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Extranodal NK/T-cell lymphoma (ENKTCL) is an Epstein-Barr virus (EBV)-related neoplasm with male dominance and a poor prognosis. A better understanding of the genetic alterations and their functional roles in ENKTCL could help improve patient stratification and treatments. In this study, we performed a comprehensive genetic analysis of 178 ENKTCL cases to delineate the landscape of mutations, copy number alterations (CNA), and structural variations, identifying 34 driver genes including six previously unappreciated ones, namely, HLA-B, HLA-C, ROBO1, CD58, POT1, and MAP2K1. Among them, CD274 (24%) was the most frequently altered, followed by TP53 (20%), CDKN2A (19%), ARID1A (15%), HLA-A (15%), BCOR (14%), and MSN (14%). Chromosome X losses were the most common arm-level CNAs in females (∼40%), and alterations of four X-linked driver genes (MSN, BCOR, DDX3X, and KDM6A) were more frequent in males and females harboring chromosome X losses. Among X-linked drivers, MSN was the most recurrently altered, and its expression was lost in approximately one-third of cases using immunohistochemical analysis. Functional studies of human cell lines showed that MSN disruption promoted cell proliferation and NF-κB activation. Moreover, MSN inactivation increased sensitivity to NF-κB inhibition in vitro and in vivo. In addition, recurrent deletions were observed at the origin of replication in the EBV genome (6%). Finally, by integrating the 34 drivers and 19 significant arm-level CNAs, nonnegative matrix factorization and consensus clustering identified two molecular groups with different genetic features and prognoses irrespective of clinical prognostic factors. Together, these findings could help improve diagnostic and therapeutic strategies in ENKTCL. Significance: Integrative genetic analyses and functional studies in extranodal NK/T-cell lymphoma identify frequent disruptions of X-linked drivers, reveal prognostic molecular subgroups, and uncover recurrent MSN alterations that confer sensitivity to NF-κB inhibition., (©2024 American Association for Cancer Research.)

Granulomatous disease is a serious complication of common variable immunodeficiency (CVID-GD) that occurs in 8-22% of these patients and can mimic sarcoidosis, with which it shares certain clinical, biological, and radiological features. However, few studies to date have compared the two pathologies immunologically and histologically. Therefore, we analyzed the immunological-histological findings for different tissue samples from ten patients with CVID-GD and compared them to those of biopsy-proven sarcoidosis. Specifically, we wanted to know whether or not the signaling abnormalities observed in sarcoidosis granulomas are also present in CVID-GD. Morphological differences were found between CVID-GD histology and classical sarcoidosis: mainly, the former's notable lymphoid hyperplasia associated with granulomas not observed in the latter. All CVID-GD involved organs contained several follicular helper-T (TFH) cells within the granulomatosis, while those cells were inconstantly and more weakly expressed in sarcoidosis. Moreover, CVID and sarcoidosis granulomas expressed the phosphorylated-signal transducer and activator of transcription (pSTAT)1 and pSTAT3 factors, regardless of the organ studied and without any significant difference between entities. Our results suggest that the macrophage-activation mechanism in CVID resembles that of sarcoidosis, thereby suggesting that Janus kinase (JAK)-STAT-pathway blockade might be useful in currently difficult-to-treat CVID-GD., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Purpose: About 25% of patients with common variable immunodeficiency disease (CVID) have splenomegaly, necessitating sometimes splenectomy whom consequences on the immunological profile of CVID patients have never been studied. We analyzed 11 CVID patients' comprehensive blood immune cell phenotypes pre- and post-splenectomy., Methods: Flow cytometry analyses of immune cell populations., Results: Among 89 CVID cohort patients, 41 with splenomegaly, splenomegaly was strongly associated with granulomatous disease, autoimmune disorders, lymphoid hyperplasia, and/or portal hypertension. CVID patients with splenomegaly have significant peripheral lymphopenia (p = 0.001), and significantly fewer peripheral class-switched memory B cells (smBs) (p = 0.001), CD4 + T lymphocytes (p = 0.001), NK (p = 0.0001) and dendritic cells (p ≤ 0.01), and significantly more circulating CD4 + and CD8 + (p = 0.00001) T cell subset activation (p = 0.00005), than CVID patients without splenomegaly. Examination of splenectomy impact on circulating lymphocyte subset distributions demonstrated the drastically enhanced total circulating lymphocyte count post-splenectomy, predominantly B lymphocytes and CD8 + T cells. However, splenectomy did not change B cell distribution, with smBs remaining persistently low, in contrast to complete inversion of the circulating T cell composition, with reversal of the CD4 + /CD8 + ratio suggesting that amplification of the CD8 + T cell compartment is a CVID characteristic in patients with splenomegaly. Our results highlight this CD8 + amplification in CVID-splenomegaly patients that might be explained by a homing effect to the spleen and/or possible chronic virus replication, which in turn could induce T cell expansions., Conclusion: Splenectomizing CVID patients with splenomegaly restores the absolute circulating lymphocyte count, suggesting that the decreased T cell count in the presence of splenomegaly cannot be used as an exclusive criterion for combined immunodeficiency., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

ALK-negative anaplastic large cell lymphoma (ALCL) comprises subgroups harboring rearrangements of DUSP22 (DUSP22- R) or TP63 (TP63-R). Two studies reported 90% and 40% 5-year overall survival (OS) rates in 21 and 12 DUSP22-R/TP63- not rearranged (NR) patients, respectively, making the prognostic impact of DUSP22-R unclear. Here, 104 newly diagnosed ALK-negative ALCL patients (including 37 from first-line clinical trials) from the LYSA TENOMIC database were analyzed by break-apart fluorescence in situ hybridization assays for DUSP22-R and TP63-R. There were 47/104 (45%) DUSP22-R and 2/93 (2%) TP63-R cases, including one DUSP22-R/TP63-R case. DUSP22-R tumors more frequently showed CD3 expression (62% vs. 35%, P=0.01), and less commonly a cytotoxic phenotype (27% vs. 82%; P<0.001). At diagnosis, DUSP22- R ALCL patients more frequently had bone involvement (32% vs. 13%, P=0.03). The patient with DUSP22-R/TP63-R ALCL had a rapidly fatal outcome. After a median follow-up of 4.9 years, 5-year progression-free survival (PFS) and OS rates of 84 patients without TP63-R treated with curative-intent anthracycline-based chemotherapy were 41% and 53%, respectively. According to DUSP22 status, 5-year PFS was 57% for 39 DUSP22-R versus 26% for 45 triple-negative (DUSP22-NR/TP63-NR/ALK-negative) patients (P=0.001). The corresponding 5-year OS rates were 65% and 41%, respectively (P=0.07). In multivariate analysis, performance status and DUSP22 status significantly affected PFS, and distinguished four risk groups, with 4-year PFS and OS ranging from 17% to 73% and 21% to 77%, respectively. Performance status but not DUSP22 status influenced OS. The use of brentuximab vedotin in relapsed/refractory patients improved OS independently of DUSP22 status. Our findings support the biological and clinical distinctiveness of DUSP22- R ALK-negative ALCL. Its relevance to outcome in patients receiving frontline brentuximab vedotin remains to be determined.

Mature tertiary lymphoid structures (mTLSs) are organized lymphoid structures containing B lymphocytes admixed to CD23+ follicular dendritic cells. Their presence has been linked to improved survival and sensitivity to immune checkpoint inhibitors in several cancers, emerging as a promising pancancer biomarker. However, the requirements for any biomarker are clear methodology, proven feasibility, and reliability. In 357 patients' samples, we studied tertiary lymphoid structures (TLSs) parameters using multiplex immunofluorescence (mIF), hematoxylin-eosin-saffron (HES) staining, double CD20/CD23 staining, and single CD23 immunohistochemistry. The cohort included carcinomas (n = 211) and sarcomas (n = 146), gathering biopsies (n = 170), and surgical specimens (n = 187). mTLSs were defined as TLSs containing either a visible germinal center on HES staining or CD23+ follicular dendritic cells. Focusing on 40 TLSs assessed using mIF, double CD20/CD23 staining was less sensitive than mIF to assess maturity in 27.5% (n = 11/40) but was rescued by single CD23 staining in 90.9% (n = 10/11). In 97 patients, several samples (n = 240) were reviewed to characterize TLS distribution. The likelihood of finding TLSs in surgical material was 6.1 higher than in biopsy and 2.0 higher in primary samples than in metastasis after adjustment with a type of sample. Interrater agreement rates over 4 examiners were 0.65 (Fleiss kappa, 95% CI [0.46, 0.90]) for the presence of TLS and 0.90 for maturity (95% CI [0.83, 0.99]). In this study, we propose a standardized method to screen mTLSs in cancer samples using HES staining and immunohistochemistry that can be applied to all specimens., (Copyright © 2023 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

