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The 2022 European Association for Haematopathology/Society for Hematopathology lymphoma workshop session on cavity-based lymphomas included sixty-eight cases in seven sections. The disease entities discussed include primary effusion lymphomas (PEL), extracavitary primary effusion lymphomas and confounding entities (ECPEL), HHV8-negative B-lineage lymphomas-effusion based (EBV-negative, EBV-positive, and plasmablastic types), diffuse large B-cell lymphoma associated with chronic inflammation, fibrin-associated diffuse large B-cell lymphoma (FA-DLBCL), breast implant-associated anaplastic large cell lymphoma (BIA-ALCL), and other lymphomas presenting as an effusion. All entities above are discussed; however, three are delved into greater detail given the challenges with classification: ECPEL, HHV8-negative effusion-based lymphomas, and FA-DLBCL. Cases exemplifying the diagnostic difficulty in differentiating ECPEL from HHV8-positive diffuse large B-cell lymphoma and germinotropic lymphoproliferative disorder were discussed. The more recently recognized effusion-based HHV8-negative large B-cell lymphoma is explored, with several cases submitted raising the question if this subset should be carved out as a specific entity, and if so, what should be the refining diagnostic criteria. Case submissions to the FA-DLBCL section yielded one of the largest case series to date, including classic cases, cases furthering the discussion on disease sites and prognosis, as well as novel concepts to be considered in this entity. The 2022 EA4HP/SH workshop cases allowed for further confirmation of the characteristics of some of the more historically accepted cavity-based lymphomas, as well as further inquiry and debate on relatively new or evolving entities., (© 2023. The Author(s).)

Cytotoxic peripheral T-cell lymphomas and EBV-positive T/NK-cell lymphoproliferative diseases were discussed at the 2022 European Association for Haematopathology/Society for Hematopathology lymphoma workshop held in Florence, Italy. This session focused on (i) primary nodal EBV-positive T and NK-cell lymphomas (primary nodal-EBV-TNKL), (ii) extranodal EBV-positive T/NK lymphoproliferative diseases (LPD) in children and adults, (iii) cytotoxic peripheral T-cell lymphomas, NOS (cPTCL-NOS), EBV-negative, and (iv) miscellaneous cases. Primary nodal-EBV-TNKL is a newly recognized entity which is rare, aggressive, and associated with underlying immune deficiency/immune dysregulation. All cases presented with lymphadenopathy but some demonstrated involvement of tonsil/Waldeyer's ring and extranodal sites. The majority of tumors are of T-cell lineage, and the most frequent mutations involve the epigenetic modifier genes, such as TET2 and DNMT3A, and JAK-STAT genes. A spectrum of EBV-positive T/NK LPD involving extranodal sites were discussed and highlight the diagnostic challenge with primary nodal-EBV-TNKL when these extranodal EBV-positive T/NK LPD cases demonstrate predominant nodal disease either at presentation or during disease progression from chronic active EBV disease. The majority of cPTCL-NOS demonstrated the TBX21 phenotype. Some cases had a background of immunosuppression or immune dysregulation. Interestingly, an unexpected association of cPTCL-NOS, EBV-positive and negative, with TFH lymphomas/LPDs was observed in the workshop cases. Similar to a published literature, the genetic landscape of cPTCL-NOS from the workshop showed frequent mutations in epigenetic modifiers, including TET2 and DNMT3A, suggesting a role of clonal hematopoiesis in the disease pathogenesis., (© 2023. The Author(s).)

Session 3 of the lymphoma workshop of the XXI joint meeting of the European Association for Haematopathology and the Society for Hematopathology took place in Florence, Italy, on September 22, 2022. The topics of this session were splenic and nodal marginal zone lymphomas, transformation in marginal zone lymphomas, and pediatric nodal marginal zone lymphomas and their differential diagnosis as well as related entities. Forty-two cases in these categories were submitted to the workshop, including splenic lymphomas (marginal zone and diffuse red pulp lymphomas), transformed marginal zone lymphomas (splenic and nodal), nodal marginal zone lymphomas with increased TFH-cells, and pediatric nodal marginal zone lymphomas. The case review highlighted some of the principal problems in the diagnosis of marginal zone lymphomas, including the difficulties in the distinction between splenic marginal zone lymphoma, splenic diffuse red pulp lymphoma, and hairy cell leukemia variant/splenic B-cell lymphoma with prominent nucleoli which requires integration of clinical features, immunophenotype, and morphology in blood, bone marrow, and spleen; cases of marginal zone lymphoma with markedly increased TFH-cells, simulating a T-cell lymphoma, where molecular studies (clonality and mutation detection) can help to establish the final diagnosis; the criteria for transformation of marginal zone lymphomas, which are still unclear and might require the integration of morphological and molecular data; the concept of an overlapping spectrum between pediatric nodal marginal zone lymphoma and pediatric-type follicular lymphoma; and the distinction between pediatric nodal marginal zone lymphoma and "atypical" marginal zone hyperplasia, where molecular studies are mandatory to correctly classify cases., (© 2023. The Author(s).)

Emerging entities and molecular subgroups in large B-cell lymphomas (LBCLs) were discussed during the 2022 European Association for Haematopathology/Society for Hematopathology workshop in Florence, Italy. This session focused on newly recognized diseases and their diagnostic challenges. High-grade/large B-cell lymphoma with 11q aberration (HG/LBCL-11q) is defined by chromosome 11q-gains and telomeric loss. FISH analysis is recommended for the diagnosis. HG/LBCL-11q can occur in the setting of immunodeficiency, including ataxia-telangiectasia, and predominates in children. The morphological spectrum of these cases is broader than previously thought with often Burkitt-like morphology and coarse apoptotic bodies. It has a Burkitt-like immunophenotype (CD10+, BCL6+, BCL2-) but MYC expression is weak or negative, lacks MYC rearrangement, and is in contrast to Burkitt lymphoma 50% of the cases express LMO2. LBCL with IRF4 rearrangement (LBCL-IRF4) occurs mainly in the pediatric population but also in adults. LBCL-IRF4 has an excellent prognosis, with distinguishing molecular findings. IRF4 rearrangements, although characteristic of this entity, are not specific and can be found in association with other chromosomal translocations in other large B-cell lymphomas. Other molecular subgroups discussed included primary bone diffuse large B-cell lymphoma (PB-DLBCL), which has distinctive clinical presentation and molecular findings, and B-acute lymphoblastic leukemia (B-ALL) with IGH::MYC translocation recently segregated from Burkitt lymphoma with TdT expression. This latter disorder has molecular features of precursor B-cells, often tetrasomy 1q and recurrent NRAS and KRAS mutations. In this report, novel findings, recommendations for diagnosis, open questions, and diagnostic challenges raised by the cases submitted to the workshop will be discussed., (© 2023. The Author(s).)

