EBV-positive HIV-associated diffuse large B cell lymphomas are characterized by JAK/STAT (STAT3) pathway mutations and unique clinicopathologic features.

Even in the era of highly active combination antiretroviral therapy (cART), patients with HIV have a disproportionate risk of developing aggressive lymphomas that are frequently Epstein-Barr virus (EBV)-related. Here, we investigate HIV-associated diffuse large B-cell lymphoma (HIV-DLBCL) and compare EBV-positive and EBV-negative cases. HIV-DLBCL were identified from two academic medical centres and characterised by immunohistochemistry, EBV status, fluorescence in situ hybridisation, cell of origin determination by gene expression profiling, and targeted deep sequencing using a custom mutation panel of 334 genes. We also applied the Lymphgen tool to determine the genetic subtype of each case. Thirty HIV-DLBCL were identified, with a median patient age of 46 years and male predominance (5:1). Thirteen cases (48%) were EBV-positive and 14 (52%) EBV-negative. Nine of the 16 tested cases (56%) had MYC rearrangement, three (19%) had BCL6 (two of which were double hit MYC/BCL6) and none had BCL2 rearrangements. Using the Lymphgen tool, half of the cases (15) were classified as other. All HIV-DLBCL showed mutational abnormalities, the most frequent being TP53 (37%), MYC (30%), STAT3 (27%), HIST1H1E (23%), EP300 (20%), TET2 (20%), SOCS1 (17%) and SGK1 (17%). EBV-negative cases were mostly of germinal centre B-cell (GCB) origin (62%), showed more frequent mutations per case (a median of 13·5/case) and significant enrichment of TP53 (57% vs. 15%; P = 0·046), SGK1 (36% vs. 0%; P = 0·04), EP300 (43% vs. 0%; P = 0·02) and histone-modifying gene (e.g. HIST1H1E, HIST1H1D, 79% vs. 31%; P = 0·02) mutations. EBV-positive cases were mostly of non-GCB origin (70%), with fewer mutations per case (median 8/case; P = 0·007), and these tumours were enriched for STAT3 mutations (P = 0·10). EBV-positive cases had a higher frequency of MYC mutations but the difference was not significant (36% vs. 15%; P = 0·38). EBV-association was more frequent in HIV-DLBCLs, arising in patients with lower CD4 counts at diagnosis (median 46·5 vs. 101, P = 0·018). In the era of cART, approximately half of HIV-DLBCL are EBV-related. HIV-DLBCL are enriched for MYC rearrangements, MYC mutations and generally lack BCL2 rearrangements, regardless of EBV status. Among HIV-DLBCL, tumours that are EBV-negative and EBV-positive appear to have important differences, the latter arising in context of lower CD4 count, showing frequent non-GCB origin, lower mutation burden and recurrent STAT3 mutations.

Chapman JR ，Bouska AC ，Zhang W ，Alderuccio JP ，Lossos IS ，Rimsza LM ，Maguire A ，Yi S ，Chan WC ，Vega F ，Song JY ... -  《-》

Gene expression profiling of Epstein-Barr virus-positive diffuse large B-cell lymphoma of the elderly reveals alterations of characteristic oncogenetic pathways.

Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma (DLBCL) of the elderly (EBV[+]DLBCL-E) is classified as a subtype of DLBCL. Until now, its molecular pathogenesis has remained unknown. To identify pathways characteristic of EBV(+)DLBCL-E, gene expression profiling of five EBV(+)DLBCL-E and seven EBV-negative DLBCL (EBV[-]DLBCL) cases was undertaken using human oligonucleotide microarray analysis. Gene set enrichment analysis and gene ontology analysis showed that gene sets of the Janus kinase-signal transducer and activator of transcription (JAK-STAT) and nuclear factor kappa B (NF-κB) pathways were enriched in EBV(+)DLBCL-E cases. To confirm the results of the expression profiles, in vitro analysis was performed. Expression profiling analysis showed that high activation of the JAK-STAT and NF-κB pathways was induced by EBV infection into DLBCL cell lines. Activation of the NF-κB pathway was confirmed in EBV-infected cell lines using an electrophoretic mobility shift assay. Western blot analysis revealed an increased protein expression level of phosphorylated signal transducer and activator of transcription 3 (STAT3) in an EBV-infected cell line. Protein expression of phosphorylated STAT3 was frequently observed in lymphoma cells of EBV(+)DLBCL-E clinical samples using immunohistochemistry (EBV[+]DLBCL-E: 80.0% [n = 20/25] versus EBV[-]DLBCL: 38.9% [n = 14/36]; P = 0.001). The results of the present study suggest that activation of the JAK-STAT and NF-κB pathways was characteristic of EBV(+)DLBCL-E, which may reflect the nature of EBV-positive tumor cells. Targeting these pathways as therapies might improve clinical outcomes of EBV(+)DLBCL-E.