Distinguishing between follicular lymphoma (FL) and nodal marginal zone lymphoma (NMZL) can be difficult when morphologic and phenotypic features are unusual and characteristic cytogenetic rearrangements are absent. We evaluated the diagnostic contribution of ancillary techniques-including fluorescence in situ hybridization (FISH)-detected 1p36 deletion; reverse-transcriptase, multiplex, ligation-dependent probe amplification (RT-MLPA); and next-generation sequencing (NGS)-for tumors that remain unclassified according to standard criteria. After review, 50 CD5-negative small B-cell lymphoid neoplasms without BCL2 and BCL6 FISH rearrangements were diagnosed as FLs (n = 27), NMZLs (n = 5), or unclassified (n = 18) based on the 2016 World Health Organization Classification of Tumours of Haematopoietic and Lymphoid Tissues. FISH helped identify the 1p36 deletion in 3 FLs and 1 unclassified tumor. Most classified FLs had an RT-MLPA germinal center B-cell (GCB) signature (93%) or were noncontributive (7%). Classified NMZLs had an RT-MLPA activated B-cell signature (20%), had an unassigned signature (40%), or were noncontributive (40%). Among unclassified tumors, the RT-MLPA GCB signature was associated with mutations most commonly found in FLs (CREBBP, EZH2, STAT6, and/or TNFRSF14) (90%). An RT-MLPA-detected GCB signature and/or NGS-detected gene mutations were considered as FL identifiers for 13 tumors. An activated B-cell signature or NOTCH2 mutation supported NMZL diagnosis in 3 tumors. Combining the RT-MLPA and NGS findings successfully discriminated 89% of unclassified tumors in favor of one or the other diagnosis. NGS-detected mutations may be of therapeutic interest. Herein, we detected 3 EZH2 and 8 CREBBP mutations that might be eligible for targeted therapies., (Copyright © 2022 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

Monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL) is a rare aggressive T-cell lymphoma most reported in Asia. We performed a comprehensive clinical, pathological and genomic study of 71 European MEITL patients (36 males, 35 females, median age 67 years). The majority presented with gastrointestinal involvement and had emergency surgery, and 40% had stage IV disease. The tumors were morphologically classified into two groups: typical (58%) and atypical (i.e., non-monomorphic or with necrosis, angiotropism or starry-sky pattern) (42%), sharing a homogeneous immunophenotypic profile (CD3+ [98%] CD4- [94%] CD5- [97%] CD7+ [97%] CD8+ [90%] CD56+ [86%] CD103+ [80%] cytotoxic marker+ [98%]) with more frequent expression of TCRgd (50%) than TCRab (32%). MYC expression (30% of cases) partly reflecting MYC gene locus alterations, correlated with non-monomorphic cytology. Almost all cases (97%) harbored deleterious mutation(s) and/or deletion of the SETD2 gene and 90% had defective H3K36 trimethylation. Other frequently mutated genes were STAT5B (57%), JAK3 (50%), TP53 (35%), JAK1 (12.5%), BCOR and ATM (11%). Both TP53 mutations and MYC expression correlated with atypical morphology. The median overall survival (OS) of 63 patients (43/63 only received chemotherapy after initial surgery) was 7.8 months. Multivariate analysis found a strong negative impact on outcome of MYC expression, TP53 mutation, STAT5B mutation and poor performance status while aberrant B-cell marker expression (20% of cases) correlated with better survival. In conclusion, MEITL is an aggressive disease with resistance to conventional therapy, predominantly characterized by driver gene alterations deregulating histone methylation and JAK/STAT signaling and encompasses genetic and morphologic variants associated with very high clinical risk.

According to expert guidelines, lymph node surgical excision is the standard of care for lymphoma diagnosis. However, core needle biopsy (CNB) has become widely accepted as part of the lymphoma diagnostic workup over the past decades. The aim of this study was to present the largest multicenter inventory of lymph nodes sampled either by CNB or surgical excision in patients with suspected lymphoma and to compare their diagnostic performance in routine pathologic practice. We reviewed 32 285 cases registered in the French Lymphopath network, which provides a systematic expert review of all lymphoma diagnoses in France, and evaluated the percentage of CNB and surgical excision cases accurately diagnosed according to the World Health Organization classification. Although CNB provided a definitive diagnosis in 92.3% and seemed to be a reliable method of investigation for most patients with suspected lymphoma, it remained less conclusive than surgical excision, which provided a definitive diagnosis in 98.1%. Discordance rates between referral and expert diagnoses were higher on CNB (23.1%) than on surgical excision (21.2%; P = .004), and referral pathologists provided more cases with unclassified lymphoma or equivocal lesion through CNB. In such cases, expert review improved the diagnostic workup by classifying ∼90% of cases, with higher efficacy on surgical excision (93.3%) than CNB (81.4%; P < 10-6). Moreover, diagnostic concordance for reactive lesions was higher on surgical excision than CNB (P = .009). Overall, although CNB accurately diagnoses lymphoma in most instances, it increases the risk of erroneous or nondefinitive conclusions. This large-scale survey also emphasizes the need for systematic expert review in cases of lymphoma suspicion, especially in those sampled by using CNB., (© 2022 by The American Society of Hematology.)

Histologically, Castleman's disease associates three subtypes: 1-the vascular hyaline (HV) subtype more often seen in unicentric forms; 2-the plasmacytic (PV) subtype, more frequently associated with the HHV8+ and idiopathic multicentric form; 3-the mixed subtype associating both HV and PV aspects that may be encountered in any type of Castleman's disease. If the diagnosis of unicentric (isolated mass) and multicentric HHV8+ Castleman's disease is easy, the diagnosis of the idiopathic multicentric form remains particularly difficult because it is at the crossroads of many other pathologies (infectious, tumoral and dysimmune), making an anatomoclinical comparison necessary. The role of the pathologist, in the context of disseminated lesions (polyadenopathy and splenomegaly), is to identify lesions that may be part of Castleman's disease, to systematically perform HHV8 testing and to perform complete phenotyping associated with molecular analysis (B and T-cell clonality) in order to rule out a lymphomatous process and certain infectious etilogies. In all cases, its role will be a warning bell and the diagnosis of Castleman's disease will be retained only after a rigorous anatomic and clinical confrontation. © 2022 Published by Elsevier Masson SAS on behalf of Société nationale française de médecine interne (SNFMI)., (Copyright © 2022 Société nationale française de médecine interne (SNFMI). Publié par Elsevier Masson SAS. Tous droits réservés.)

Clinicians are sometimes confronted with the diagnostic difficulties of the idiopathic form of Castleman's Disease (iMCD). As this review reports with demonstrative clinical cases, iMCD can mimic various serious systemic pathologies such as certain autoimmune diseases, Still's disease, POEMS syndrome, and malignant lymphoproliferations, sharing a very similar histology and identical symptoms. To make a diagnosis of iMCD, the clinician must eliminate all the pathologies mentioned above, but he must first think of it and evoke this diagnosis of rare disease before the first symptoms but also know how to evoke this diagnosis again even after several years of evolution of a disease like those mentioned above whose evolution is not favorable. © 2022 Published by Elsevier Masson SAS on behalf of Société nationale française de médecine interne (SNFMI)., (Copyright © 2022 Société nationale française de médecine interne (SNFMI). Publié par Elsevier Masson SAS. Tous droits réservés.)

