Publications by authors named "Lorenzo Leoncini"

94 Publications

MiR-200c-3p Contrasts PD-L1 Induction by Combinatorial Therapies and Slows Proliferation of Epithelial Ovarian Cancer through Downregulation of β-Catenin and c-Myc.

Cells 2021 Mar 1;10(3). Epub 2021 Mar 1.

Department of Experimental Medicine, Sapienza University of Rome, 00161 Rome, Italy.

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.
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http://dx.doi.org/10.3390/cells10030519DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7998372PMC
March 2021

Follicular lymphoma t(14;18)-negative is genetically a heterogeneous disease.

Blood Adv 2020 11;4(22):5652-5665

Institute of Pathology and Neuropathology, Eberhard Karls University of Tübingen-Comprehensive Cancer Center, University Hospital Tübingen, Tübingen, Germany.

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.
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http://dx.doi.org/10.1182/bloodadvances.2020002944DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7686888PMC
November 2020

Prognostic impact of tumor-associated macrophages, lymphocyte-to-monocyte and neutrophil-to-lymphocyte ratio in diffuse large B-cell lymphoma.

Am J Blood Res 2020 25;10(4):97-108. Epub 2020 Aug 25.

Unit of Pathology, Department of Medical Biotechnologies, University of Siena Siena, Italy.

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.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7486489PMC
August 2020

The Binding of CD93 to Multimerin-2 Promotes Choroidal Neovascularization.

Invest Ophthalmol Vis Sci 2020 07;61(8):30

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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.
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http://dx.doi.org/10.1167/iovs.61.8.30DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425738PMC
July 2020

Frequent traces of EBV infection in Hodgkin and non-Hodgkin lymphomas classified as EBV-negative by routine methods: expanding the landscape of EBV-related lymphomas.

Mod Pathol 2020 12 1;33(12):2407-2421. Epub 2020 Jun 1.

Section of Pathology, Department of Medical Biotechnology, University of Siena, Siena, Italy.

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.
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http://dx.doi.org/10.1038/s41379-020-0575-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7685982PMC
December 2020

Immune landscape in Burkitt lymphoma reveals M2-macrophage polarization and correlation between PD-L1 expression and non-canonical EBV latency program.

Infect Agent Cancer 2020 6;15:28. Epub 2020 May 6.

11Department of Cellular Pathology, University College Hospital, London, London UK.

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.
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http://dx.doi.org/10.1186/s13027-020-00292-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7201729PMC
May 2020

Correction to: IGHV mutational status of nodal marginal zone lymphoma by NGS reveals distinct pathogenic pathways with different prognostic implications.

Virchows Arch 2020 07;477(1):169

Department of Medical Biotechnologies, Anatomic Pathology Division, University of Siena, Via delle Scotte, 6, 53100, Siena, Italy.

This error was caused due to the author's oversight and this does not change the views or the results presented in the manuscript.
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http://dx.doi.org/10.1007/s00428-020-02794-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7645535PMC
July 2020

IGHV mutational status of nodal marginal zone lymphoma by NGS reveals distinct pathogenic pathways with different prognostic implications.

Virchows Arch 2020 Jul 4;477(1):143-150. Epub 2019 Dec 4.

Department of Medical Biotechnologies, Anatomic Pathology Division, University of Siena, Via delle Scotte, 6, 53100, Siena, Italy.

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.
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http://dx.doi.org/10.1007/s00428-019-02712-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7320062PMC
July 2020

Molecular switch from MYC to MYCN expression in MYC protein negative Burkitt lymphoma cases.

Blood Cancer J 2019 11 20;9(12):91. Epub 2019 Nov 20.

Department of Medical Biotechnology, University of Siena, Siena, Italy.