Follicular helper T-cell lymphomas (TFH lymphomas) were discussed in session V of the lymphoma workshop of the European Association for Haematopathology (EA4HP)/Society for Hematopathology (SH) 2022 meeting in Florence, Italy. The session focused on the morphologic spectrum of TFH lymphoma, including its three subtypes: angioimmunoblastic-type (AITL), follicular-type, and not otherwise specified (NOS). The submitted cases encompassed classic examples of TFH lymphoma and unusual cases such as those with early or indolent presentations, associated B-cell proliferations, or Hodgkin/Reed-Sternberg-like cells. The relationship between TFH lymphoma and clonal hematopoiesis was highlighted by several cases documenting divergent evolution of myeloid neoplasm and AITL from shared clonal mutations. The distinction between TFH lymphoma and peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS), was stressed, and many challenging examples were presented. Various cases highlighted the difficulties of differentiating TFH lymphoma from other established types of lymphoma and reactive conditions. Cutaneous T-cell lymphoma expressing TFH markers, particularly when resulting in lymph node involvement, should be distinguished from TFH lymphomas. Additional immunophenotyping and next-generation sequencing studies were performed on various cases in this session, highlighting the importance of these technologies to our current understanding and classification of TFH lymphomas., (© 2023. The Author(s).)

With the progressive lengthening of the average age of the population, especially in some countries such as Italy, vaccination of the elderly is a fixed point on which most of the public health efforts are concentrating as epidemic infectious diseases, especially those of the winter, have a major impact on the progression of severe disease, hospitalization, and death. The protection of the elderly against acute infectious diseases should not only limit mortality but also have a positive impact on the fragility of these people in terms of less disability and fewer care needs. However, vaccination of the elderly population differs in efficacy and safety compared to that of other population categories since aging and the consequent loss of efficiency of the immune system lead to a reduction in the immunogenicity of vaccines without achieving a lasting antibody coverage. There are various strategies to avoid the failure of immunization by vaccines such as resorting to supplementary doses with adjuvant vaccines, increasing the dosage of the antigen used, or choosing to inoculate the serum relying on various routes of administration of the vaccine. Vaccination in the elderly is also an important factor in light of growing antibiotic resistance because it can indirectly contribute to combating antibiotic resistance, reducing theoretically the use of those agents. Furthermore, vaccination in old age reduces mortality from infectious diseases preventable with vaccines and reduces the same rate of resistance to antibiotics. Given the importance and complexity of the topic, in this review, we will deal with the main aspects of vaccination in the elderly and how it can influence mortality and healthcare costs, especially in those countries where population aging is more evident. Therefore, we conducted a systematic literature search in PubMed to identify all types of studies published up to 31 May 2023 that examined the association between vaccination and the elderly. Data extraction and quality assessment were conducted by two reviewers (PC and TC) who independently extracted the following data and assessed the quality of each study.

Epstein-Barr virus (EBV), defined as a group I carcinogen by the World Health Organization (WHO), is present in the tumour cells of patients with different forms of B-cell lymphoma, including Burkitt lymphoma, Hodgkin lymphoma, post-transplant lymphoproliferative disorders, and, most recently, diffuse large B-cell lymphoma (DLBCL). Understanding how EBV contributes to the development of these different types of B-cell lymphoma has not only provided fundamental insights into the underlying mechanisms of viral oncogenesis, but has also highlighted potential new therapeutic opportunities. In this review, we describe the effects of EBV infection in normal B-cells and we address the germinal centre model of infection and how this can lead to lymphoma in some instances. We then explore the recent reclassification of EBV+ DLBCL as an established entity in the WHO fifth edition and ICC 2022 classifications, emphasising the unique nature of this entity. To that end, we also explore the unique genetic background of this entity and briefly discuss the potential role of the tumour microenvironment in lymphomagenesis and disease progression. Despite the recent progress in elucidating the mechanisms of this malignancy, much work remains to be done to improve patient stratification, treatment strategies, and outcomes.

Burkitt lymphoma (BL) is an aggressive form of B cell lymphoma that can affect children and adults. The study of BL led to the identification of the first recurrent chromosomal aberration in lymphoma, t(8;14)(q24;q32), and subsequent discovery of the central role of MYC and Epstein-Barr virus (EBV) in tumorigenesis. Most patients with BL are cured with chemotherapy but those with relapsed or refractory disease usually die of lymphoma. Historically, endemic BL, non-endemic sporadic BL and the immunodeficiency-associated BL have been recognized, but differentiation of these epidemiological variants is confounded by the frequency of EBV positivity. Subtyping into EBV + and EBV - BL might better describe the biological heterogeneity of the disease. Phenotypically resembling germinal centre B cells, all types of BL are characterized by dysregulation of MYC due to enhancer activation via juxtaposition with one of the three immunoglobulin loci. Additional molecular changes commonly affect B cell receptor and sphingosine-1-phosphate signalling, proliferation, survival and SWI-SNF chromatin remodelling. BL is diagnosed on the basis of morphology and high expression of MYC. BL can be effectively treated in children and adolescents with short durations of high dose-intensity multiagent chemotherapy regimens. Adults are more susceptible to toxic effects but are effectively treated with chemotherapy, including modified versions of paediatric regimens. The outcomes in patients with BL are good in high-income countries with low mortality and few late effects, but in low-income and middle-income countries, BL is diagnosed late and is usually treated with less-effective regimens affecting the overall good outcomes in patients with this lymphoma., (© 2022. Springer Nature Limited.)