Background: Intravascular large B-cell lymphoma (lVLBCL) is a very rare type of large B-cell lymphoma., Methods: We conducted a retrospective study on IVLBCL patients treated from 2000 to 2016 in LYSA cooperative group centers., Results: Sixty-five patients were identified in 23 centers. Median age at diagnosis was 69 years (range 23-92). Thirty-four patients (64%) had an IPI score >3 and 40 patients (67%) had a performance status ≥2. The most frequent extra-nodal locations were bone marrow (n = 34; 52%), central nervous system (n = 25; 39%), and skin (n = 21; 33%). Nodal involvement and endocrine system were observed in 34% (n = 22) and 18% (n = 12) of all cases, respectively. Twenty-six patients (41%) had macrophage activation syndrome. Tumor cells were frequently CD5 positive (52%) with a non-germinal center origin (86%). BCL2 was expressed in 87% of all samples analyzed (n = 20) and 43% of patients had a MYC/BCL2 double expression. Fifty-six patients were treated with a regimen of chemotherapy containing rituximab, among whom 73% reached complete remission. The median progression-free survival (PFS) and median overall survival (OS) were 29.4 months and 63.8 months, respectively. History of autoimmune disorder (Hazard ratio [HR] 3.3 [1.4-7.8]; p < 0.01), nodal involvement (HR 2.6 [1.4-5.1]; p < 0.01), lack of anthracycline (HR 0.1 [0-0.4] for use; p < 0.001), or no intensification at first-line regimen (p = 0.02) were associated with worse PFS. High-dose methotrexate use was not associated with better PFS or OS., Conclusions: Our study highlights the aggressive clinical picture of IVLBCL, in particular the frequency of macrophage activation syndrome, and the need for new therapies despite a response to R-CHOP-like regimen similar to non-intravascular diffuse large B-cell lymphomas., (© 2022 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Nodal peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS) with cytotoxic phenotype is overall rare, with most reports coming from Asia. Given its elusive pathobiology, we undertook a clinicopathological and molecular study of 54 Western patients diagnosed with PTCL, NOS expressing cytotoxic molecules, within a lymph node. More commonly males (M/F-2,6/1) with median age of 60 years were affected. Besides lymphadenopathy, 87% of patients had ≥1 involved extranodal site. High-stage disease (III-IV), International Prognostic Index >2, B symptoms, LDH level, and cytopenia(s) were observed in 92, 63, 67, 78, and 66% of cases, respectively. Ten patients had a history of B-cell malignancies, one each of myeloid neoplasm, breast or prostate cancer, and 4 others had underlying immune disorders. Most patients (70%) died, mostly of disease, with a median overall survival of 12.7 months. Immunophenotypically, the neoplastic lymphocytes were T-cell receptor (TCR) αβ + (47%), TCR-silent (44%) or TCRγδ+ (10%), commonly CD8 + (45%) or CD4-CD8- (32%). All except one had an activated cytotoxic profile, and 95% were subclassified into PTCL-TBX21 subtype based on CXCR3, TBX21, and GATA3 expression pattern. Seven patients (13%) disclosed EBER + tumor cells. Targeted DNA deep-sequencing (33 cases) and multiplex ligation-dependent reverse transcription-polymerase chain reaction assay (43 cases) identified frequent mutations in epigenetic modifiers (73%), including TET2 (61%) and DNMT3A (39%), recurrent alterations affecting the TCR (36%) and JAK/STAT (24%) signaling pathways and TP53 mutations (18%). Fusion transcripts involving VAV1 were identified in 6/43 patients (14%). Patients with nodal cytotoxic PTCL, NOS have an aggressive behavior and frequently present in a background of impaired immunity, although the association with Epstein-Barr virus is rare. The recurrent alterations in genes involved in DNA methylation together with genes related to cytokine or TCR signaling, suggest that co-operation of epigenetic modulation with cell-signaling pathways plays a critical role in the pathogeny of these lymphomas., (© 2022. The Author(s), under exclusive licence to United States & Canadian Academy of Pathology.)

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).)

Despite the impressive efficacy of chimeric antigen receptor (CAR) T-cell therapy (CART) in B-cell non-Hodgkin lymphomas, durable responses are uncommon. The histopathologic and molecular features associated with treatment failure are still largely unknown. Therefore, we have analyzed 19 sequential tumor samples from 9 patients, prior anti-CD19 CART (pre-CART) and at relapse (post-CART), using immunohistochemistry, fluorescence in situ hybridization, array comparative genomic hybridization, next-generation DNA and RNA sequencing, and genome-scale DNA methylation. The initial diagnosis was diffuse large B-cell lymphoma (n=6), double-hit high-grade B-cell lymphoma (n=1), and Burkitt lymphoma (n=2). Histopathologic features were mostly retained at relapse in 7/9 patients, except the frequent loss of 1 or several B-cell markers. The remaining 2 cases (1 diffuse large B-cell lymphoma and 1 Burkitt lymphoma) displayed a dramatic phenotypic shift in post-CART tumors, with the drastic downfall of B-cell markers and emergence of T-cell or histiocytic markers, despite the persistence of identical clonal immunoglobulin gene rearrangements. The post-CART tumor with aberrant T-cell phenotype showed reduced mRNA expression of most B-cell genes with increased methylation of their promoter. Fluorescence in situ hybridization and comparative genomic hybridization showed global stability of chromosomal alterations in all paired samples, including 17p/TP53 deletions. New pathogenic variants acquired in post-CART samples included mutations triggering the PI3K pathway (PIK3R1, PIK3R2, PIK3C2G) or associated with tumor aggressiveness (KRAS, INPP4B, SF3B1, SYNE1, TBL1XR1). These results indicate that CART-resistant B-cell non-Hodgkin lymphomas display genetic remodeling, which may result in profound dysregulation of B-cell differentiation. Acquired mutations in the PI3K and KRAS pathways suggest that some targeted therapies could be useful to overcome CART resistance., Competing Interests: Conflicts of Interest and Source of Funding: Supported in part by the Lymphoma Study Association (LYSA) and the Institut Carnot Lymphome (CALYM), the Laboratoire d’Excellence Toulouse Cancer (TOUCAN) (contract ANR11-LABX). The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

HHV-8 is an oncogenic Gammaherpesvirinae discovered in 1994 during the HIV pandemic. It is the causative agent of Kaposi's sarcoma, and is also associated with the occurrence of several aggressive B lymphoproliferative disorders. Most of them occur in an immunosuppression setting, usually due to HIV infection. Multicentric HHV8-associated Castleman's disease and KSHV Inflammatory Cytokine Syndrome (KICS) are primarily reactive entities with prominent systemic features. They illustrate the cytokinic storm induced by HHV-8 in its cell host. On the other hand, HHV-8 can drive proliferation and lymphomagenesis of its plasmablastic cell host, and is associated with a risk to develop aggressive lymphomas with plasmacytic differenciation. Primary effusion lymphoma usually localizes in body cavities and may affect other extra-nodal sites ; its prognostic is poor. Diffuse large B-cell lymphoma HHV-8, NOS affect more commonly nodes and blood and evolve from infected cell of HHV-8 associated Castleman disease. On the contrary, germinotropic lymphoproliferative disorders presents mainly as localized adenopathy with indolent course, and show polyclonality. Histology plays a key role in distinguishing these different entities and need expert reviewing, especially since they may be associated with each other. Besides lymphoproliferative disorders, HHV8 is associated with various hematological manifestations. The aim of this review is to provide an update on the presentation, diagnosis, and management of immunologic and hematologic complications associated with HHV-8., (Copyright © 2021 Société Nationale Française de Médecine Interne (SNFMI). Published by Elsevier Masson SAS. All rights reserved.)

Primary cutaneous large B-cell lymphomas (PCLBCL) represent a diagnostic challenge because they are classified as PCLBCL, leg type (PCLBCL, LT) or primary cutaneous follicle centre lymphoma, large cell (PCFCL, LC), which differ by prognosis and therapeutic requirement. Unclassified cases with discordant clinical presentations, morphologies, and immunophenotypes may be classified into the not otherwise specified (PCLBCL, NOS) category based on ancillary molecular analyses. Cell-of-origin profiling as germinal centre (GC) type or non-GC type by immunohistochemistry is not considered reproducible because of variable CD10 expression. In a series of 55 PCLBCL cases with > 80% large cells, we reported 21 PCFCL, LC cases as GC-type and 27 PCLBCL, LT as non-GC-type; 7 cases were considered PCLBCL, NOS. Here, we demonstrate the accuracy of molecular profiling of PCLBCL as GC or non-GC type using a reverse transcriptase multiplex ligation assay (RT-MLPA). RT-MLPA classified the seven PCLBCL, NOS cases in accordance with their mutational profile. An integrative principal component analysis confirmed the main criteria and the relevance of genomic profiling of PCFCL, LC as GC-derived, and PCLBCL, LT as non-GC-derived. Both the cell-of-origin classification of PCLBCL and the integrative analysis identified two clinically relevant subgroups according to overall survival, which may help to standardize PCLBCL diagnosis and patient management., Competing Interests: The authors have declared that no competing interests exist.

