant and B-prolymphocytic leukaemia, a clear new start to define their pathobiology; (2) how best to classify BCL2 rearrangement negative follicular lymphoma including those with BCL6 rearrangement, integrating the emerging new knowledge on various germinal centre B-cell subsets; (3) what is the spectrum of non-IG gene partners of MYC translocation in diffuse large B-cell lymphoma/high-grade B-cell lymphoma and how they impact MYC expression and clinical outcome; how best to investigate this in a routine clinical setting; and (4) how best to define high-grade B-cell lymphoma not otherwise specified and high-grade B-cell lymphoma with 11q aberrations to distinguish them from their mimics and characterise their molecular pathogenetic mechanism. Addressing these questions would provide more robust evidence to better define these entities/subtypes, improve their diagnosis and/or prognostic stratification, leading to better patient care. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Non-IG::MYC in diffuse large B-cell lymphoma confers variable genomic configurations and MYC transactivation potential.

MYC translocation occurs in 8-14% of diffuse large B-cell lymphoma (DLBCL), and may concur with BCL2 and/or BCL6 translocation, known as double-hit (DH) or triple-hit (TH). DLBCL-MYC/BCL2-DH/TH are largely germinal centre B-cell like subtype, but show variable clinical outcome, with IG::MYC fusion significantly associated with inferior survival. While DLBCL-MYC/BCL6-DH are variable in their cell-of-origin subtypes and clinical outcome. Intriguingly, only 40-50% of DLBCL with MYC translocation show high MYC protein expression (>70%). We studied 186 DLBCLs with MYC translocation including 32 MYC/BCL2/BCL6-TH, 75 MYC/BCL2-DH and 26 MYC/BCL6-DH. FISH revealed a MYC/BCL6 fusion in 59% of DLBCL-MYC/BCL2/BCL6-TH and 27% of DLBCL-MYC/BCL6-DH. Targeted NGS showed a similar mutation profile and LymphGen genetic subtype between DLBCL-MYC/BCL2/BCL6-TH and DLBCL-MYC/BCL2-DH, but variable LymphGen subtypes among DLBCL-MYC/BCL6-DH. MYC protein expression is uniformly high in DLBCL with IG::MYC, but variable in those with non-IG::MYC including MYC/BCL6-fusion. Translocation breakpoint analyses of 8 cases by TLC-based NGS showed no obvious genomic configuration that enables MYC transactivation in 3 of the 4 cases with non-IG::MYC, while a typical promoter substitution or IGH super enhancer juxtaposition in the remaining cases. The findings potentially explain variable MYC expression in DLBCL with MYC translocation, and also bear practical implications in its routine assessment.

The 5th edition of the World Health Organization Classification of mature lymphoid and stromal tumors - an overview and update.

The purpose of this review is to give an overview on the conceptual framework and major developments of the upcoming 5th edition of the World Health Organization (WHO) Classification of Haematolymphoid tumours (WHO-HAEM5) and to highlight the most significant changes made in WHO-HAEM5 compared with the revised 4th edition (WHO-HAEM4R) of lymphoid and stromal neoplasms. The changes from the revised 4th edition include the reorganization of entities by means of a hierarchical system that is realized throughout the 5th edition of the WHO classification of tumors of all organ systems, a modification of nomenclature for some entities, the refinement of diagnostic criteria or subtypes, deletion of certain entities, and introduction of new entities. For the first time, tumor-like lesions, mesenchymal lesions specific to lymph node and spleen, and germline predisposition syndromes associated with the lymphoid neoplasms are included in the classification.

Relapses in early-stage follicular lymphoma frequently develop via a divergent evolution from their clonally related precursor cells.

Follicular lymphoma (FL) develops through a stepwise acquisition of cooperative genetic changes with t(14;18)(q32;q21)/IGH::BCL2 occurring early at the pre-B stage of B-cell development. Patients with FL typically show an indolent clinical course, remitting and relapsing with the eventual development of resistance to treatments. Interestingly, the majority of transformed FL do not progress directly from FL but originate from their clonally related lymphoma precursor (CLP) cells. To examine whether such divergent tumour evolution also underpins the relapses in patients with early-stage FL, we investigated by targeted next-generation sequencing 13 cases (stage I = 9, stage II = 4), who showed complete remission (mean: 5 years; range: 1-11.5 years) following local radiotherapy but subsequently relapsed (≥2 in 5). A clonal relationship between the diagnostic FL and relapses was confirmed in 11 cases. In six cases, common and distinct variants were seen between the paired diagnostic and relapsed lymphomas, indicating their divergent evolution from a CLP. In two cases, different B-cell clones were involved in the diagnostic and relapsed lymphomas, including one case involving two different BCL2 translocations. In the remaining five cases, the relapsed lymphoma developed via a linear progression (n = 4) or a mixed evolutionary path (n = 1). These findings may bear important implications in the routine diagnosis and management of relapsed FL. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Divergent evolution of metachronous follicular lymphoma and extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue from a common precursor.