We herein present an overview of the upcoming 5 th 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 4 th edition. These include reorganization of entities by a hierarchical system as is adopted throughout the 5 th 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., (© 2022. The Author(s).)

(1) Purpose: To describe first-line pharmacotherapy and overall survival in non-resectable non-small cell lung cancer (nrNSCLC) patients by gender. (2) Methods: Incident cases of nrNSCLC recorded between 2009 and 2019 (cohort entry) in the pathology registry of the regional administrative healthcare database of Tuscany were identified. Records of antineoplastic therapies delivered up to 4 months following cohort entry were classified as chemotherapy, target therapies, immunotherapies, and undefined monoclonal antibodies. First-line treatment and survival of patients receiving drug treatment was described. Analyses were stratified according to histology, gender, and cohort entry year. (3) Results: 4393 incident cases of nrNSCLC were included. Women with non-squamous-NSCLC received target-therapy more frequently than men (14.9% vs. 6.5%). Immunotherapy incidence of use varied between 3.8% (2017) and 9.1% (2019). The 2-year survival rate increased over time: for non-squamous-NSCLC, it was 22.3% (2009-2011) and 30.6% (2018-2019), while for squamous-NSCLC, it was 13.5% and 22.5%, respectively. After multivariate analysis, a low reduction in mortality risk in 2018-2019 vs. 2009-2011 was found (non-squamous: HR: 0.95 CI95%: 0.92-0.98; squamous: HR: 0.94 CI95%: 0.90-0.98). Among non-squamous NSCLC, median survival was longer in women than in men (389 vs. 276 days). (4) Conclusion: In light of sex-related biomolecular differences, among non-squamous NSCLC, women received target-therapy more frequently than men. Survival seemed to slightly improve over the study period for both histologies, despite a poor reduction in mortality risk was still observed.

Background: Human herpesvirus-8 (HHV8) is a lymphotropic virus associated with different lymphoproliferative disorders, including primary effusion lymphoma (PEL), multicentric Castleman's disease (MCD), diffuse large B-cell lymphomas, not otherwise specified, and the rare entity known as germinotropic lymphoproliferative disorder (GLPD). In PELs and GLPD the neoplastic cells also contain Epstein-Barr virus (EBV). In addition, occasional cases with atypical and overlapping features among these entities have been recognised, suggesting that the spectrum of the HHV8-related lesions may not be fully characterised., Aims: Here, we report two cases of lymphoproliferative disorder associated with HHV8 and EBV that further expand the spectrum of HHV8/EBV-positive lymphoproliferative disease., Methods and Results: Case 1 represented HHV8/EBV-positive extracavitary nodal PEL followed by pleural PEL. The striking characteristic of this case was the almost focal and intrasinusoidal localisation of the neoplastic cells and the association with Castleman's disease features. In the second case, we found the entire spectrum of HHV8-related disorders, i.e. MCD, GLPD, and PEL, coexisting in the same lymph node, underlining the variability, possible overlap and evolution among these entities. Both cases were well analysed with immunohistochemistry, determination of the EBV latency programme, and molecular analysis for clonality of immnoglobulin genes. In both patients, the disease followed an unexpected indolent course, both being still alive after 8 and 12 months, respectively., Conclusion: Our findings represent further evidence of the overlap among HHV8/EBV-positive lymphoproliferative disorders, and underline a grey zone that requires further study; they further confirm the experimental evidence that lytic EBV replication influences HHV8-related tumorigenesis., (© 2021 The Authors. Histopathology published by John Wiley & Sons Ltd.)

Altered cellular energetic metabolism has recently emerged as important feature of neoplastic cells. Indeed, interfering with cancer cell metabolism might represent a suitable therapeutic strategy. In this study, we aimed to assess glucose metabolism activation in human lymphomas and evaluate how metformin can exert its action on lymphoma cells. We studied a large series of human lymphomas (N = 252) and an in vitro model of Burkitt lymphoma (BL) cells. We combined molecular biology techniques, including global gene expression profiling (GEP) analysis, quantitative PCR (qPCR) and Western blotting, and biochemical assays, aimed to assess pentose phosphate pathway, tricarboxylic acid (TCA) cycle, and aerobic glycolysis rates. We found that glucose metabolism is overall enhanced in most lymphoma subtypes, based on gene expression profiling (GEP), with general shift to aerobic glycolysis. By contrast, normal B cells only showed an overall increase in glucose usage during germinal center transition. Interestingly, not only highly proliferating aggressive lymphomas but also indolent ones, like marginal zone lymphomas, showed the phenomenon. Consistently, genes involved in glycolysis were confirmed to be overexpressed in BL cells by qPCR. Biochemical assays showed that while aerobic glycolysis is increased, TCA cycle is reduced. Finally, we showed that metformin can induce cell death in BL cells by stressing cellular metabolism through the induction of GLUT1, PKM2, and LDHA. In conclusion, we unveiled glucose metabolism abnormalities in human lymphomas and characterized the mechanism of action of metformin in Burkitt lymphoma model., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Bagaloni, Visani, Biagiotti, Ruzzo, Navari, Etebari, Mundo, Granai, Lazzi, Isidori, Loscocco, Li, Leoncini, Visani, Magnani and Piccaluga.)

EBV is a direct causative agent in around 1.5% of all cancers. The oncogenic properties of EBV are related to its ability to activate processes needed for cellular proliferation, survival, migration, and immune evasion. The EBV latency program is required for the immortalization of infected B cells and involves the expression of non-coding RNAs (ncRNAs), including viral microRNAs. These ncRNAs have different functions that contribute to virus persistence in the asymptomatic host and to the development of EBV-associated cancers. In this review, we discuss the function and potential clinical utility of EBV microRNAs and other ncRNAs in EBV-associated malignancies. This review is not intended to be comprehensive, but rather to provide examples of the importance of ncRNAs.