Purpose: The AHL2011 study (ClinicalTrials.gov identifier: NCT01358747) demonstrated that a positron emission tomography (PET)-driven de-escalation strategy after two cycles of bleomycin, etoposide, doxorubicin, cyclophosphamide, vincristine, procarbazine, and prednisone (BEACOPP) provides similar progression-free survival (PFS) and overall survival (OS) and reduces early toxicity compared with a nonmonitored standard treatment. Here, we report, with a prolonged follow-up, the final study results., Methods: Patients with advanced Hodgkin lymphoma (stage III, IV, or IIB with mediastinum/thorax ratio > 0.33 or extranodal involvement) age 16-60 years were prospectively randomly assigned between 6 × BEACOPP and a PET-driven arm after 2 × BEACOPP delivering 4 × ABVD (doxorubicin, bleomycin, vinblastine, and dacarbazine) in PET2- and 4 × BEACOPP in PET2+ patients. PET performed after four cycles of chemotherapy had to be negative to complete the planned treatment., Results: In total, 823 patients were enrolled including 413 in the standard arm and 410 in the PET-driven arm. With a 67.2-month median follow-up, 5-year PFS (87.5% v 86.7%; hazard ratio [HR] = 1.07; 95% CI, 0.74 to 1.57; P = .67) and OS (97.7% in both arms; HR = 1.012; 95% CI, 0.50 to 2.10; P = .53) were similar in both randomization arms. In the whole cohort, full interim PET assessment predicted patients' 5-year PFS (92.3% in PET2-/PET4-, 75.4% [HR = 3.26; 95% CI, 18.3 to 5.77] in PET2+/PET4- and 46.5% [HR = 12.4; 95% CI, 7.31 to 19.51] in PET4+ patients, respectively; P < .0001) independent of international prognosis score. Five-year OS was also affected by interim PET results, and PET2+/PET4- patients (93.5%; HR = 3.3; 95% CI, 1.07 to 10.1; P = .036) and PET4+ patients (91.9%; HR = 3.756; 95% CI, 1.07 to 13.18; P = .038) had a significant lower OS than PET2-/PET4- patients (98.2%). Twenty-two patients (2.7%) developed a second primary malignancy, 13 (3.2%) and 9 (2.2%) in the standard and experimental arms, respectively., Conclusion: The extended follow-up confirms the continued efficacy and favorable safety of AHL2011 PET-driven strategy, which is noninferior to standard six cycles of BEACOPP. PET4 provides additional prognostic information to PET2 and allows identifying patients with particularly poor prognosis., Competing Interests: René-Olivier CasasnovasHonoraria: Roche/Genentech, Takeda, Gilead Sciences, Bristol Myers Squibb, Merck, AbbVie, Celgene, Janssen, AmgenConsulting or Advisory Role: Roche/Genentech, Takeda, Gilead Sciences, Bristol Myers Squibb, Merck, AbbVie, Celgene, Janssen, IncyteResearch Funding: Roche/Genentech (Inst), Gilead Sciences (Inst), Takeda (Inst)Travel, Accommodations, Expenses: Roche/Genentech, Takeda, Gilead Sciences, Janssen Pauline BriceResearch Funding: Takeda, BMSTravel, Accommodations, Expenses: Roche, Amgen, AbbVie/Genentech Julien LazaroviciTravel, Accommodations, Expenses: Mundipharma Hervé GhesquieresHonoraria: Gilead Sciences, Janssen, Celgene, RocheConsulting or Advisory Role: Gilead Sciences, Celgene, Roche, MundipharmaTravel, Accommodations, Expenses: Roche, Gilead Sciences, Celgene, Takeda Aspasia StamatoullasTravel, Accommodations, Expenses: Pfizer Jehan DupuisConsulting or Advisory Role: AstraZenecaTravel, Accommodations, Expenses: Gilead Sciences Thomas GastinneHonoraria: Pfizer, Takeda, Gilead SciencesConsulting or Advisory Role: Takeda, Gilead SciencesTravel, Accommodations, Expenses: Roche, Pfizer Krimo BouabdallahHonoraria: Roche, Takeda Science Foundation, AbbVie, Kite/GileadConsulting or Advisory Role: Roche, Takeda, Kite/GileadTravel, Accommodations, Expenses: Roche, Takeda Pierre FeugierHonoraria: Roche/Genentech, Janssen, Gilead Sciences, Amgen, AbbVieConsulting or Advisory Role: Roche/Genentech, Janssen, AbbVie, Gilead Sciences, Amgen, AstraZenecaSpeakers' Bureau: Roche/Genentech, AbbVie, Amgen, Janssen, Gilead SciencesResearch Funding: Roche/Genentech, Gilead Sciences, Janssen, AbbVie, AmgenTravel, Accommodations, Expenses: Amgen, Gilead Sciences, Janssen, Roche/Genentech, AbbVie Franck MorschhauserConsulting or Advisory Role: Roche/Genentech, Gilead Sciences, Celgene, Bristol Myers Squibb, AbbVie, Epizyme, ServierSpeakers' Bureau: RocheExpert Testimony: Roche/Genentech David SibonConsulting or Advisory Role: Takeda, Iqone healthcare, Janssen, Roche, AbbVieTravel, Accommodations, Expenses: Takeda, Janssen Christophe BonnetConsulting or Advisory Role: Roche Diane DamotteResearch Funding: AstraZeneca/MedImmune Marc AndréConsulting or Advisory Role: Takeda, BMSiResearch Funding: Takeda (Inst), Roche (Inst)Travel, Accommodations, Expenses: Roche, Celgene, Gilead Sciences Michel MeignanHonoraria: RocheTravel, Accommodations, Expenses: Roche Cédric RossiConsulting or Advisory Role: Janssen, AbbVie, Roche, TakedaResearch Funding: TakedaTravel, Accommodations, Expenses: AbbVieNo other potential conflicts of interest were reported.

Although follicular lymphoma (FL) is usually graded as FL1-2, FL3A, and FL3B, some borderline cases can be observed and led us to investigate the clinicopathologic diversity of grade 3 FL (FL3). Among 2449 FL patients enrolled in Lymphoma Study Association (LYSA) trials, 1921 cases with sufficient material underwent a central pathologic review. The resulting diagnoses comprised 89.6% FL1-2 (n=1723), 7.2% FL3A (n=138), and 0.5% purely follicular FL3B (n=9). The remaining 51 unclassifiable cases (2.7%) exhibited high-grade features but did not meet WHO criteria for either FL3A or FL3B; and were considered as "unconventional" high-grade FL (FL3U). FL3U morphological pattern consisted of nodular proliferation of large cleaved cells or small-sized to medium-sized blast cells. Compared with FL3A, FL3U exhibited higher MUM1 and Ki67 expression, less BCL2 breaks and more BCL6 rearrangements, together with a higher number of cases without any BCL2, BCL6 or MYC rearrangement. FL3U harbored less frequent mutations in BCL2, KMT2D, KMT2B, and CREBBP than FL3A. MYC and BCL2 were less frequently mutated in FL3U than FL3B. Rituximab cyclophosphamide, doxorubicin, vincristine, and prednisone treated FL3U patients had a worse survival than FL1-2 patients with similar follicular lymphoma international prognostic index and treatment. These results suggest that high-grade FLs encompass a heterogeneous spectrum of tumors with variable morphology and genomic alterations, including FL3U cases that do not strictly fit WHO criteria for either FL3A or FL3B, and display a worse outcome than FL1-2. The distinction of FL3U may be useful to allow a better comprehension of high-grade FLs and to design clinical trials., Competing Interests: Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

BCR-ABL-fusion-negative myeloproliferative neoplasms (MPNs) with myelofibrosis (MF) include primary MF, post-polycythemia vera MF and post-essential thrombocythemia MF. Clonal extramedullary hematopoiesis (EMH) can occur during MPN pathogenesis. Although histopathological bone-marrow (BM) features during clonal EMH have been investigated, those of the spleen have been poorly described. We analyzed splenectomy samples from 28 patients with MF and BM samples from 20 of them. Slides were stained with hematoxylin and eosin, reticulin, and trichrome, with immunohistochemical labeling of glycophorin A, myeloperoxidase, CD61, CD34, and CD117. We also subjected splenectomy and BM samples from six patients and spleen samples from seven patients to next-generation sequencing (NGS). Megakaryocyte-rich spleen nodules (MRSNs), seen in seven of the 28 patients, were significantly associated with megakaryocyte proliferation in the spleen (p = 0.04). We devised a grading system for spleen fibrosis (SF) and found that SF was increased in 20 of 28 patients. Notably, patients with SF were more likely to have MRSNs, suggesting that megakaryocytes might participate in SF, as previously described in BM. Comparisons of spleen and BM NGS findings of six patients' specimens revealed identical mutational status in the two organs for half of the patients. We observed additional mutations in the spleen of two patients. However, the meaning of this finding remains unknown since there was a long interval between BM and spleen samplings (68 and 82 months, respectively)., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

