Primary effusion lymphoma (PEL) is a rare subtype of non-Hodgkin's lymphoma, which is associated with infection by Kaposi's sarcoma herpesvirus (KSHV)/human herpesvirus-8. The c-Myc transcription ...factor plays an important role in cellular proliferation, differentiation and apoptosis. Lymphomas frequently have deregulated c-Myc expression owing to chromosomal translocations, amplifications or abnormal stabilization. However, no structural abnormalities were found in the c-myc oncogene in PEL. Given that c-Myc is often involved in lymphomagenesis, we hypothesized that it is deregulated in PEL. We report that PEL cells have abnormally stable c-Myc protein. The turnover of c-Myc protein is stringently regulated by post-transcriptional modifications, including phosphorylation of c-Myc threonine 58 (T58) by glycogen synthase kinase-3beta (GSK-3beta). Our data show that the impaired c-Myc degradation in PEL cells is associated with a significant underphosphorylation of c-Myc T58. The KSHV latency-associated nuclear antigen (LANA) is responsible for this deregulation. Overexpression of LANA in human embryonic kidney 293 or peripheral blood B cells leads to post-transcriptional deregulation of c-Myc protein. Conversely, when LANA is eliminated from PEL cells using RNA interference, GSK-3beta-mediated c-Myc T58 phosphorylation is restored. The presence of c-Myc and LANA in GSK-3beta-containing complexes in PEL cells further confirms the significance of these interactions in naturally KSHV-infected cells.
Aims: Kaposi sarcoma herpesvirus (KSHV) is aetiologically related to Kaposi sarcoma, classical and extracavitary primary effusion lymphoma (PEL; EC‐PEL) and multicentric Castleman disease (MCD), ...entities preferentially occurring in HIV‐infected individuals. Characterization of HIV‐associated PELs/EC‐PELs suggests that the KSHV‐infected malignant cells originate from a pre‐terminal stage of B‐cell differentiation. However, only limited phenotypic studies have been performed on HIV+ MCD, including for PR domain containing 1 with zinc finger domain/B lymphocyte‐induced maturation protein 1 (PRDM1/BLIMP1), a key regulator of terminal B‐cell differentiation. The aim was to characterize KSHV‐infected cells in 17 cases of HIV+ MCD.
Methods and results: Double immunohistochemistry and immunohistochemistry–in situ hybridization were used to characterize the KSHV‐infected cells in MCD; the results were compared with the phenotypic profiles of 39 PELs/EC‐PELs and seven PEL cell lines. Whereas the immunophenotype of KSHV‐infected cells in MCD and malignant KSHV+ PEL cells was similar (PAX5, Bcl‐6−; PRDM1/BLIMP1, IRF4/MUM1+; Ki67+), the MCD KSHV‐infected cells differed, as they expressed OCT2, cytoplasmic λ immunoglobulin; variably expressed CD27; lacked CD138; and were Epstein–Barr virus negative.
Conclusions: Although both PEL and MCD originate from KSHV‐infected pre‐terminally differentiated B cells, these findings, with previously reported genetic studies, indicate HIV+ MCD may arise from extrafollicular B cells, whereas PELs may originate from cells that have traversed the germinal centre.
Kaposi sarcoma–associated herpesvirus (KSHV), or human herpervirus 8 (HHV-8), is a γ-herpesvirus that infects human lymphocytes and is associated with primary effusion lymphoma (PEL). Currently, the ...role of viral infection in the transformation of PEL cells is unknown. One possibility is that KSHV, like the lymphotropic viruses Epstein-Barr virus (EBV) and human T-cell leukemia virus I (HTLV-I), activates the transcription factor NF-κB to promote survival and proliferation of infected lymphocytes. To examine this possibility, we assessed NF-κB activity in KSHV-infected PEL cell lines and primary tumor specimens by electrophoretic mobility shift assay (EMSA). We observed that NF-κB is constitutively activated in all KSHV-infected lymphomas, and consists of 2 predominant complexes, p65/p50 heterodimers and p50/p50 homodimers. Inhibition experiments demonstrated that Bay 11-7082, an irreversible inhibitor of IκBα phosphorylation, completely and specifically abrogated the NF-κB/DNA binding in PEL cells. PEL cells treated with Bay 11 demonstrated down-regulation of the NF-κB inducible cytokine interleukin 6 (IL-6), and apoptosis. These results suggest that NF-κB activity is necessary for survival of KSHV-infected lymphoma cells, and that pharmacologic inhibition of NF-κB may be an effective treatment for PEL.