Background: The present study aimed to classify lymphoid neoplasms according to the latest World Health Organization (WHO) classification and outlining the distribution in Nigeria of different entities. Additionally, the study describes the prevalence of lymphoid neoplasms associated with Epstein-Barr virus (EBV) infection in the Nigerian population., Methods: We collected 152 formalin-fixed paraffin-embedded (FFPE) tissues diagnosed as lymphoma from 2008 to 2018, coming from three different institutions located within three geopolitical zone in Nigeria. These institutions included the University College Hospital (UCH), Ibadan, Oyo State, the Enugu State University of Science and Technology Teaching Hospital (ESUTH), Enugu, Enugu State, and the Meena Histopathology and Cytology Laboratory (MHCL), Jos, Plateau State., Results: From the total 152 cases retrieved, 50 were excluded due to insufficient tissue materials or inconclusive antigen reactivity. We confirmed 66 (64.7%) cases as lymphomas out of the remaining 102 FFPE with a male to female ratio of 2:1 and a mean age of 44.4 years. Ten entities were identified, and of these, chronic lymphocytic leukemia (CLL) was the most prevalent category (34.8%). For the diffuse large B-cell lymphomas not otherwise specified (DLBCL, NOS), the germinal centre B-cell type was the most common (71.4%). Ten lymphoma cases (15.2%) were positive for Epstein-Barr virus (EBV), most of which were Hodgkin lymphoma (HL). CLL was common in the Hausa ethnic group, HL in the Yoruba ethnic group, while the Igbo ethnic group had an equal distribution of CLL, HL, and DLBCL diagnosis., Conclusion: Although the distribution of lymphomas in Nigeria shares some similarities with those of other countries, we described distinct features of some subtypes of lymphomas. Also, the study underscores the need for a more precise diagnosis and classification of lymphoid neoplasms in Nigeria using the latest WHO classification.

Background: The global active prevalence of migraines is approximately 14.7%. Oxygen therapy may reduce the use of non-steroidal anti-inflammatory drugs (NSAIDs) which often have various negative side effects. The purpose of this systematic review is to analyze the literature on the efficacy of high flow oxygen for the management of headache disorders, compared to placebo treatment., Methods: Studies were identified by PubMed, Web of Science and Scopus database from 1980 to the 30 October 2020. The search included the following terms: "oxygen therapy" and "headache" and "migraine". Studies were included if high flow oxygen was used in the treatment of headache disorders. All selected studies were qualitatively analyzed., Results: Our literature search identified 71 studies, of which 65 were discarded and 6 were included in the meta-analysis. The random effect model did not show a pooled significant resolution of headache disorders (OR 2.08 (95% CI 0.92-4.70), p < 0.0001) in the oxygen therapy group compared to the placebo group., Conclusion: In our systematic review of six studies, there were no significant differences between high flow oxygen and placebo treatment groups.

Conventional/targeted chemotherapies and ionizing radiation (IR) are being used both as monotherapies and in combination for the treatment of epithelial ovarian cancer (EOC). Several studies show that these therapies might favor oncogenic signaling and impede anti-tumor responses. MiR-200c is considered a master regulator of EOC-related oncogenes. In this study, we sought to investigate if chemotherapy and IR could influence the expression of miR-200c-3p and its target genes, like the immune checkpoint PD-L1 and other oncogenes in a cohort of EOC patients' biopsies. Indeed, PD-L1 expression was induced, while miR-200c-3p was significantly reduced in these biopsies post-therapy. The effect of miR-200c-3p target genes was assessed in miR-200c transfected SKOV3 cells untreated and treated with olaparib and IR alone. Under all experimental conditions, miR-200c-3p concomitantly reduced PD-L1, c-Myc and β-catenin expression and sensitized ovarian cancer cells to olaparib and irradiation. In silico analyses further confirmed the anti-correlation between miR-200c-3p with c-Myc and β-catenin in 46 OC cell lines and showed that a higher miR-200c-3p expression associates with a less tumorigenic microenvironment. These findings provide new insights into how miR-200c-3p could be used to hold in check the adverse effects of conventional chemotherapy, targeted therapy and radiation therapy, and offer a novel therapeutic strategy for EOC.

The Epstein-Barr virus (EBV) is linked to various B-cell lymphomas, including Burkitt lymphoma (BL), classical Hodgkin lymphoma (cHL) and diffuse large B-cell lymphoma (DLBCL) at frequencies ranging, by routine techniques, from 5 to 10% of cases in DLBCL to >95% in endemic BL. Using higher-sensitivity methods, we recently detected EBV traces in a few EBV-negative BL cases, possibly suggesting a "hit-and-run" mechanism. Here, we used routine and higher-sensitivity methods (qPCR and ddPCR for conserved EBV genomic regions and miRNAs on microdissected tumor cells; EBNA1 mRNA In situ detection by RNAscope) to assess EBV infection in a larger lymphoma cohort [19 BL, 34 DLBCL, 44 cHL, 50 follicular lymphomas (FL), 10 T-lymphoblastic lymphomas (T-LL), 20 hairy cell leukemias (HCL), 10 mantle cell lymphomas (MCL)], as well as in several lymphoma cell lines (9 cHL and 6 BL). qPCR, ddPCR, and RNAscope consistently documented the presence of multiple EBV nucleic acids in rare tumor cells of several cases EBV-negative by conventional methods that all belonged to lymphoma entities clearly related to EBV (BL, 6/9 cases; cHL, 16/32 cases; DLBCL, 11/30 cases), in contrast to fewer cases (3/47 cases) of FL (where the role of EBV is more elusive) and no cases (0/40) of control lymphomas unrelated to EBV (HCL, T-LL, MCL). Similarly, we revealed traces of EBV infection in 4/5 BL and 6/7 HL cell lines otherwise conventionally classified as EBV negative. Interestingly, additional EBV-positive cases (1 DLBCL, 2 cHL) relapsed as EBV-negative by routine methods while showing EBNA1 expression in rare tumor cells by RNAscope. The relapse specimens were clonally identical to their onset biopsies, indicating that the lymphoma clone can largely loose the EBV genome over time but traces of EBV infection are still detectable by high-sensitivity methods. We suggest EBV may contribute to lymphoma pathogenesis more widely than currently acknowledged.