In this retrospective study, we report 70 cases of Epstein-Barr virus (EBV)+ diffuse large B-cell lymphoma not otherwise specified (DLBCL-NOS) among 1696 DLBCL-NOS cases diagnosed between 2006 and 2019 (prevalence of 4.1%). At diagnosis, median age was 68.5 years; 79% of the cases presented with an advanced-stage disease (III-IV), 48% with extranodal lesions, and 14% with an hemophagocytic lymphohistiocytosis (HLH) (8 at diagnosis and 1 on therapy). A total of 46 cases presented a polymorphic pattern, and 21 were monomorphic. All had a non-germinal center B phenotype, with the majority of tumor cells expressing CD30 and programmed death ligand 1 (98% and 95%, respectively). Type II and III EBV latency was seen in 88% and 12% of the cases, respectively. Patients were treated with immunochemotherapy (59%) or chemotherapy (22%), and 19% received palliative care due to advanced age and altered performance status. After a median follow-up of 48 months, progression-free survival (PFS) and overall survival (OS) at 5 years were 52.7% and 54.8%, respectively. Older age (>50 years) and HLH were associated with shorter PFS and OS in multivariate analysis (PFS: hazard ratio [HR], 14.01; 95% confidence interval [CI], 2.34-83.97; and HR, 5.78; 95% CI, 2.35-14.23; OS: HR, 12.41; 95% CI, 1.65-93.53; and HR, 6.09; 95% CI, 2.42-15.30, respectively). Finally, using a control cohort of 425 EBV- DLBCL-NOS, EBV positivity was associated with a shorter OS outcome within patients >50 years (5-year OS, 53% [95% CI, 38.2-74] vs 60.8% [95% CI, 55.4-69.3], P = .038), but not in younger patients., (© 2021 by The American Society of Hematology.)

The genetic basis of peripheral T-cell lymphoma (PTCL) is complex and encompasses several recurrent fusion transcripts discovered over the past years by means of massive parallel sequencing. However, there is currently no affordable and rapid technology for their simultaneous detection in clinical samples. Herein, we developed a multiplex ligation-dependent RT-PCR-based assay, followed by high-throughput sequencing, to detect 33 known PTCL-associated fusion transcripts. Anaplastic lymphoma kinase (ALK) fusion transcripts were detected in 15 of 16 ALK-positive anaplastic large-cell lymphomas. The latter case was further characterized by a novel SATB1&#95;ALK fusion transcript. Among 239 other PTCLs, representative of nine entities, non-ALK fusion transcripts were detected in 24 samples, mostly of follicular helper T-cell (TFH) derivation. The most frequent non-ALK fusion transcript was ICOS&#95;CD28 in nine TFH-PTCLs, one PTCL not otherwise specified, and one adult T-cell leukemia/lymphoma, followed by VAV1 rearrangements with multiple partners (STAP2, THAP4, MYO1F, and CD28) in five samples (three PTCL not otherwise specified and two TFH-PTCLs). The other rearrangements were CTLA4&#95;CD28 (one TFH-PTCL), ITK&#95;SYK (two TFH-PTCLs), ITK&#95;FER (one TFH-PTCL), IKZF2&#95;ERBB4 (one TFH-PTCL and one adult T-cell leukemia/lymphoma), and TP63&#95;TBL1XR1 (one ALK-negative anaplastic large-cell lymphoma). All fusions detected by our assay were validated by conventional RT-PCR and Sanger sequencing in 30 samples with adequate material. The simplicity and robustness of this targeted multiplex assay make it an attractive tool for the characterization of these heterogeneous diseases., (Copyright © 2021 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Activating mutations of MYD88 ( MYD88 L265P being the far most frequent) are found in most cases of Waldenström macroglobulinemia (WM) as well as in various aggressive B-cell lymphoma entities with features of plasma cell (PC) differentiation, such as activated B-cell type diffuse large B-cell lymphoma (DLBCL). To understand how MYD88 activation exerts its transformation potential, we developed a new mouse model in which the MYD88 L252P protein, the murine ortholog of human MYD88 L265P , is continuously expressed in CD19 positive B-cells together with the Yellow Fluorescent Protein (Myd88 L252P mice). In bone marrow, IgM B and plasma cells were expanded with a CD138 expression continuum from IgM high CD138 low to IgM low CD138 high cells and the progressive loss of the B220 marker. Serum protein electrophoresis (SPE) longitudinal analysis of 40 Myd88 L252P mice (16 to 56 weeks old) demonstrated that ageing was first associated with serum polyclonal hyper gammaglobulinemia (hyper Ig) and followed by a monoclonal immunoglobulin (Ig) peak related to a progressive increase in IgM serum levels. All Myd88 L252P mice exhibited spleen enlargement which was directly correlated with the SPE profile and was maximal for monoclonal Ig peaks. Myd88 L252P mice exhibited very early increased IgM PC differentiation. Most likely due to an early increase in the Ki67 proliferation index, IgM lymphoplasmacytic (LP) and plasma cells continuously expanded with age being first associated with hyper Ig and then with monoclonal Ig peak. This peak was consistently associated with a spleen LP-like B-cell lymphoma. Clonal expression of both membrane and secreted µ chain isoforms was demonstrated at the mRNA level by high throughput sequencing. The Myd88 L252P tumor transcriptomic signature identified both proliferation and canonical NF-κB p65/RelA activation. Comparison with MYD88 L265P WM showed that Myd88 L252P tumors also shared the typical lymphoplasmacytic transcriptomic signature of WM bone marrow purified tumor B-cells. Altogether these results demonstrate for the first time that continuous MYD88 activation is specifically associated with clonal transformation of differentiating IgM B-cells. Since MYD88 L252P targets the IgM PC differentiation continuum, it provides an interesting preclinical model for development of new therapeutic approaches to both WM and aggressive MYD88 associated DLBCLs., 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 Ouk, Roland, Gachard, Poulain, Oblet, Rizzo, Saintamand, Lemasson, Carrion, Thomas, Balabanian, Espéli, Parrens, Soubeyran, Boulin, Faumont, Feuillard and Vincent-Fabert.)

The RYTHMIC network, supported by the French National Cancer Institute is dedicated to the management of patients with thymic epithelial tumors through regional and national multidisciplinary tumor boards. Tumor board decisions are based on the initial pathology diagnoses. However, following clinical inclusion in the network, a central pathology review is organized, implicating a panel of pathologists, for histotype and stage classification, which is different from a classical second opinion from pathologist to pathologist for a difficult case. Thanks to the participation of all French pathologists, more than 1000 cases have been reviewed by the panel. The aim of this review is to share with the French pathology community, the experience of the group. It underlines the importance of macroscopy and surgeon-pathologist involvement to allow a good central review, the main histopathological and immunophenotypical patterns of the most frequent thymomas and thymic carcinoma types, the differential diagnoses, as well as the difficulties for the panel to reproducibly assess on slides, stage, for some cases., (Copyright © 2020 Elsevier Masson SAS. All rights reserved.)

Angioimmunoblastic T-cell lymphoma (AITL) is a frequent T-cell lymphoma in the elderly population that has a poor prognosis when treated with cyclophosphamide, doxorubicin, vincristine, and prednisone  (CHOP) therapy. Lenalidomide, which has been safely combined with CHOP to treat B-cell lymphoma, has shown efficacy as a single agent in AITL treatment. We performed a multicentric phase 2 trial combining 25 mg lenalidomide daily for 14 days per cycle with 8 cycles of CHOP21 in previously untreated AITL patients aged 60 to 80 years. The primary objective was the complete metabolic response (CMR) rate at the end of treatment. Seventy-eight of the 80 patients enrolled were included in the efficacy and safety analysis. CMR was achieved in 32 (41%; 95% confidence interval [CI], 30%-52.7%) patients, which was below the prespecified CMR rate of 55% defined as success in the study. The 2-year progression-free survival (PFS) was 42.1% (95% CI, 30.9%-52.8%), and the 2-year overall survival was 59.2% (95% CI, 47.3%-69.3%). The most common toxicities were hematologic and led to treatment discontinuation in 15% of patients. This large prospective and uniform series of AITL treatment data was used to perform an integrative analysis of clinical, pathologic, biologic, and molecular data. TET2, RHOA, DNMT3A, and IDH2 mutations were present in 78%, 54%, 32%, and 22% of patients, respectively. IDH2 mutations were associated with distinct pathologic and clinical features and DNMT3A was associated with shorter PFS. In conclusion, the combination of lenalidomide and CHOP did not improve the CMR in AITL patients. This trial clarified the clinical impact of recurrent mutations in AITL. This trial was registered at www.clincialtrials.gov as #NCT01553786., (© 2021 by The American Society of Hematology.)