The translocation t(14;18)(q32:q21)/IGH::BCL2 occurs at the pre-B stage of B-cell development in the bone marrow and is insufficient for malignant transformation, although it leads to the formation of in situ follicular B-cell neoplasia (ISFN). Despite that, the translocation is the genetic hallmark of follicular lymphoma (FL), it occurs infrequently in metachronous/synchronous lymphomas, including extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (EMZL), mantle cell lymphoma, and Hodgkin's lymphoma. In each of these scenarios, the two lymphomas often appear to be clonally related by analyses of IGH::BCL2 and/or rearranged IG genes. However, it remains largely unknown whether one lymphoma originates from the other or they develop independently. We studied five cases of metachronous EMZL and FL. In four cases, the two lymphomas were clonally related, as shown by identical IGH::BCL2 and/or rearranged IG genes or shared mutations. There were common and unique mutations between the paired EMZL and FL, indicating that they developed independently from a common premalignant cell population, harbouring IGH::BCL2 in three cases. Furthermore, case 1 presented with three metachronous FLs, and all of them originated from a common precursor cell population via divergent evolution. Our findings highlight the multi-malignant potential of IGH::BCL2-positive B-cells. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Clinical relevance of molecular aspects in extranodal marginal zone lymphoma: a critical appraisal.

Extranodal marginal zone B-cell lymphoma of the mucosa-associated lymphoid tissue (MALT lymphoma) is among the more common types of lymphoma accounting for up to 8% of newly diagnosed lymphoma cases. As opposed to other B-cell lymphomas, however, no predominant genetic hallmark has been defined in MALT lymphoma, but different localizations appear to be affected by different, sometimes distinct changes. Nonetheless, a high proportion of these genetic changes reported in MALT lymphomas dysregulate the pathways leading to NF-kB activation. t(11;18)(q21;q21)/BIRC3::MALT1 appears to be MALT lymphoma specific and is found in 24% of gastric and 40% of pulmonary MALT lymphomas. The translocation is associated with more disseminated disease in gastric MALT lymphoma and is found in a large percentage of patients whose lymphoma is unresponsive to antibiotic eradication of Helicobacter pylori. In addition to t(11;18)(q21;q21), nuclear expression of BCL10 or NF-kB appears to be highly associated with lymphoma cell survival independence of H. pylori-mediated stimulations. Antibiotic eradication, however, is the recommended therapy of choice irrespective of genetic findings, and molecular analysis is not required before initiation of therapy. The influence of genetic translocations including t(11;18)(q21;q21) on systemic therapies, however, is less clearly defined. While small series have shown no influence on the outcome for treatment with the anti-CD20 antibody rituximab (R) or treatment with cladribine (2-CdA), conflicting data have been reported for alkylating agents, especially chlorambucil and the combination of R + chlorambucil. None of other genetic changes seen in MALT lymphoma to date has discernible value in routine clinical applications, but recent data suggest that changes in TNFAIP3(A20), KMTD2 and CARD11 might be associated with response to Bruton kinase inhibitors.

Differential Efficacy From the Addition of Bortezomib to R-CHOP in Diffuse Large B-Cell Lymphoma According to the Molecular Subgroup in the REMoDL-B Study With a 5-Year Follow-Up.