Fifty-five cases of t(14;18)- follicular lymphoma (FL) were genetically characterized by targeted sequencing and copy number (CN) arrays. t(14;18)- FL predominated in women (M/F 1:2); patients often presented during early clinical stages (71%), and had excellent prognoses. Overall, t(14;18)- FL displayed CN alterations (CNAs) and gene mutations carried by conventional t(14;18)+ FL (cFL), but with different frequencies. The most frequently mutated gene was STAT6 (57%) followed by CREBBP (49%), TNFRSF14 (39%), and KMT2D (27%). t(14;18)- FL showed significantly more STAT6 mutations and lacked MYD88, NOTCH2, MEF2B, and MAP2K1 mutations compared with cFL, nodal marginal zone lymphoma (NMZL), and pediatric-type FL (PTFL). We identified 2 molecular clusters. Cluster A was characterized by TNFRSF14 mutations/1p36 alterations (96%) and frequent mutations in epigenetic regulators, with recurrent loss of 6q21-24 sharing many features with cFL. Cluster B showed few genetic alterations; however, a subgroup with STAT6 mutations concurrent with CREBBP mutations/16p alterations without TNFRSF14 and EZH2 mutations was noted (65%). These 2 molecular clusters did not distinguish cases by inguinal localization, growth pattern, or presence of STAT6 mutations. BCL6 rearrangements were demonstrated in 10 of 45 (22%) cases and did not cluster together. Cases with predominantly inguinal presentation (20 of 50; 40%) had a higher frequency of diffuse growth pattern, STAT6 mutations, CD23 expression, and a lower number of CNAs, in comparison with noninguinal cases (5.1 vs 9.1 alterations per case; P < .05). STAT6 mutations showed a positive correlation with CD23 expression (P < .001). In summary, t(14;18)- FL is genetically a heterogeneous disorder with features that differ from cFL, NMZL, and PTFL.

Introduction: Microenvironment has a prognostic influence in diffuse large B-cell lymphoma (DLBCL); among its components, tumor-associated macrophages (TAM) play a leading role. TAM can be classified into M1 (anti-tumor) and M2 (pro-tumor). Another prognostic factor could be represented by lymphocyte-to-monocyte and neutrophil-to-lymphocyte ratio (LMR and NLR)., Objective: The aim of the study is to evaluate the prognostic impact of M1 and M2 TAM subtypes, LMR and NLR in DLBCL., Methods: We analyzed 37 consecutive patients between 2009 and 2013. Out of 37 patients, 28/37 (75.6%) received R-CHOP/CHOP-like regimens, 9/37 (24.4%) less intensive therapies. Immunohistochemistry was performed with antibodies against CD68 and CD163. We divided our cohort into 2 categories according to the Steidl score. TAM who coexpressed CD68 and CD163 were considered as M2. For LMR and NLR we used previously published cut-offs of 2.71 and 2.81., Results: CR rate was 70.3%; we did not record a significant correlation between CD68+ TAM, CD163+ TAM, CD68+/CD163+ TAM, LMR, NLR and CR. We observed a reduced PFS in patients with IPI ≥ 2 and high M2 TAM expression and a trend between higher expression of CD68+ TAM and improved PFS., Conclusion: M2 TAM could have a prognostic role for IPI ≥ 2 DLBCL patients receiving R-CHOP, which thus warrants further investigation., Competing Interests: None., (AJBR Copyright © 2020.)

The precise B cell of origin and molecular pathogenesis of nodal marginal zone lymphoma (NMZL) remain poorly defined. To date, due to the rarity of NMZL, the vast majority of already-published studies have been conducted on a limited number of samples and the technical approach to analyze the immunoglobulin genes was of amplifying rearranged variable region genes with the classical direct sequencing of the PCR products followed by cloning. Here, we studied the B cell Ig heavy-chain repertoires by next-generation sequencing (NGS) in 30 NMZL cases. Most of the cases were mutated (20/28; 71.5%) with homologies to the respective germ line genes ranging from 85 to 97, 83%, whereas 8/28 (28.5%) were unmutated. In addition, our results show that NMZL cases have a biased usage of specific immunoglobulin heavy-chain variable (IGHV) region genes. Moreover, we documented intraclonal diversity in all (100%) of the mutated cases and ongoing somatic hypermutations (SHM) have been confirmed by hundreds of reads. We analyzed the mutational pattern to detect and quantify antigen selection pressure and we found a positive selection in 4 cases, whereas in the remaining cases there was an unspecific stimulation. Finally, the disease-specific survival and the progression-free survival were significantly different between cases with mutated and unmutated IGHV genes, pointing out mutational status as a possible new biomarker in NMZL.

This error was caused due to the author's oversight and this does not change the views or the results presented in the manuscript.

Purpose: The purpose of this study was to investigate the involvement of CD93 and Multimerin-2 in three choroidal neovascularization (CNV) models and to evaluate their contribution in the neovascular progression of age-related macular degeneration (AMD)., Methods: Choroidal neovascular membranes collected during surgery from AMD patients were analyzed by microscopy methods. Laser-induced CNV mouse models and choroid sprouting assays (CSAs) were carried out using the CD93 knockout mouse model. An original ex vivo CSA of vascular angiogenesis, employing choroid tissues isolated from human donors, was developed., Results: In contrast to healthy choroid endothelium, hyperproliferative choroidal endothelial cells (ECs) of AMD patients expressed high levels of CD93, and Multimerin-2 was abundantly deposited along the choroidal neovasculature. CD93 knockout mice showed a significant reduced neovascularization after laser photocoagulation, and their choroidal ECs displayed a decreased ability to produce sprouts in ex vivo angiogenesis assays. Moreover, the presence of an antibody able to hamper the CD93/Multimerin-2 interaction reduced vascular sprouting in the human CSA., Conclusions: Our results demonstrate that CD93 and its interaction with Multimerin-2 play an important role in pathological vascularization of the choroid, disclosing new possibilities for therapeutic intervention to neovascular AMD.