Background: Classification of thymic epithelial tumours (TETs) is known to be challenging; however, the level of discordances at a nationwide level between initial and expert diagnosis and their clinical consequences are currently unknown. RYTHMIC is a national network dedicated to the management of TET based on initial histological diagnosis, followed by an additional expert review of all cases. Our aim was to evaluate the discordances between initial and expert diagnoses and whether they would have led to different clinical management., Patients and Methods: We conducted a retrospective analysis of the cohort of patients discussed at RYTHMIC tumour board from January 2012 to December 2016. Assessment of disagreement was made for histological typing and for staging. The discordances were classified as major or minor based on whether they would have changed or not the proposed therapeutic strategy, respectively. Follow-up of the patients with major discordances was conducted until December 2018., Results: Four hundred sixty-seven patients were reviewed, and 183 (39%) discordances were identified either related to histological subtype (132) and/or stage (72). Major discordances were identified in 27 patients (6%). They included 16 patients with TET for whom treatment recommendation based on the central review would have been post-operative radiotherapy, whereas it had not been the case. However, follow-up did not show any progression among the 15 patients with high-grade histology and/or stage resected thymomas. On the other hand, among the remaining 11 patients including 7 with a diagnosis other than TET, the overall management or follow-up would have been completely different with the expert diagnosis., Conclusion: Our real-life cohort reveals a high level of discordances considering TET diagnosis and supports expert review for optimal clinical management., Competing Interests: Conflict of interest statement The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this article., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Peripheral T cell lymphomas are rare malignancies with aggressive course, with several different subtype described in the 2016 WHO classification. Their distribution across the world is heterogenous, with marked difference between Western and Asian country. Their clinical presentation often comprise extra-nodal involvement, B symptoms and immune system disorder which can lead to wrong diagnosis orientation. Make a right diagnosis need a experienced pathologist in close collaboration with clinical datas. Peripheral T cell lymphomas are in general associated with poor prognosis when treated with anthracyclines-based regimen, and several studies and trials focused on the use of intensified regimen or novel targeted agents, whose proper indication still remain to be clarified., (Copyright © 2020. Published by Elsevier Masson SAS.)

In humans, histiocytic sarcoma (HS) is an aggressive cancer involving histiocytes. Its rarity and heterogeneity explain that treatment remains a challenge. Sharing high clinical and histopathological similarities with human HS, the canine HS is conversely frequent in specific breeds and thus constitutes a unique spontaneous model for human HS to decipher the genetic bases and to explore therapeutic options. We identified sequence alterations in the MAPK pathway in at least 63.9% (71/111) of HS cases with mutually exclusive BRAF (0.9%; 1/111), KRAS (7.2%; 8/111) and PTPN11 (56.75%; 63/111) mutations concentrated at hotspots common to human cancers. Recurrent PTPN11 mutations are associated to visceral disseminated HS subtype in dogs, the most aggressive clinical presentation. We then identified PTPN11 mutations in 3/19 (15.7%) human HS patients. Thus, we propose PTPN11 mutations as key events for a specific subset of human and canine HS: the visceral disseminated form. Finally, by testing drugs targeting the MAPK pathway in eight canine HS cell lines, we identified a better anti-proliferation activity of MEK inhibitors than PTPN11 inhibitors in canine HS neoplastic cells. In combination, these results illustrate the relevance of naturally affected dogs in deciphering genetic mechanisms and selecting efficient targeted therapies for such rare and aggressive cancers in humans., (© 2020 UICC.)

Peripheral T-cell lymphoma comprises a heterogeneous group of mature non-Hodgkin lymphomas. Their diagnosis is challenging, with up to 30% of cases remaining unclassifiable and referred to as "not otherwise specified". We developed a reverse transcriptase-multiplex ligation-dependent probe amplification gene expression profiling assay to differentiate the main T-cell lymphoma entities and to study the heterogeneity of the "not specified" category. The test evaluates the expression of 20 genes, including 17 markers relevant to T-cell immunology and lymphoma biopathology, one Epstein-Barr virus-related transcript, and variants of RHOA (G17V) and IDH2 (R172K/T). By unsupervised hierarchical clustering, our assay accurately identified 21 of 21 ALK-positive anaplastic large cell lymphomas, 16 of 16 extranodal natural killer (NK)/T-cell lymphomas, 6 of 6 hepatosplenic T-cell lymphomas, and 13 of 13 adult T-cell leukemia/lymphomas. ALK-negative anaplastic lymphomas (n=34) segregated into one cytotoxic cluster (n=10) and one non-cytotoxic cluster expressing Th2 markers (n=24) and enriched in DUSP22 -rearranged cases. The 63 T FH -derived lymphomas divided into two subgroups according to a predominant T FH (n=50) or an enrichment in Th2 (n=13) signatures. We next developed a support vector machine predictor which attributed a molecular class to 27 of 77 not specified T-cell lymphomas: 17 T FH , five cytotoxic ALK-negative anaplastic and five NK/T-cell lymphomas. Among the remaining cases, we identified two cell-of-origin subgroups corresponding to cytotoxic/Th1 (n=19) and Th2 (n=24) signatures. A reproducibility test on 40 cases yielded a 90% concordance between three independent laboratories. This study demonstrates the applicability of a simple gene expression assay for the classification of peripheral T-cell lymphomas. Its applicability to routinely-fixed samples makes it an attractive adjunct in diagnostic practice., (Copyright© 2020 Ferrata Storti Foundation.)

The prognostic value of cell of origin (COO) classification and BCL2 expression is not well established in diffuse large B-cell lymphoma (DLBCL) patients with human immunodeficiency virus (HIV) infection in the recent era. Phenotypic patterns were determined by immunohistochemistry (IHC) of pathological samples from patients with HIV-associated DLBCL prospectively enrolled in the French AIDS and Viral Hepatitis CO16 Lymphovir cohort between 2008 and 2015. Molecular subgroup classification into germinal centre B-cell (GCB) and non-GCB subtypes was determined using the Hans algorithm. Among 52 samples of systemic DLBCL subjected to centralized pathological analysis, 25 of the 42 tested for BCL2 expression were positive. Samples were further classified into GCB (n = 19) and non-GCB (n = 16) subtypes and 17 remained unclassified. In multivariable analysis, BCL2 expression was an independent pejorative prognostic biomarker [4-year progression-free survival (PFS): 52% for BCL2 + vs. 88% for BCL2 - , P = 0·02] and tended to reduce 4-year overall survival (OS) (63% for BCL2 + vs. 88% for BCL2 - , P = 0·06). The difference between CGB and non-GCB subtypes on PFS and OS did not reach significance (4-year PFS: 79% for GCB vs. 53% for non-GCB, P = 0·24 and 4-year OS: 78% for GCB vs. 69% for non-GCB, P = 0·34). BCL2 expression determined by IHC is an independent pejorative prognostic biomarker in HIV-associated DLBCL in the recent era. This supports the investigation of new therapeutic strategies in patients with BCL2 expression., (© 2019 British Society for Haematology and John Wiley & Sons Ltd.)

The major weakness of most knock-in JAK2V617F mouse models is the presence of the JAK2 mutation in all rather than in a few hematopoietic stem cells (HSC), such as in human "early-stage" myeloproliferative neoplasms (MPN). Understanding the mechanisms of disease initiation is critical as underscored by the incidence of clonal hematopoiesis of indeterminate potential associated with JAK2V617F. Currently, such studies require competitive transplantation. Here, we report a mouse model obtained by crossing JAK2V617F/WT knock-in mice with PF4iCre transgenic mice. As expected, PF4iCre;JAK2V617F/WT mice developed an early thrombocytosis resulting from the expression of JAK2V617F in the megakaryocytes. However, these mice then developed a polycythemia vera-like phenotype at 10 weeks of age. Using mT/mG reporter mice, we demonstrated that Cre recombination was present in all hematopoietic compartments, including in a low number of HSC. The frequency of mutated cells increased along hematopoietic differentiation mimicking the clonal expansion observed in essential thrombocythemia and polycythemia vera patients. This model thus mimics the HSC compartment observed in early-stage MPN, with a small number of JAK2V617F HSC competing with a majority of JAK2WT HSC. PF4iCre;JAK2V617F/WT mice are a promising tool to investigate the mechanisms that regulate clonal dominance and progression to myelofibrosis., (© 2019 by The American Society of Hematology.)