Clinical trials frequently include multiple end points that mature at different times. The initial report, typically based on the primary end point, may be published when key planned co-primary or secondary analyses are not yet available. Clinical Trial Updates provide an opportunity to disseminate additional results from studies, published in JCO or elsewhere, for which the primary end point has already been reported.The REMoDL-B phase III adaptive trial compared rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone (R-CHOP) versus R-CHOP + bortezomib (RB-CHOP) in patients with diffuse large B-cell lymphoma (DLBCL), stratified by molecular subtype. Primary analysis at a median follow-up of 30 months found no effect of bortezomib on progression-free survival (PFS) or overall survival (OS). Retrospective analysis using a gene expression-based classifier identified a molecular high-grade (MHG) group with worse outcomes. We present an updated analysis for patients successfully classified by the gene expression profile (GEP). Eligible patients were age older than 18 years with untreated DLBCL, fit enough for full-dose chemotherapy, and with adequate biopsies for GEP. Of 1,077 patients registered, 801 were identified with Activated B-Cell (ABC), Germinal Center B-cell, or MHG lymphoma. At a median follow-up of 64 months, there was no overall benefit of bortezomib on PFS or OS (5-year PFS hazard ratio [HR], 0.81; P = .085; OS HR, 0.86; P = .32). However, improved PFS and OS were seen in ABC lymphomas after RB-CHOP: 5-year OS 67% with R-CHOP versus 80% with RB-CHOP (HR, 0.58; 95% CI, 0.35 to 0.95; P = .032). Five-year PFS was higher in MHG lymphomas: 29% versus 55% (HR, 0.46; 95% CI, 0.26 to 0.84). Patients with ABC and MHG DLBCL may benefit from the addition of bortezomib to R-CHOP in initial therapy.

The 5th edition of the World Health Organization Classification of Haematolymphoid Tumours: Lymphoid Neoplasms.

We herein present an overview of the upcoming 5th edition of the World Health Organization Classification of Haematolymphoid Tumours focussing on lymphoid neoplasms. Myeloid and histiocytic neoplasms will be presented in a separate accompanying article. Besides listing the entities of the classification, we highlight and explain changes from the revised 4th edition. These include reorganization of entities by a hierarchical system as is adopted throughout the 5th edition of the WHO classification of tumours of all organ systems, modification of nomenclature for some entities, revision of diagnostic criteria or subtypes, deletion of certain entities, and introduction of new entities, as well as inclusion of tumour-like lesions, mesenchymal lesions specific to lymph node and spleen, and germline predisposition syndromes associated with the lymphoid neoplasms.

CCR6 activation links innate immune responses to mucosa-associated lymphoid tissue lymphoma development.

The genesis of extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT) is driven by oncogenic co-operation among immunological stimulations and acquired genetic changes. We previously identified recurrent CCR6 mutations in MALT lymphoma, with majority predicted to result in truncated proteins lacking the phosphorylation motif important for receptor desensitization. Functional consequences of these mutational changes, the molecular mechanisms of CCR6 activation and how this receptor signaling contributes to MALT lymphoma development remain to be investigated. In the present study, we demonstrated that these mutations impaired CCR6 receptor internalization and were activating changes, being more potent in apoptosis resistance, malignant transformation, migration and intracellular signaling, particularly in the presence of the ligands CCL20, HBD2 (human b defensin 2) and HD5 (human a defensin 5). CCR6 was highly expressed in malignant B cells irrespective of the lymphoma sites. HBD2 and CCL20 were constitutively expressed by the duct epithelial cells of salivary glands, and also those involved in lymphoepithelial lesions (LEL) in salivary gland MALT lymphoma. While in the gastric setting, HBD2, and HD5, to a less extent CCL20, were highly expressed in epithelial cells of pyloric and intestinal metaplasia respectively including those involved in LEL, which are adaptive responses to chronic Helicobacter pylori infection. These findings suggest that CCR6 signaling is most likely active in MALT lymphoma, independent of its mutation status. The observations explain why the emergence of malignant B cells and their clonal expansion in MALT lymphoma are typically around LEL, linking the innate immune responses to lymphoma genesis.

Parallel evolution of two distinct lymphoid proliferations in clonal haematopoiesis.

AIMS: Angioimmunoblastic T-cell lymphoma (AITL) is genetically characterized by TET2 and DNMT3A mutations occurring in haematopoietic progenitor cells, and late events (e.g. the RHOA-G17V mutation) associated with malignant transformation. As TET2/DNMT3A-mutated progenitor cells can differentiate into multilineage progenies and give rise to both AITL and myeloid neoplasms, they may also have the potential to lead to other metachronous/synchronous neoplasms. We report two cases showing parallel evolution of two distinct potentially neoplastic lymphoid proliferations from a common mutated haematopoietic progenitor cell population. METHODS AND RESULTS: Both cases presented with generalized lymphadenopathy. In case 1 (a 67-year-old female), an initial lymph node (LN) biopsy was dismissed as reactive, but a repeat biopsy showed a nodal marginal zone lymphoma (NMZL)-like proliferation with an increase in the number of T-follicular helper (TFH) cells. Immunohistochemistry, and clonality and mutational analyses by targeted sequencing of both whole tissue sections and microdissected NMZL-like lesions, demonstrated a clonal B-cell proliferation that harboured the BRAF-G469R mutation and shared TET2 and DNMT3A mutations with an underlying RHOA-G17V-mutant TFH proliferation. Review of the original LN biopsy showed histological and immunophenotypic features of AITL. In case 2 (a 66-year-old male), cytotoxic T-cell lymphoma with an increase in the number of Epstein-Barr virus-positive large B cells was diagnosed on initial biopsy. On review together with the relapsed biopsy, we identified an additional occult neoplastic TFH proliferation/smouldering AITL. Both T-cell proliferations shared TET2 and DNMT3A mutations while RHOA-G17V was confined to the smouldering AITL. CONCLUSIONS: In addition to demonstrating diagnostic challenges, these cases expand the potential of clonal haematopoiesis in the development of different lineage neoplastic proliferations.