Background: The Tumor Microenviroment (TME) is a complex milieu that is increasingly recognized as a key factor in multiple stages of disease progression and responses to therapy as well as escape from immune surveillance. However, the precise contribution of specific immune effector and immune suppressor components of the TME in Burkitt lymphoma (BL) remains poorly understood., Methods: In this paper, we applied the computational algorithm CIBERSORT to Gene Expression Profiling (GEP) datasets of 40 BL samples to draw a map of immune and stromal components of TME. Furthermore, by multiple immunohistochemistry (IHC) and multispectral immunofluorescence (IF), we investigated the TME of additional series of 40 BL cases to evaluate the role of the Programmed Death-1 and Programmed Death Ligand-1 (PD-1/PD-L1) immune checkpoint axis., Results: Our results indicate that M2 polarized macrophages are the most prominent TME component in BL. In addition, we investigated the correlation between PD-L1 and latent membrane protein-2A (LMP2A) expression on tumour cells, highlighting a subgroup of BL cases characterized by a non-canonical latency program of EBV with an activated PD-L1 pathway., Conclusion: In conclusion, our study analysed the TME in BL and identified a tolerogenic immune signature highlighting new potential therapeutic targets., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2020.)

An accurate diagnosis of clinically distinct subgroups of aggressive mature B cell lymphomas is crucial for the choice of proper treatment. Presently, precise recognition of these disorders relies on the combination of morphological, immunophenotypical, and cytogenetic/molecular features. The diagnostic workup in such situations implies the application of costly and time-consuming analyses, which are not always required, since an intensified treatment option is reasonably reserved to fit patients. The Italian Group of Haematopathology proposes herein a practical algorithm for the diagnosis of aggressive mature B cell lymphomas based on a stepwise approach, aimed to select cases deserving molecular analysis, in order to optimize time and resources still assuring the optimal management for any patient.

The Shc family adaptor p66Shc acts as a negative regulator of proliferative and survival signals triggered by the B-cell receptor and, by enhancing the production of reactive oxygen species, promotes oxidative stress-dependent apoptosis. Additionally, p66Shc controls the expression and function of chemokine receptors that regulate lymphocyte traffic. Chronic lymphocytic leukemia cells have a p66Shc expression defect which contributes to their extended survival and correlates with poor prognosis. We analyzed the impact of p66Shc ablation on disease severity and progression in the Eμ-TCL1 mouse model of chronic lymphocytic leukemia. We showed that Eμ-TCL1/p66Shc -/- mice developed an aggressive disease that had an earlier onset, occurred at a higher incidence and led to earlier death compared to that in Eμ-TCL1 mice. Eμ-TCL1/p66Shc -/- mice displayed substantial leukemic cell accumulation in both nodal and extranodal sites. The target organ selectivity correlated with upregulation of chemokine receptors whose ligands are expressed therein. This also applied to chronic lymphocytic leukemia cells, where chemokine receptor expression and extent of organ infiltration were found to correlate inversely with these cells' level of p66Shc expression. p66Shc expression declined with disease progression in Eμ-TCL1 mice and could be restored by treatment with the Bruton tyrosine kinase inhibitor ibrutinib. Our results highlight p66Shc deficiency as an important factor in the progression and severity of chronic lymphocytic leukemia and underscore p66Shc expression as a relevant therapeutic target., (Copyright© 2019 Ferrata Storti Foundation.)

The histone modifier lysine (K)-specific demethylase 2B (KDM2B) plays a role in the differentiation of hematopoietic cells, and its expression appears to be deregulated in certain cancers of hematological and lymphoid origins. We have previously found that the KDM2B gene is differentially methylated in cell lines derived from Epstein-Barr virus (EBV)-associated endemic Burkitt lymphoma (eBL) compared with that in EBV-negative sporadic Burkitt lymphoma-derived cells. However, whether KDM2B plays a role in eBL development has not been previously investigated. Oncogenic viruses have been shown to hijack the host cell epigenome to complete their life cycle and to promote the transformation process by perturbing cell chromatin organization. Here, we investigated whether EBV alters KDM2B levels to enable its life cycle and promote B-cell transformation. We show that infection of B cells with EBV leads to downregulation of KDM2B levels. We also show that LMP1, one of the main EBV transforming proteins, induces increased DNMT1 recruitment to the KDM2B gene and augments its methylation. By altering KDM2B levels and performing chromatin immunoprecipitation in EBV-infected B cells, we show that KDM2B is recruited to the EBV gene promoters and inhibits their expression. Furthermore, forced KDM2B expression in immortalized B cells led to altered mRNA levels of some differentiation-related genes. Our data show that EBV deregulates KDM2B levels through an epigenetic mechanism and provide evidence for a role of KDM2B in regulating virus and host cell gene expression, warranting further investigations to assess the role of KDM2B in the process of EBV-mediated lymphomagenesis. IMPORTANCE In Africa, Epstein-Barr virus infection is associated with endemic Burkitt lymphoma, a pediatric cancer. The molecular events leading to its development are poorly understood compared with those leading to sporadic Burkitt lymphoma. In a previous study, by analyzing the DNA methylation changes in endemic compared with sporadic Burkitt lymphoma cell lines, we identified several differential methylated genomic positions in the proximity of genes with a potential role in cancer, and among them was the KDM2B gene. KDM2B encodes a histone H3 demethylase already shown to be involved in some hematological disorders. However, whether KDM2B plays a role in the development of Epstein-Barr virus-mediated lymphoma has not been investigated before. In this study, we show that Epstein-Barr virus deregulates KDM2B expression and describe the underlying mechanisms. We also reveal a role of the demethylase in controlling viral and B-cell gene expression, thus highlighting a novel interaction between the virus and the cellular epigenome., (Copyright © 2019 American Society for Microbiology.)