Background: Immunotherapy is currently under investigation in B3 Thymoma (TB3) and Thymic Carcinoma (TC). PD-L1 expression has been evaluated on a limited number of patients with selected antibodies. We aimed to analyze cohort of TB3 and TC with a panel of antibodies to assess the prevalence of PD-L1 expression, its prognostic value and to set up a reproducible test., Methods: We retrospectively studied 103 patients samples of FFPE histologically confirmed TB3 ( n  = 53) and TC ( n  = 50) by expert pathologists within the RYTHMIC national network. We compared PD-L1, PD1, CD8 and PD-L2 expression and performed correlation with tumor types and patients outcomes. Four PD-L1 antibodies were tested, three of them validated as companion tests in lung cancer, one tested on two automates on whole section of tumors. We evaluated the percentage and intensity of both epithelial and immune stained cells., Results: TB3 epithelial cells had a higher and more diffuse expression of PD-L1 than TC regardless the antibodies tested ( p  < 0.0001). Three out of four antibodies targeting PD-L1 tested on the DAKO autostainer gave similar staining. Concordance between antibodies was lower for PD-L1 staining on immune cells with no significant difference between TB3 and TC except on E1L3N antibody. PD-L2 antibody stained no tumor epithelial cells. High PD-L1 expression was correlated with a better overall survival for TB3 and was not correlated with tumor staging., Conclusion: Frequent PD-L1 expression, particularly in TB3, paves the way for immunotherapy in TET (Thymic Epithelial Tumor). Otherwise, we have set up three reproducible LDT (laboratory-developed test) for four PD-L1 antibodies., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s). 2019.)

Objective: Angioimmunoblastic T-cell lymphoma (AITL) is frequently associated with autoimmune cytopenia (AIC). Whether such patients have a particular phenotype and require particular management is unclear., Method: Angioimmunoblastic T-cell lymphoma patients from the multicentric database of the Lymphoma Study Association presenting with AIC during disease course were included and matched to AITL patients without AIC (1/5 ratio)., Results: At diagnosis, AIC patients (n = 28) had more spleen and bone marrow involvement (54% vs 19% and 71% vs 34%, P < 0.001), Epstein-Barr virus replication (89% vs 39%, P < 0.001), gamma globulin titers (median 23 vs 15 g/L, P = 0.002), and proliferating B cells and plasmablasts in biopsies, as compared to control patients (n = 136). The 28 AIC patients had 41 episodes of AIC, diagnosed concomitantly with AITL in 23 (82%) cases. After a median follow-up of 24 months (range 3-155), 10 patients relapsed, all associated with AITL relapse., Conclusion: Our results provide new insight into AIC associated with AITL by highlighting the significant interplay between AITL and B-cell activation leading to subsequent autoimmunity., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Aims: We applied the 2017 World Health Organization (WHO) classification criteria to categorise a series of 64 primary cutaneous large B-cell lymphomas (PCLBCLs), containing a majority (≥80%) of large cells and a proliferative rate of ≥40%, raising the problem of the differential diagnosis between PCLBCL, leg type (PCLBCL-LT) and primary cutaneous follicle centre lymphoma, large cell (PCFCL-LC). The aims were to determine the reproducibility and prognostic relevance of the 2017 WHO criteria., Methods and Results: Morphology and phenotype identified 32 PCLBCLs-LT and 25 PCFCLs-LC; seven cases (11%) remained unclassified. Morphology was less reproducible than immunophenotype. Pertinent markers for the differential diagnosis were MUM1, FOXP1, CD10, and IgM. bcl-2 and bcl-6 were expressed by both PCFCLs-LC and PCLBCLs-LT at substantial levels. Neither Ki67 expression nor p63 expression was of diagnostic value. MYD88 was found to be mutated only in PCLBCLs-LT (n = 22, 69%). According to Hans/Hans modified algorithms, 23 of 25 PCFCLs-LC had germinal centre (GC) status, and the 32 PCLBCLs-LT had non-GC status. Overall survival was poorer for PCLBCLs-LT than PCFCLs-LC (P = 0.0002). Non-GC cases had poorer overall survival than GC cases (P = 0.0007). In PCLBCLs-LT, MYC expression was associated with cutaneous relapses (P = 0.014). When GC/non-GC status was applied to unclassified cases, only a single case remained discordant., Conclusions: Our results support the 2017 WHO classification criteria for PCLBCL diagnosis. The Hans modified algorithm using CD10 and MUM1 distinguished PCFCLs-LC from PCLBCLs-LT with optimal diagnostic value without requiring bcl-6 immunolabelling (poorly reproducible). Rare unclassified cases may constitute a provisionally heterogeneous subgroup for which GC/non-GC status (relevant for prognosis) may guide therapeutic decisions., (© 2019 John Wiley & Sons Ltd.)

Essentials To reliably study the respective roles of blood and endothelial cells in hemostasis, mouse models with a strong and specific endothelial expression of the Cre recombinase are needed. Using mT/mG reporter mice and conditional JAK2 V617F/WT mice, we compared Pdgfb-iCreERT2 and Cdh5(PAC)-CreERT2 with well-characterized Tie2-Cre mice. Comparison of recombination efficiency and specificity towards blood lineage reveals major differences between endothelial transgenic mice. Cre-mediated recombination occurs in a small number of adult hematopoietic stem cells in Pdgfb-iCreERT2;JAK2 V617F/WT transgenic mice. SUMMARY: Background The vessel wall, and particularly blood endothelial cells (BECs), are intensively studied to better understand hemostasis and target thrombosis. To understand the specific role of BECs, it is important to have mouse models that allow specific and homogeneous expression of genes of interest in all BEC beds without concomitant expression in blood cells. Inducible Pdgfb-iCreERT2 and Cdh5(PAC)-CreERT2 transgenic mice are widely used for BEC targeting. However, issues remain in terms of recombination efficiency and specificity regarding hematopoietic cells. Objectives To determine which mouse model to choose when strong expression of a transgene is required in adult BECs from various organs, without concomitant expression in hematopoietic cells. Methods Using mT/mG reporter mice to measure recombination efficiency and conditional JAK2 V617F/ WT mice to assess specificity regarding hematopoietic cells, we compared Pdgfb-iCreERT2 and Cdh5(PAC)-CreERT2 with well-characterized Tie2-Cre mice. Results Adult Cdh5(PAC)-CreERT2 mice are endothelial specific but require a dose of 10 mg of tamoxifen to allow constant Cre expression. Pdgfb-iCreERT2 mice injected with 5 mg of tamoxifen are appropriate for most endothelial research fields except liver studies, as hepatic sinusoid ECs are not recombined. Surprisingly, 2 months after induction of Cre-mediated recombination, all Pdgfb-iCreERT2;JAK2 V617F/ WT mice developed a myeloproliferative neoplasm that is related to the presence of JAK2V617F in hematopoietic cells, showing for the first time that Cre-mediated recombination occurs in a small number of adult hematopoietic stem cells in Pdgfb-iCreERT2 transgenic mice. Conclusion This study provides useful guidelines for choosing the best mouse line to study the role of BECs in hemostasis and thrombosis., (© 2019 International Society on Thrombosis and Haemostasis.)

Background: Although a prognosis of recurrent or refractory childhood Hodgkin lymphoma (HL) is associated with poor outcomes despite intensive therapy, the immune checkpoint inhibitors PD-1/PD-L1 appear to be therapeutic alternatives for advanced adult cases. However, these pharmacotherapies are yet to be studied in a pediatric population., Procedure: The present study measured the expression of PD-1/PD-L1 in diagnostic samples of children with classical HL, according to the disease course. This study included two groups of patients treated at the Department of Pediatric Oncology, Bordeaux University Hospital-a group of cured or in-remission cases and a group of relapsed or refractory cases. Immunohistochemical analyses of anti-PD-1 and anti-PD-L1 (clone 28-8, companion test for nivolumab) were performed on baseline and follow-up biopsies., Results: Of the 42 included patients, 31 were cured or in remission and 11 were categorized as relapsed or refractory. At the time of diagnosis, PD-1 expression was low (1-3% of intratumoral lymphocytes labeled) in <20% of cases, whereas PD-L1 was expressed by tumor cells in all cases, and strongly (≥50%) in most cases. There were no significant differences in the expression levels of the two checkpoint molecules between the groups. Initial biopsies showed strong expression of PD-L1, whereas expression of PD-1 was rare., Conclusions: The identical labeling profiles of the cured and relapsed/refractory patients suggest that comparable responses to inhibitors of the PD1/PDL1 immunological checkpoints could be expected in patients undergoing first-, second-, or third-line therapy., (© 2019 Wiley Periodicals, Inc.)