MYC is the most altered oncogene in human cancer, and belongs to a large family of genes, including MYCN and MYCL. Recently, while assessing the degree of correlation between MYC gene rearrangement and MYC protein expression in aggressive B-cell lymphomas, we observed few Burkitt lymphoma (BL) cases lacking MYC protein expression despite the translocation involving the MYC gene. Therefore, in the present study we aimed to better characterize such cases. Our results identified two sub-groups of MYC protein negative BL: one lacking detectable MYC protein expression but presenting MYCN mRNA and protein expression; the second characterized by the lack of both MYC and MYCN proteins but showing MYC mRNA. Interestingly, the two sub-groups presented a different pattern of SNVs affecting MYC gene family members that may induce the switch from MYC to MYCN. Particulary, MYCN-expressing cases show MYCN SNVs at interaction interface that stabilize the protein associated with loss-of-function of MYC. This finding highlights MYCN as a reliable diagnostic marker in such cases. Nevertheless, due to the overlapping clinic, morphology and immunohistochemistry (apart for MYC versus MYCN protein expression) of both sub-groups, the described cases represent bona fide BL according to the current criteria of the World Health Organization.
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http://dx.doi.org/10.1038/s41408-019-0252-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6868231PMC
November 2019

status in chronic lymphocytic leukemia identify ethnic groups with an aggressive clinical course .

Haematologica 2019 10;104(10):e493

Section of Pathology, Department of Medical Biotechnology, University of Siena, Siena, Italy

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http://dx.doi.org/10.3324/haematol.2019.219618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6886440PMC
October 2019

Interplay between the Epigenetic Enzyme Lysine (K)-Specific Demethylase 2B and Epstein-Barr Virus Infection.

J Virol 2019 07 14;93(13). Epub 2019 Jun 14.

International Agency for Research on Cancer, World Health Organization, Lyon, France

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 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 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. 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 gene. 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.
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http://dx.doi.org/10.1128/JVI.00273-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6580945PMC
July 2019

p66Shc deficiency in the Eμ-TCL1 mouse model of chronic lymphocytic leukemia enhances leukemogenesis by altering the chemokine receptor landscape.

Haematologica 2019 10 28;104(10):2040-2052. Epub 2019 Feb 28.

Department of Life Sciences, University of Siena, Siena

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.
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http://dx.doi.org/10.3324/haematol.2018.209981DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6886430PMC
October 2019

The mutational landscape of Burkitt-like lymphoma with 11q aberration is distinct from that of Burkitt lymphoma.

Blood 2019 02 19;133(9):962-966. Epub 2018 Dec 19.

Institute of Human Genetics, Ulm University and Ulm University Medical Center, Ulm, Germany.

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- 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 -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 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 or An exception is , 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.
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http://dx.doi.org/10.1182/blood-2018-07-864025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6396176PMC
February 2019

Granulysin, a novel marker for extranodal NK/T cell lymphoma, nasal type.

Virchows Arch 2018 Dec 27;473(6):749-757. Epub 2018 Aug 27.

Department of Histopathology, University College London, University College Hospital, Rockefeller Building, UCL Site, 21 University Street, London, WC1E 6JJ, UK.

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.
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http://dx.doi.org/10.1007/s00428-018-2434-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6267266PMC
December 2018

Epstein-Barr Virus-Induced Metabolic Rearrangements in Human B-Cell Lymphomas.

Front Microbiol 2018 8;9:1233. Epub 2018 Jun 8.

Section of Pathology, Department of Medical Biotechnology, University of Siena, Siena, Italy.

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.
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http://dx.doi.org/10.3389/fmicb.2018.01233DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6002739PMC
June 2018

Pathobiologic Roles of Epstein-Barr Virus-Encoded MicroRNAs in Human Lymphomas.

Int J Mol Sci 2018 Apr 12;19(4). Epub 2018 Apr 12.

Department of Experimental, Diagnostic, and Experimental Medicine, Bologna University School of Medicine, 40126 Bologna, Italy.

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 () and Bam HI fragment H rightward open reading frame 1 (), 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.
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http://dx.doi.org/10.3390/ijms19041168DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5979337PMC
April 2018

MicroRNAs sequencing unveils distinct molecular subgroups of plasmablastic lymphoma.

Oncotarget 2017 Dec 31;8(64):107356-107373. Epub 2017 Oct 31.