The new recently described provisional lymphoma category Burkitt-like lymphoma with 11q aberration comprises cases similar to Burkitt lymphoma (BL) on morphological, immunophenotypic and gene-expression levels but lacking the IG- MYC translocation. They are characterized by a peculiar imbalance pattern on chromosome 11, but the landscape of mutations is not yet described. Thus, we investigated 15 MYC -negative Burkitt-like lymphoma with 11q aberration (mnBLL,11q,) cases by copy-number analysis and whole-exome sequencing. We refined the regions of 11q imbalance and identified the INO80 complex-associated gene NFRKB as a positional candidate in 11q24.3. Next to recurrent gains in 12q13.11-q24.32 and 7q34-qter as well as losses in 13q32.3-q34, we identified 47 genes recurrently affected by protein-changing mutations (each ≥3 of 15 cases). Strikingly, we did not detect recurrent mutations in genes of the ID3-TCF3 axis or the SWI/SNF complex that are frequently altered in BL, or in genes frequently mutated in germinal center-derived B-cell lymphomas like KMT2D or CREBBP An exception is GNA13 , which was mutated in 7 of 15 cases. We conclude that the genomic landscape of mnBLL,11q, differs from that of BL both at the chromosomal and mutational levels. Our findings implicate that mnBLL,11q, is a lymphoma category distinct from BL at the molecular level., (© 2019 by The American Society of Hematology.)

Granulysin is a cytolytic protein expressed in cytotoxic T and natural killer (NK) cells. Abnormal serum levels of granulysin in lymphomas with NK and cytotoxic phenotype have been shown to correlate with tumour progression. In this study, we investigated the expression pattern of granulysin in routine sections of normal and reactive lymphoid tissues as well as in a large series of lymphomas. In normal tissues, granulysin labelled a small population of cells that double immunostaining revealed to belong to the pool of cytotoxic T/NK cells. Among lymphoid neoplasms, the highest expression of granulysin (71%) was found in extranodal NK/T cell lymphomas of nasal type (ENKTL). To note is that 29% of ENKTLs, which were negative for one or more of classical cytotoxic markers strongly expressed granulysin. Furthermore, expression of granulysin was observed in rare cases of T cell lymphomas with a cytotoxic phenotype (i.e. ALK-negative anaplastic large cell lymphoma (26%), enteropathy-associated T cell lymphoma (12%) and peripheral T cell lymphoma, NOS (4%)). None of the investigated non-Hodgkin B cell lymphomas, Hodgkin lymphoma and plasma cell myeloma were granulysin positive. The results suggest granulysin as a novel marker for a subset of cytotoxic NK cell derived malignancies and its usefulness is highlighted in those ENKTLs that lack expression of other cytotoxic markers but retain granulysin expression.

Tumor metabolism has been the object of several studies in the past, leading to the pivotal observation of a consistent shift toward aerobic glycolysis (so-called Warburg effect). More recently, several additional investigations proved that tumor metabolism is profoundly affected during tumorigenesis, including glucose, lipid and amino-acid metabolism. It is noticeable that metabolic reprogramming can represent a suitable therapeutic target in many cancer types. Epstein-Barr virus (EBV) was the first virus linked with cancer in humans when Burkitt lymphoma (BL) was described. Besides other well-known effects, it was recently demonstrated that EBV can induce significant modification in cell metabolism, which may lead or contribute to neoplastic transformation of human cells. Similarly, virus-induced tumorigenesis is characterized by relevant metabolic abnormalities directly induced by the oncoviruses. In this article, the authors critically review the most recent literature concerning EBV-induced metabolism alterations in lymphomas.

Epstein-Barr virus (EBV) is a human γ-herpesvirus implicated in several human malignancies, including a wide range of lymphomas. Several molecules encoded by EBV in its latent state are believed to be related to EBV-induced lymphomagenesis, among which microRNAs-small RNAs with a posttranscriptional regulating role-are of great importance. The genome of EBV encodes 44 mature microRNAs belonging to two different classes, including BamHI-A rightward transcript ( BART ) and Bam HI fragment H rightward open reading frame 1 ( BHRF1 ), with different expression levels in different EBV latency types. These microRNAs might contribute to the pathogenetic effects exerted by EBV through targeting self mRNAs and host mRNAs and interfering with several important cellular mechanisms such as immunosurveillance, cell proliferation, and apoptosis. In addition, EBV microRNAs can regulate the surrounding microenvironment of the infected cells through exosomal transportation. Moreover, these small molecules could be potentially used as molecular markers. In this review, we try to present an updated and extensive view of the role of EBV-encoded miRNAs in human lymphomas., Competing Interests: The authors declare no conflict of interest.

Objectives: Chronic lymphocytic leukemia (CLL) is the most common type of leukemia in Western populations, being rarer in Asian and African people. It has been suggested that patients with CLL from Africa might have a more aggressive disease compared with white patients. In this study, we aimed to identify genetic factors that may account for this difference., Methods: We analyzed immunoglobulin heavy chain (IGH) genes' mutational status by performing next-generation sequencing in 25 Senegalese and 50 Italian patients with CLL., Results: We found that Senegalese patients more frequently had adverse prognostic factors and an unmutated profile. Furthermore, we documented that IGHV1 (IGHV1-69), IGHD3, and IGHJ6 were significantly more frequent in Senegalese patients, whereas IGHV3-30 was common and limited to the Italian cohort. Stereotyped receptors commonly detected in the white population were not recorded in our Senegalese series., Conclusions: The different IGH repertoire we observed in the Senegalese cohort may reflect the diverse genetic and microenvironmental (ie, polymicrobial stimulation) background., (© American Society for Clinical Pathology, 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com)

Plasmablastic lymphoma (PBL) is an aggressive lymphoma, often arising in the context of immunodeficiency and associated with Epstein-Barr virus (EBV) infection. The most frequently detected genetic alteration is the deregulation of MYC gene through the translocation - t(8;14)(q24;q32). The diagnosis of PBL is often challenging because it has an overlap in morphology, immunophenotype, cytogenetics and virus association with other lymphomas and plasma cell neoplasms; further, its molecular basis remains elusive. In the present study we aimed to better define the possible contribution of EBV infection as well as miRNA deregulation in PBL pathogenesis. We studied 23 cases of PBL, 19 Burkitt lymphomas (BL), and 17 extra-medullary plasmacytoma (EMPC). We used qPCR and immunohistochemistry to assess EBV latency patterns, while micro-RNA (miRNA) profiling was performed by next generation sequencing (Illumina) and validated by qPCR. Our analysis revealed a non-canonical EBV latency program with the partial expression of some proteins characterizing latency II and the activation of an abortive lytic cycle. Moreover, we identified miRNA signatures discriminating PBL from BL and EMPC. Interestingly, based on the miRNA profile, PBL appeared constituted by two discrete subgroups more similar to either BL or EMPC, respectively. This pattern was confirmed in an independent set of cases studied by qPCR and corresponded to different clinico-pathological features in the two groups, including HIV infection, MYC rearrangement and disease localization. In conclusion, we uncovered for the first time 1) an atypical EBV latency program in PBL; 2) a miRNA signature distinguishing PBL from the closest malignant counterparts; 3) the molecular basis of PBL heterogeneity., Competing Interests: CONFLICTS OF INTEREST The authors declare no competing interests.

