Pathogenic Variants Associated with Epigenetic Control and the NOTCH Pathway Are Frequent in Classic Hodgkin Lymphoma.

Classic Hodgkin lymphoma (cHL) constitutes a B-cell neoplasm derived from germinal center lymphocytes. Despite high cure rates (80-90%) obtained with the current multiagent protocols, a significant proportion of cHL patients experience recurrences, characterized by a lower sensitivity to second-line treatments. The genomic background of chemorefractory cHL is still poorly understood, limiting personalized treatment strategies based on molecular features. In this study, using a targeted next-generation sequencing (NGS) panel specifically designed for cHL research, we compared chemosensitive and chemorefractory diagnostic tissue samples of cHL patients. Furthermore, we longitudinally examined paired diagnosis-relapsesamples of chemorefractory cHL in order to define patterns of dynamic evolution and clonal selection. Pathogenic variants in NOTCH1 and NOTCH2 genes frequently arise in cHL. Mutations in genes associated with epigenetic regulation (CREBBP and EP300) are particularly frequent in relapsed/refractory cHL. The appearance of novel clones characterized by mutations previously not identified at diagnosis is a common feature in cHL cases showing chemoresistance to frontline treatments. Our results expand current molecular and pathogenic knowledge of cHL and support the performance of molecular studies in cHL prior to the initiation of first-line therapies.

Whole slide imaging of tumour microenvironment in classical Hodgkin's lymphoma: development of a clinical prediction model based on programmed death-ligand 1 and tumorous Reed-Sternberg cells.

AIMS: The prognostic impact of programmed death-ligand 1 (PD-L1) cells in classic Hodgkin lymphoma (cHL) tumour microenvironment remains undefined. METHODS: Model development via Transparent Reporting of a multivariable prediction model for Individual Prognosis Or Diagnosis guidelines were followed. PD-L1+ and CD30+ tumoral Reed-Sternberg cells were quantified through whole slide imaging and digital image analysis in 155 digital histopathological slides of cHL. Univariate and multivariate survival analyses were performed. The analyses were reproduced for patients with advanced stages (IIB, III and IV) using the Advanced-stage cHL International Prognostic Index. RESULTS: The PD-L1/CD30 ratio was statistically significantly associated with survival outcomes. Patients with a PD-L1/CD30 ratio above 47.1 presented a shorter overall survival (mean OS: 53.7 months; 95% CI: 28.7 to 78.7) in comparison with patients below this threshold (mean OS: 105.4 months; 95% CI: 89.6 to 121.3) (p=0.04). When adjusted for covariates, the PD-L1/CD30 ratio retained prognostic impact, both for the OS (HR: 1.005; 95% CI: 1.002 to 1.008; p=0.000) and the progression-free survival (HR: 3.442; 95% CI: 1.045 to 11.340; p=0.04) in a clinical and histopathological multivariate model including the male sex (HR: 3.551; 95% CI: 0.986 to 12.786; p=0.05), a percentage of tumoral cells ≥10.1% (HR: 1.044; 95% CI: 1.003 to 1.087; p=0.03) and high risk International Prognostic Score (≥3 points) (HR: 6.453; 95% CI: 1.970 to 21.134; p=0.002). CONCLUSIONS: The PD-L1/CD30 ratio identifies a group of cHL patients with an increased risk of treatment failure. Its clinical application can be performed as it constitutes an easy to implement pathological information in the diagnostic work-up of patients with cHL.

Identification of prognostic factors in classic Hodgkin lymphoma by integrating whole slide imaging and next generation sequencing.

Digital pathology and genomics are increasingly used to improve our understanding of lymphoid neoplasms. Algorithms for quantifying cell populations in the lymph node and genetics can be integrated to identify new biomarkers with prognostic impact in classic Hodgkin lymphoma (cHL). In 16 cHL patients, we have performed whole slide imaging (WSI) analysis and quantification of CD30+, CD20+, CD3+ and MUM1+ cells in whole tissue slides, and Next Generation Sequencing (NGS) in formalin fixed paraffin-embedded (FFPE) tissue, using a widely used NSG panel (Oncomine® Focus Assay) to define genetic variants underlying tumor development. The different cell populations could be successfully identified in scanned slides of cHL, supporting the inclusion of WSI in the histopathological evaluation of cHL as an adequate method for the quantification of different cell populations. We also performed genetic profiling in FFPE samples of cHL leading to the identification of copy number variations in the Neurofibromin 1 gene (17q11.2) and the Androgen Receptor gene (Xq12) accompanied by chromosomal gains and losses in CDK4, KRAS and FGFR2 genes. Progression-free survival (PFS) was statistically significantly higher in cHL patients with amplification in the NF1 gene combined with CD3+ cells above 28.6% (p = 0.006) and MUM1+ cells above 21.8% (p < 0.001). Moreover, patients with MUM1+ cells above 21.8% showed a statistically significantly higher PFS when combined with amplification of the AR gene (p < 0.001) and wild-type KRAS (p < 0.001). The integration of WSI analysis and DNA sequencing could be useful to improve our understanding of the biology of cHL and define risk subgroups.