Kato H ，Karube K ，Yamamoto K ，Takizawa J ，Tsuzuki S ，Yatabe Y ，Kanda T ，Katayama M ，Ozawa Y ，Ishitsuka K ，Okamoto M ，Kinoshita T ，Ohshima K ，Nakamura S ，Morishima Y ，Seto M ... -  《-》

Clinical Significance of BCL2, C-MYC, and BCL6 Genetic Abnormalities, Epstein-Barr Virus Infection, CD5 Protein Expression, Germinal Center B Cell/Non-Germinal Center B-Cell Subtypes, Co-expression of MYC/BCL2 Proteins and Co-expression of MYC/BCL2/BCL6 P

Ting CY ，Chang KM ，Kuan JW ，Sathar J ，Chew LP ，Wong OJ ，Yusuf Y ，Wong L ，Samsudin AT ，Pana MNBM ，Lee SK ，Gopal NSR ，Puri R ，Ong TC ，Bahari SK ，Goh AS ，Teoh CS ... -  《International Journal of Medical Sciences》

pSTAT3 and MYC in Epstein-Barr virus-positive diffuse large B-cell lymphoma.

Teoh SH ，Khoo JJ ，Abdul Salam DSD ，Peh SC ，Cheah SC ... -  《Malaysian Journal of Pathology》

Epstein-Barr Virus Infection of Cell Lines Derived from Diffuse Large B-Cell Lymphomas Alters MicroRNA Loading of the Ago2 Complex.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive lymphoid tumor which is occasionally Epstein-Barr virus (EBV) positive and is further subtyped as activated B-cell DLBCL (ABC-DLBCL) and germinal center B-cell DLBCL (GCB-DLBCL), which has implications for prognosis and treatment. We performed Ago2 RNA immunoprecipitation followed by high-throughput RNA sequencing (Ago2-RIP-seq) to capture functionally active microRNAs (miRNAs) in EBV-negative ABC-DLBCL and GCB-DLBCL cell lines and their EBV-infected counterparts. In parallel, total miRNA profiles of these cells were determined to capture the cellular miRNA profile for comparison with the functionally active profile. Selected miRNAs with differential abundances were validated using real-time quantitative PCR (RT-qPCR) and Northern blotting. We found 6 miRNAs with differential abundances (2 upregulated and 4 downregulated miRNAs) between EBV-negative and -positive ABC-DLBCL cells and 12 miRNAs with differential abundances (3 upregulated and 9 downregulated miRNAs) between EBV-negative and -positive GCB-DLBCL cells. Eight and twelve miRNAs were confirmed using RT-qPCR in ABC-DLBCL and GCB-DLBCL cells, respectively. Selected miRNAs were analyzed in additional type I/II versus type III EBV latency DLBCL cell lines. Furthermore, upregulation of miR-221-3p and downregulation of let7c-5p in ABC-DLBCL cells and upregulation of miR-363-3p and downregulation of miR-423-5p in GCB-DLBCL cells were verified using RIP-Northern blotting. Our comprehensive sequence analysis of the DLBCL miRNA profiles identified sets of deregulated miRNAs by Ago2-RIP-seq. Our Ago2-IP-seq miRNA profile could be considered an important data set for the detection of deregulated functionally active miRNAs in DLBCLs and could possibly lead to the identification of miRNAs as biomarkers for the classification of DLBCLs or even as targets for personalized targeted treatment.IMPORTANCE Diffuse large B-cell lymphoma (DLBCL) is a highly aggressive tumor of lymphoid origin which is occasionally Epstein-Barr virus (EBV) positive. MicroRNAs are found in most multicellular organisms and even in viruses such as EBV. They regulate the synthesis of proteins by binding to their cognate mRNA. MicroRNAs are tethered to their target mRNAs by "Argonaute" proteins. Here we compared the overall miRNA content of the Ago2 complex by differential loading to the overall content of miRNAs in two DLBCL cell lines and their EBV-converted counterparts. In all cell lines, the Ago2 load was different from the overall expression of miRNAs. In addition, the loading of the Ago2 complex was changed upon infection with EBV. This indicates that the virus not only changes the overall content of miRNAs but also influences the expression of proteins by affecting the Ago complexes.

Ayoubian H ，Ludwig N ，Fehlmann T ，Menegatti J ，Gröger L ，Anastasiadou E ，Trivedi P ，Keller A ，Meese E ，Grässer FA ... -  《-》