Gray-zone lymphoma (GZL) with features intermediate between classic Hodgkin lymphoma (cHL) and large B-cell lymphoma (LBCL) was introduced as a provisional entity into the World Health Organization classification in 2008. However, as diagnostic criteria are imprecise, reliable identification of GZL cases remains challenging. Here, we describe the histopathologic features of 139 GZL cases from a retrospective Lymphoma Study Association (LYSA) study with the goal to improve classification accuracy. Inclusion criteria were based on literature review and an expert consensus opinion of the LYSA hematopathologist panel. We observed 86 cases with a morphology more closely related to cHL, but with an LBCL immunophenotype based on strong and homogenous B-cell marker expression (CD20 and/or CD79a, OCT2, BOB1, PAX5) on all tumor cells (cHL-like GZL). Fifty-three cases were morphologically more closely related to LBCL but harbored a cHL immunophenotype (LBCL-like GZL). Importantly, we observed a continuous morphologic and immunophenotypic spectrum within these 2 GZL categories. The majority of cases presented genetic immune escape features with CD274/PDCD1LG2 and/or CIITA structural variants by fluorescence in situ hybridization. Patients without mediastinal involvement at diagnosis (17%) were older than those with mediastinal tumors (median: 56 vs. 39 y). Cases associated with Epstein-Barr virus (24%) presented with similar patient characteristics and outcome as Epstein-Barr virus negative cases. In summary, we provide refined diagnostic criteria that contribute to a more precise pathologic and clinical characterization of GZL within a broad spectrum from cHL-like to LBCL-like disease.

Herein we report the case of a 41-year-old woman who presented with pelvic pain. Magnetic Resonance Imaging exhibited a single pelvic mass, measuring 50mm long axis, alongside the right iliac vessels. Histological examination of the excision specimen showed a lymphoid tumor with features of localized Castleman disease, hyaline vascular type. Moreover we identified multiple interfollicular dark clusters, composed of cells morphologically resembling cortical thymocytes. Their immunophenotype was consistent with an intermediate stage of T-cell differentiation, with the expression of CD3, CD4, CD8, TdT, CD1a, CD99, CD2, CD5, CD7 and CD10, with 40% Ki67. After integration of clinical and molecular data, the retained diagnosis was an indolent T-cell lymphoblastic proliferation associated with hyaline vascular localized Castleman disease. The clinical course confirmed the indolent nature of the proliferation, despite a late local recurrence at 7 years of the initial diagnosis, without histological modification, due to an incomplete initial resection surgery., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Common variable immunodeficiency (CVID) is characterized by low levels of circulating immunoglobulins and defects in B cell maturation leading to an increased susceptibility to infections. Some patients develop complications such as autoimmune diseases, enteropathy, and lymphoproliferation, resulting in higher morbidity and mortality. Follicular helper T (Tfh) cells are specialized in helping B cell differentiation into Ig-producing cells. Three subsets have been described, namely non B-cell helper Tfh1 and the two B-helper cell subsets Tfh2 and Tfh17. We determined that circulating Tfh cells were elevated in CVID patients and skewed toward a Tfh1 profile. Interestingly, elevated levels of Tfh1 cells were significant only in patients harboring non-infectious complications regardless of the type of complication and inversely correlated with switched memory B cells. Moreover, CXCR3 + cells are increased in splenic CVID germinal centers. Our observations suggest that the altered balance in Tfh subsets in CVID is linked to a more severe disease., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Immune thrombocytopenia (ITP) is an acquired antibody-mediated disease, for which splenectomy remains a curative treatment. We analyzed histology and phenotypes of ITP-splenectomy specimens from 41 adult patients, with different previous ITP-specific treatments, including B-cell-depleting rituximab (RTX) or not, in an attempt to predict splenectomy success or failure on the basis of day 56 postoperative platelet counts. RTX-naive ITP-spleen samples, compared with those from a 20-patient control trauma cohort, contained the following nonspecific, ITP-evocative, white-pulp lesions: follicular helper T-cell (programmed death-1 and inducible T-cell COStimulator) expansion in reactive follicles (P=0.01 and 0.03, respectively) and regulatory T-cell (FOXP3) expansion in the T-cell zone (P=0.049). On comparing ITP-splenectomy samples that would be successful with those that would be failures, only marginal zone hyperplasia differed (P=0.017). Indeed, 13/21 (61.9%) successful splenectomy samples exhibited marginal zone hyperplasia, as opposed to 1/9 (11.1%) failed splenectomy specimens. RTX impact on ITP-splenectomy samples was characterized by white-pulp (P=0.03) and marginal zone atrophies (P=0.01), and periarteriolar T-cell-zone hyperplasia (P<0.0001). The results of this novel comparative study of the histologic patterns of 41 ITP patients' evocative splenic lesions enabled clear description of different ITP morphologies and phenotypes, as a function of prior treatment and splenectomy success or failure.

Skin biopsies of 41 angioimmunoblastic T-cell lymphoma patients were retrospectively analyzed for the expression of follicular helper T-cell (TFH) markers, Epstein-Barr virus (EBV), and the presence of RHOA (p.G17V) and IDH2 (p.R172K/S) mutations using allele-specific polymerase chain reaction. We categorized cases into 4 distinctive patterns: (1) low-density lymphocytic perivascular infiltrates (n=11), (2) dense perivascular infiltrates with atypical cells and occasional inflammatory cells (n=13), (3) diffuse infiltrates reminiscent of angioimmunoblastic T-cell lymphoma (n=4), or (4) other aspects (n=13). Two EBV and 2 plasmacytoid lymphoproliferative disorders were seen. We observed variable expression of TFH markers (CD10 [50%], BCLB6 [84%], PD1 [94%], CXCL13 [84%], and ICOS [97.5%]), and EBV B-blasts (26%). A TFH phenotype was identified in 82% and 73%, respectively, of cases with the most challenging patterns 1 and 2. TFH markers and EBV can thus help for diagnosis and are detected in samples with low-density infiltrates. We found RHOA G17V and IDH2 R172K/S mutations in the skin in 14/18 (78%) and 3/16 (19%) cases, respectively. The RHOA G17V mutation was identified in a proportion of biopsies with patterns 1 and 2, which represent a diagnostic challenge. The RHOA G17V mutation was detected both in the skin and lymph node (LN) biopsies in 7/9 (64%) cases, and in only the skin or the LN of 1 sample each. The frequency of RHOA G17V mutation was similar to that reported in LNs. It may represent a sensitive diagnostic marker in the skin, helpful in cases with low-density infiltrates.

Background: Immunophenotype of primary cutaneous diffuse large B-cell lymphoma, leg-type (PCLBCL-LT) suggests a germinal center-experienced B lymphocyte (BCL2+ MUM1+ BCL6+/-)., Objectives: As maturation history of B-cell is "imprinted" during B-cell development on the immunoglobulin gene sequence, we studied the structure and sequence of the variable part of the genes (IGHV, IGLV, IGKV), immunoglobulin surface expression and features of class switching in order to determine the PCLBCL-LT cell of origin., Methods: Clonality analysis with BIOMED2 protocol and VH leader primers was done on DNA extracted from frozen skin biopsies on retrospective samples from 14 patients. The clonal DNA IGHV sequence of the tumor was aligned and compared with the closest germline sequence and homology percentage was calculated. Superantigen binding sites were studied. Features of selection pressure were evaluated with the multinomial Lossos model., Results: A functional monoclonal sequence was observed in 14 cases as determined for IGHV (10), IGLV (2) or IGKV (3). IGV mutation rates were high (>5%) in all cases but one (median:15.5%), with superantigen binding sites conservation. Features of selection pressure were identified in 11/12 interpretable cases, more frequently negative (75%) than positive (25%). Intraclonal variation was detected in 3 of 8 tumor specimens with a low rate of mutations. Surface immunoglobulin was an IgM in 12/12 cases. FISH analysis of IGHM locus, deleted during class switching, showed heterozygous IGHM gene deletion in half of cases. The genomic PCR analysis confirmed the deletions within the switch μ region. IGV sequences were highly mutated but functional, with negative features of selection pressure suggesting one or more germinal center passage(s) with somatic hypermutation, but superantigen (SpA) binding sites conservation. Genetic features of class switch were observed, but on the non functional allele and co-existing with primary isotype IgM expression., Conclusion: These data suggest that cell-of origin is germinal center experienced and superantigen driven selected B-cell, in a stage between germinal center B-cell and plasma cell., (Copyright © 2017 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)