Department of Medical Biotechnology, Section of Pathology, University of Siena, Siena, Italy.

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 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, 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.
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http://dx.doi.org/10.18632/oncotarget.22219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5746073PMC
December 2017

Preferential Usage of Specific Immunoglobulin Heavy Chain Variable Region Genes With Unmutated Profile and Advanced Stage at Presentation Are Common Features in Patients With Chronic Lymphocytic Leukemia From Senegal.

Am J Clin Pathol 2017 Nov;148(6):545-554

Department of Experimental, Diagnostic, and Experimental Medicine, Bologna University School of Medicine, Bologna, Italy.

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.
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http://dx.doi.org/10.1093/ajcp/aqx105DOI Listing
November 2017

Unveiling Another Missing Piece in EBV-Driven Lymphomagenesis: EBV-Encoded MicroRNAs Expression in EBER-Negative Burkitt Lymphoma Cases.

Front Microbiol 2017 1;8:229. Epub 2017 Mar 1.

Section of Pathology, Department of Medical Biotechnology, University of Siena Siena, Italy.

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 . 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 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) 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 ( = 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.
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http://dx.doi.org/10.3389/fmicb.2017.00229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5331039PMC
March 2017

Optimal Minimal Panels of Immunohistochemistry for Diagnosis of B-Cell Lymphoma for Application in Countries With Limited Resources and for Triaging Cases Before Referral to Specialist Centers.

Am J Clin Pathol 2016 May;145(5):687-95

From the Department of Histopathology, Imperial College Healthcare NHS Trust & Imperial College, London, United Kingdom

Objectives: Establish and validate optimal minimal immunohistochemistry panels for usage in a staged algorithmic manner for precise diagnosis of B-cell lymphomas in countries with limited resources. Suggest short panels of immunostains to be used in referring units that refer suspected lymphomas to specialist diagnostic centers in resourceful countries.

Methods: Significant proportion of six B-cell lymphomas has characteristic morphology requiring a short panel of confirmatory immunostains. The rest would go through five different algorithms.

Results: 812 cases in which a B-cell lymphoma or an HIV-associated lymphoma was suspected on morphological grounds were evaluated. This led to arriving at a specific diagnosis of 799 B-cell lymphomas. A correct diagnosis was achievable in 69% cases with the application of three to five antibodies; others required additional work-up.

Conclusions: The panels/algorithms assist pathologists in practicing lymphoma diagnostics in countries with limited resources and in making lymphoma referrals to specialist centers.
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http://dx.doi.org/10.1093/ajcp/aqw060DOI Listing
May 2016

The cell of origin of Burkitt lymphoma: germinal centre or not germinal centre?

Histopathology 2016 11 15;69(5):885-886. Epub 2016 Jul 15.

Department of Medical Biotechnology, Section of Pathology, University of Siena, Siena, Italy.

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http://dx.doi.org/10.1111/his.12997DOI Listing
November 2016

Clonality Analysis of Immunoglobulin Gene Rearrangement by Next-Generation Sequencing in Endemic Burkitt Lymphoma Suggests Antigen Drive Activation of BCR as Opposed to Sporadic Burkitt Lymphoma.

Am J Clin Pathol 2016 Jan;145(1):116-27

From the Department of Medical Biotechnologies, University of Siena, Siena, Italy.

Objectives: Recent studies using next-generation sequencing (NGS) analysis disclosed the importance of the intrinsic activation of the B-cell receptor (BCR) pathway in the pathogenesis of sporadic Burkitt lymphoma (sBL) due to mutations of TCF3/ID3 genes. Since no definitive data are available on the genetic landscape of endemic Burkitt (eBL), we first assessed the mutation frequency of TCF3/ID3 in eBL compared with sBL and subsequently the somatic hypermutation status of the BCR to answer whether an extrinsic activation of BCR signaling could also be demonstrated in Burkitt lymphoma.

Methods: We assessed the mutations of TCF3/ID3 by RNAseq and the BCR status by NGS analysis of the immunoglobulin genes (IGs).