Early detection of T-cell lymphoma with T follicular helper phenotype by RHOA mutation analysis.

Angioimmunoblastic T-cell lymphoma (AITL) and peripheral T-cell lymphoma with T follicular helper phenotype (PTCL-TFH) are a group of complex clinicopathological entities that originate from T follicular helper cells and share a similar mutation profile. Their diagnosis is often a challenge, particularly at an early stage, because of a lack of specific histological and immunophenotypic features, paucity of neoplastic T cells and prominent polymorphous infiltrate. We investigated whether the lymphoma-associated RHOA Gly17Val (c.50G>T) mutation, occurring in 60% of cases, is present in the early "reactive" lesions, and whether mutation analysis could help to advance the early diagnosis of lymphoma. The RHOA mutation was detected by quantitative polymerase chain reaction with a locked nucleic acid probe specific to the mutation, and a further peptide nucleic acid clamp oligonucleotide to suppress the amplification of the wild-type allele. The quantitative polymerase chain reaction assay was highly sensitive and specific, detecting RHOA Gly17Val at an allele frequency of 0.03%, but not other changes in Gly17, nor in 61 controls. Among the 37 cases of AITL and PTCL-TFH investigated, RHOA Gly17Val was detected in 62.2% (23/37) of which 19 had multiple biopsies including preceding biopsies in ten and follow-up biopsies in 11 cases. RHOA Gly17Val was present in each of these preceding or follow-up biopsies including 18 specimens that showed no evidence of lymphoma by combined histological, immunophenotypic and clonality analyses. The mutation was seen in biopsies 0-26.5 months (mean 7.87 months) prior to the lymphoma diagnosis. Our results show that RHOA Gly17Val mutation analysis is valuable in the early detection of AITL and PTCL-TFH.

GPR34 activation potentially bridges lymphoepithelial lesions to genesis of salivary gland MALT lymphoma.

GPR34 translocation and mutation are specifically associated with salivary gland MALT lymphoma (SG-MALT-lymphoma). The majority of GPR34 mutations are clustered in its C-terminus, resulting in truncated proteins lacking the phosphorylation motif important for receptor desensitization. It is unclear why GPR34 genetic changes associate with SG-MALT-lymphoma and how these mutations contribute to the development of lymphoma. We generated isogenic Flp-InTRex293 cell lines that stably expressed a single copy of GPR34 or its various mutants and performed a range of in vitro assays. We found that the GPR34 Q340X truncation, but not the R84H and D151A mutants, conferred a significantly increased resistance to apoptosis and greater transforming potential than the GPR34 wild type. The GPR34 truncation mutant had a significantly delayed internalization compared with the wild type after ligand (lysophosphatidylserine) stimulation. Among the 9 signaling pathways examined, the GPR34 Q340X truncation, and to a lesser extent the D151A mutant, significantly activated CRE, NF-κB, and AP1 reporter activities, particularly in the presence of ligand stimulation. We further described the enhanced activities of phospholipase-A1/2 in the culture supernatant of Flp-InTRex293 cells that expressed the GPR34 Q340X mutant, as well as their potential to catalyze the synthesis of lysophosphatidylserine from phosphatidylserine. Importantly, phospholipase-A1 was abundantly expressed in the duct epithelium of salivary glands and those involved in lymphoepithelial lesions (LELs). Our findings advocate a model of paracrine stimulation of malignant B cells via GPR34, in which phospholipase A is released by LELs and hydrolyzes the phosphatidylserine exposed on apoptotic cells, generating lysophosphatidylserine, the ligand for GPR34. Thus, GPR34 activation potentially bridges LELs to genesis of SG-MALT-lymphoma.