Epstein-Barr virus (EBV) is a gammaherpesvirus linked to a number of lymphoid and epithelial malignancies, including Burkitt lymphoma (BL) in which its frequency ranges from 30% in sporadic cases to 100% in the endemic ones. The possible contribution of EBV to BL pathogenesis is largely unknown. It has been suggested that EBV may be associated with all of the cases, including those diagnosed as EBV negative by a mechanism of hit-and-run . Early during oncogenesis, viral genes are essential for initiating disease. Progressively, viral genome is lost to escape the immune system and host mutations accumulate in proto-oncogenic cell. The main problem with the hit-and-run hypothesis is the lack of evidence in primary tumors. The routine methods applied to detect the virus [i.e., immunohistochemistry and EBV-encoded RNAs (EBER) in situ hybridization (ISH)] have a low specificity and accuracy. The aim of this study was to identify the most suitable method to detect EBV infection in pathology samples by applying conventional and non-conventional methods (i.e., EBV-microRNAs detection and EBV viral load measurement). We investigated a total of 10 cases and we found that all the samples ( n = 6) diagnosed as EBV negative by immunohistochemistry and EBER-ISH demonstrated the presence of EBV-microRNAs and EBV genome. This points at the possibility that EBV might have contributed to lymphomagenesis in all our patients, and propose microRNAs detection as the most specific and sensitive tool to recognize EBV vestiges. It is worth noting that our data would have considerable implications for EBV-related diseases control. By using anti-EBV vaccines, one could potentially prevent also some cancers less suspected of a viral origin because of viral genome loss.

In Hodgkin Lymphoma (HL), about 20% of patients still have relapsed/refractory disease and late toxic effects rate continue to rise with time. 'Early FDG-PET' and tissue macrophage infiltration (TAM) emerged as powerful prognostic predictors. The primary endpoint was to investigate the prognostic role of both early FDG-PET and TAM; the secondary endpoint was to test if early FDG-PET positivity could correlate with high TAM score. A cohort of 200 HL patients was analysed. Induction treatment plan consisted of two to six courses of ABVD and, if indicated, involved field radiation therapy. All patients repeated CT scan and FDG-PET after two cycles and after the completion of therapy. TAM in diagnostic specimens was determined by immunohistochemistry with a monoclonal antibody (anti-CD68 KP1). Overall, early FDG-PET was negative in 163 patients (81.5%) and positive in 37 patients (18.5%), showing a significant correlation with the achievement of CR (p < 0.0001). After a median follow-up of 40 months, progression free survival (PFS) was significantly better for PET negative patients (p < 0.0001). CD68 expression was low, intermediate or high in 26 (13%), 100 (50%) and 74 (37%) cases, without difference in the distribution between responders and non-responders. PFS analysis showed no significant difference in any score group. TAM score did not show any correlation with early FDG-PET result. This study confirms that early FDG-PET has a high prognostic power, while TAM score does not seem to influence the outcome; in contrast to our original hypothesis, it does not correlate with FDG-PET assessment. Copyright © 2015 John Wiley & Sons, Ltd., (Copyright © 2015 John Wiley & Sons, Ltd.)

Background: In 2000, an Italian non-governmental organisation (NGO) began a 9-year project to establish a surgical pathology laboratory at the Bugando Medical Centre (BMC) in Mwanza, Tanzania, a country with a low Human Development Index (HDI), and as of 2009, the laboratory was operating autonomously. The present survey aims to evaluate the reproducibility of histological and cytological diagnoses assigned in the laboratory's early years of autonomous activity. We selected a random sample of 196 histological and cytological diagnoses issued in 2010-2011 at the BMC surgical pathology laboratory. The corresponding samples were sent to Italy for review by Italian senior pathologists, who were blinded to the local results. Samples were classified into four diagnostic categories: malignant, benign, inflammatory, and suspicious. The two-observer kappa-statistic for categorised (qualitative) data was then calculated to measure diagnostic concordance between the local Tanzanian pathologists and Italian senior pathologists. The k-Cohen was calculated for concordance in the overall study sample. Concordance and discordance rates were also stratified by subset: general adult, paediatric/adolescent, and lymphoproliferative histopathological diagnoses; fluid and fine needle aspiration (FNA) cytological diagnoses; and PAP tests. Discordance was also categorised by the corresponding hypothetical clinical implications: high, intermediate, and not significant., Results: Overall concordance was 85.2% (167 of 196 diagnoses), with a k-Cohen of 0.7691 ( P  = 0.0000). Very high concordance was observed in the subsets of adult general pathological diagnoses (90%) and paediatric/adolescent pathological diagnoses (91.18%). Concordance in the subset of PAP tests was 75%, and for fluid/FNA cytological diagnoses it was 56.52%. Concordance among 12 histological subtypes of lymphoma was 75.86%, with substantial discordance observed in the diagnosis of Burkitt lymphoma (five cases diagnosed by Italian pathologists versus 2 by local pathologists). The overall proportion of discordance with high hypothetical clinical implications was 6.1% (12 diagnoses)., Conclusion: This blind review of diagnoses assigned in Tanzania, a country with low HDI, and in Italy, a country with a very high HDI, seemed to be a sensitive and effective method to identify areas of potential error and may represent a reference point for future, more detailed quality control processes or audits of surgical pathology services located in limited-resource regions.