Whole-slide image analysis identifies a high content of Hodgkin Reed-Sternberg cells and a low content of T lymphocytes in tumor microenvironment as predictors of adverse outcome in patients with classic Hodgkin lymphoma treated with ABVD.

Classic Hodgkin lymphoma (cHL) constitutes the most frequent lymphoma in young adults. Its histopathology is unique as a scattered tumor population, termed Hodgkin Reed-Sternberg (HRS) cells is diluted in a prominent tumor microenvironment (TME) composed of T lymphocytes, B lymphocytes, macrophages, neutrophils, eosinophils and histiocytes. Traditionally, the identification of prognostic biomarkers in the cHL TME has required visual inspection and manual counting by pathologists. The advent of whole-slide imaging (WSI) and digital image analysis methods could significantly contribute to improve this essential objective in cHL research, as a 10-20% of patients are still refractory or relapsed after conventional chemotherapy. In this work, we have digitized a total of 255 diagnostic cHL slides and quantified the proportion of HRS cells (CD30), B cells (CD20) and T cells (CD3) by digital image analysis. Data obtained where then correlated with the overall survival (OS) and progression free survival (PFS) of cHL patients. Quantification of HRS cells, B cells and T cells reflects the biological heterogeneity of the different cHL histological subtypes analyzed. A percentage of 2.00% of HRS cells statistically significantly discriminated between patients achieving a complete metabolic response (CMR) and refractory or relapsed (R/R) patients both for the OS (P=0.001) and PFS (P=0.005). Furthermore, patients with a percentage of T cells below the 26.70% in the TME showed a statistically significantly shorter OS (P=0.019) and PFS (P=0.041) in comparison with patients above this threshold. A subgroup of patients with a low content of T cells and high content of HRS cells exhibited a special aggressive clinical course. Currently, there is the need to implement quantitative and easy scalable methods to enhance clinical translation, as the cHL TME plays a central role in the clinical course of the disease. The results of this study could contribute to the identification of prognostic biomarkers specifically looking at the cHL TME and their inclusion in future clinical trials.

The Need for Standardization in Next-Generation Sequencing Studies for Classic Hodgkin Lymphoma: A Systematic Review.

Classic Hodgkin lymphoma (cHL) constitutes a B cell-derived neoplasm defined by a scarce tumoral population, termed Hodgkin and Reed-Sternberg (HRS) cells, submerged into a histologically heterogeneous microenvironment. The paucity of HRS cells has historically hampered genetic studies, rendering the identification of the recurrent genetic lesions and molecular pathways deregulated in this lymphoma difficult. The advent of high-throughput sequencing methods such as next-generation sequencing (NGS) could sensibly optimize the identification of the mutational landscape of cHL. However, there is no current consensus either in the design of panels for targeted NGS or in its most relevant clinical applications. In this work, we systematically review the current state of NGS studies of cHL, stressing the need for standardization both in the candidate genes to be analyzed and the bioinformatic pipelines. As different institutions have developed and implemented their own customized NGS-based protocols, to compare and systematically review the major findings of this ongoing research area could be of added value for centers that routinely perform diagnostic, monitoring and genotyping strategies in cHL samples. The results of this systematic review should contribute to the interdepartmental harmonization and achievement of a consensus in the current clinical applications of NGS studies of cHL.

Prognostic Role of the Expression of Latent-Membrane Protein 1 of Epstein-Barr Virus in Classical Hodgkin Lymphoma.

The prognostic impact of the presence of Epstein-Barr virus (EBV) in classical Hodgkin lymphoma (cHL) is controversial. Previous studies reported heterogeneous results, rendering difficult the clinical validation of EBV as a prognostic biomarker in this lymphoma. The objective of this study was to evaluate the survival impact of the expression of EBV Latent-Membrane Protein 1 (EBV-LMP1) in tumoral Hodgkin-Reed-Sternberg (HRS) cells of primary diagnostic samples of cHL. Formalin-Fixed Paraffin-Embedded (FFPE) lymph node samples from 88 patients with cHL were analyzed. Patients were treated with the standard first-line chemotherapy (CT) with Adriamycin, Bleomycin, Vinblastine and Dacarbazine (ABVD) followed by radiotherapy. The Kaplan-Meier method and the Cox proportional hazards model were used for carrying out the survival analysis. In order to investigate whether the influence of EBV was age-dependent, analyses were performed both for patients of all ages and for age-stratified subgroups. In bivariate analysis, the expression of EBV was associated with older age (p = 0.011), mixed cellularity subtype cHL (p < 0.001) and high risk International Prognostic Score (IPS) (p = 0.023). Overall survival (OS) and progression-free survival (PFS) were associated with the presence of bulky disease (p = 0.009) and advanced disease at diagnosis (p = 0.016). EBV-positive cases did not present a significantly lower OS and PFS in comparison with EBV-negative cases, for all ages and when stratifying for age. When adjusted for covariates, absence of bulky disease at diagnosis (HR: 0.102, 95% CI: 0.02-0.48, p = 0.004) and limited disease stages (I-II) (HR: 0.074, 95% CI: 0.01-0.47, p = 0.006) were associated with a significant better OS. For PFS, limited-disease stages also retained prognostic impact in the multivariate Cox regression (HR: 0.145, 95% CI: 0.04-0.57, p = 0.006). These results are of importance as the early identification of prognostic biomarkers in cHL is critical for guiding and personalizing therapeutic decisions. The prognostic role of EBV in cHL could be modulated by the type of CT protocol employed and interact with the rest of presenting features.