Results: We detected mutations of TCF3/ID3 in about 30% of the eBL cases. This rate is significantly lower than that detected in sBL (64%). The NGS analysis of IGs revealed intraclonal diversity, suggesting an active targeted somatic hypermutation process in eBL compared with sBL.

Conclusions: These findings support the view that the antigenic pressure plays a key role in the pathogenetic pathways of eBL, which may be partially distinct from those driving sBL development.
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http://dx.doi.org/10.1093/ajcp/aqv011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4778259PMC
January 2016

A look into the evolution of Epstein-Barr virus-induced lymphoproliferative disorders: a case study.

Am J Clin Pathol 2015 Nov;144(5):817-22

From the Department of Medical Biotechnologies, Section of Pathology, University of Siena, Siena, Italy;

Objectives: Epstein-Barr virus (EBV)-induced lymphoproliferative disorders (LPDs) are lymphoid proliferations arising as a result of the loss of an effective EBV-specific cytotoxic T-cell response. LPDs may occur for primary or acquired impairment of the immune system, as well as in some persons without documented immunodeficiency.

Methods: In this article, we describe the case of a human immunodeficiency virus-positive patient affected by an EBV-LPD of the stomach who developed a nodal diffuse large B-cell lymphoma with complex morphologic and molecular features.

Results: GeneScan analysis of the gastric specimen identified two different heavy-chain immunoglobulin gene (IGH) rearrangements characterized by a dominant peak of 285 base pairs (bp) in length and a smaller peak of 266 bp in length. In the lymph node sample, IGH evaluation also demonstrated two different peaks; however, the main peak corresponded to the minor peak detected in the EBV-LPD specimen at the diagnosis. In addition, a monoclonal immunoglobulin light chain gene (IGL) rearrangement was also found. We also demonstrated that the major peak in the stomach corresponded to the EBV-positive population observed in the histologic sections.

Conclusions: This case may provide additional insights to better understanding the "hit-and-run" role for EBV in lymphomagenesis. However, we could not exclude that our findings represent the co-occurrence of two unrelated B-cell neoplasms rather than a progression from an EBV-positive neoplasm to an EBV-negative one.
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http://dx.doi.org/10.1309/AJCP2G0VKTKPNPRRDOI Listing
November 2015

Distinct Viral and Mutational Spectrum of Endemic Burkitt Lymphoma.

PLoS Pathog 2015 Oct 15;11(10):e1005158. Epub 2015 Oct 15.

Department of Systems Biology, Columbia University College of Physicians and Surgeons, New York, New York, United States of America; Department of Biomedical Informatics, Columbia University College of Physicians and Surgeons, New York, New York, United States of America.

Endemic Burkitt lymphoma (eBL) is primarily found in children in equatorial regions and represents the first historical example of a virus-associated human malignancy. Although Epstein-Barr virus (EBV) infection and MYC translocations are hallmarks of the disease, it is unclear whether other factors may contribute to its development. We performed RNA-Seq on 20 eBL cases from Uganda and showed that the mutational and viral landscape of eBL is more complex than previously reported. First, we found the presence of other herpesviridae family members in 8 cases (40%), in particular human herpesvirus 5 and human herpesvirus 8 and confirmed their presence by immunohistochemistry in the adjacent non-neoplastic tissue. Second, we identified a distinct latency program in EBV involving lytic genes in association with TCF3 activity. Third, by comparing the eBL mutational landscape with published data on sporadic Burkitt lymphoma (sBL), we detected lower frequencies of mutations in MYC, ID3, TCF3 and TP53, and a higher frequency of mutation in ARID1A in eBL samples. Recurrent mutations in two genes not previously associated with eBL were identified in 20% of tumors: RHOA and cyclin F (CCNF). We also observed that polyviral samples showed lower numbers of somatic mutations in common altered genes in comparison to sBL specimens, suggesting dual mechanisms of transformation, mutation versus virus driven in sBL and eBL respectively.
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http://dx.doi.org/10.1371/journal.ppat.1005158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4607508PMC
October 2015