Translational Applications of Artificial Intelligence and Machine Learning for Diagnostic Pathology in Lymphoid Neoplasms: A Comprehensive and Evolutive Analysis.

Genomic analysis and digitalization of medical records have led to a big data scenario within hematopathology. Artificial intelligence and machine learning tools are increasingly used to integrate clinical, histopathological, and genomic data in lymphoid neoplasms. In this study, we identified global trends, cognitive, and social framework of this field from 1990 to 2020. Metadata were obtained from the Clarivate Analytics Web of Science database in January 2021. A total of 525 documents were assessed by document type, research areas, source titles, organizations, and countries. SciMAT and VOSviewer package were used to perform scientific mapping analysis. Geographical distribution showed the USA and People's Republic of China as the most productive countries, reporting up to 190 (36.19%) of all documents. A third-degree polynomic equation predicts that future global production in this area will be three-fold the current number, near 2031. Thematically, current research is focused on the integration of digital image analysis and genomic sequencing in Non-Hodgkin lymphomas, prediction of chemotherapy response and validation of new prognostic models. These findings can serve pathology departments to depict future clinical and research avenues, but also, public institutions and administrations to promote synergies and optimize funding allocation.

Chemokine receptor expression on infiltrating lymphocytes from abdominal aortic aneurysms: role of CXCR4-CXCL12 in lymphoid recruitment.

This is a study to define the profile of chemokine receptors expressed on isolated infiltrating lymphocytes in human abdominal aortic aneurysms (AAAs), and to examine their role in lymphoid recruitment. AAA T-lymphocytes were CXCR4-positive, CCR7-negative and partially CXCR3 and CCR5-positive. Functionally, AAA T-cells were proinflammatory effector cells, as they produced IFN-gamma and granzyme A. AAA B-lymphocytes were CXCR4-positive and exhibited low CXCR5 expression. A relevant feature of infiltrating T- and B-lymphocytes was the high intensity of CXCR4 expression and the capability to migrate to CXCL12. CXCL12-producing cells were found in the adventitia of AAA. These cells were CD45-negative and partially VCAM-1 and DR-positive. In summary, the present results suggest that CXCR4, expressed on infiltrating lymphocytes, and CXCL12, expressed on stromal cells, is involved in the recruitment of lymphocytes within the arterial wall in AAA.

[Cronkhite-Canada syndrome: an acquired, potentially reversible polyposis]. / Síndrome de Cronkhite-Canada: una poliposis adquirida y potencialmente reversible.

Cronkhite-Canada syndrome is an infrequent, nonadenomatous, acquired polyposis that is associated with ectodermic alterations. The etiopathogenesis and optimal therapeutic management are unknown. We present a case of Cronkhite-Canada syndrome in a 58-year-old man who, after combined treatment with corticosteroids, disodium cromoglycate, loratadine, ciprofloxacin, and zinc, showed complete clinical and partial endoscopic remission. The polypoid lesions of Cronkhite-Canada syndrome can be reversed with medical treatment. Consequently, the natural history of the disease can be modified and its prognosis improved.

Characterisation of T and B lymphocytes infiltrating abdominal aortic aneurysms.

Cellular infiltrates consisting mainly of lymphocytes are commonly present in the arterial wall in abdominal aortic aneurysm (AAA). However, despite this, the precise nature of these populations has to date not been described in detail. The aim of this study was to perform an exhaustive phenotypic characterisation of the infiltrating lymphocytes in order to define the inflammatory process that develops in AAA. For this purpose, AAA-infiltrating lymphocytes from 25 fresh aneurysm wall samples and, as a control, peripheral blood (PB) lymphocytes from the same patients were purified. The expression of lineage specific markers, maturation molecules, activation antigens and adhesion molecules on T and B lymphocytes was examined by triple-colour immunofluorescence and flow cytometry analysis. The phenotype of AAA-infiltrating lymphocytes was compared with that of PB of the patients with AAA. The results reveal that AAA-infiltrating B and T lymphocytes consist of activated memory cells, with specific homing properties. In addition, they express molecules of B-T co-stimulation and, occasionally, exhibit phenotypical features indicative of the ectopic formation of lymphoid structures. These findings are discussed in the light of the similarities shared with the lymphoid infiltration occurring in other chronic autoimmune/inflammatory disorders.