Kaposi sarcoma-associated herpesvirus (KSHV), also called human herpesvirus 8 (HHV-8), is associated with a specific subset of lymphoproliferative disorders. These include two main categories. The ...first is primary effusion lymphomas and related solid variants. The second is multicentric Castleman disease, from which KSHV-positive plasmablastic lymphomas can arise. KSHV contributes to lymphomagenesis by subverting the host cell molecular signaling machinery to deregulate cell growth and survival. KSHV expresses a selected set of genes in the lymphoma cells, encoding viral proteins that play important roles in KSHV lymphomagenesis. Deregulation of the NF-kappaB pathway is an important strategy used by KSHV to promote lymphoma cell survival, and the viral protein vFLIP is essential for this process. Two other viruses that are well documented to be causally associated with lymphoid neoplasia in humans are Epstein-Barr virus (EBV/HHV-4) and human T-cell lymphotropic virus (HTLV-1). Both of these are similar to KSHV in their use of viral proteins to promote cell survival by deregulating the NF-kappaB pathway. Here we review the basic information and recent developments that have contributed to our knowledge of lymphomas caused by KSHV and other viruses. The understanding of the mechanisms of viral lymphomagenesis should lead to the identification of novel therapeutic targets and to the development of rationally designed therapies.
The recently identified Kaposi's sarcoma-associated herpesvirus (KSHV), also known as human herpesvirus 8 (HHV-8), has been found to be consistently associated with an unusual subset of acquired ...immunodeficiency syndrome-related lymphomas, the so-called body cavity-based lymphomas (BCBL) or primary effusion lymphomas (PEL). These lymphomas are characterized by a unique spectrum of morphologic and molecular characteristics, and grow as lymphomatous effusions without an identifiable contiguous tumor mass. Until now, efforts to delineate the role of KSHV in the pathogenesis of PELs have been hampered by the lack of appropriate model systems and the concomitant presence of Epstein-Barr virus (EBV) in nearly all cases examined, and in all previously established cell lines. We now report the establishment and characterization of a novel PEL cell line, BC-3, which is KSHV+ by polymerase chain reaction (PCR) but EBV- as assessed by a variety of methods including PCR for EBER, EBNA-2, and EBNA-3C. This cell line was established from a lymphomatous effusion obtained from an HIV- patient, and has immunophenotypic and molecular features consistent with the diagnosis of PEL, including an indeterminate immunophenotype with a B-cell immunogenotype and lack of c-myc proto-oncogene rearrangements. Pulsed-field gel electrophoresis shows an intact KSHV genome of about 170 kb both in the cell line and in the viral isolate, whereas herpesvirus-like capsids are visible by electron microscopy. Consequently, the BC-3 cell line represents an invaluable tool as a source of KSHV, for both the evaluation of the pathogenic potential of this virus and the mechanistic characterization of its role in the development of Kaposi's sarcoma and malignant lymphoma.
Two unique DNA fragments were recently identified in over 90% of acquired immunodeficiency syndrome (AIDS)-related Kaposi's sarcoma tissues. Sequence analysis suggests that these fragments belong to ...a previously unidentified human herpesvirus, Kaposi's sarcoma-associated herpesvirus (KSHV). These fragments have also been identified in a subset of non-Hodgkin's lymphomas in human immunodeficiency virus-positive patients; specifically, in body cavity-based lymphomas (AIDS-BCBLs). We have established two cell lines derived from AIDS-BCBLs, BC-1 and BC-2, which retain the KSHV sequences, and have used them to further characterize this putative viral genome. In this report, we demonstrate that the KSHV sequences represent a portion of a much larger DNA molecule that is located predominantly in the nucleus of the infected cells. In situ hybridization of metaphase spreads made from both of these cell lines show these sequences in episomal structures. Their presence in the cells as large nuclear episomes supports previous sequence homology data suggesting that KSHV belongs to the herpesvirus family. These cell lines provide a continuous source of KSHV sequences. Thus, they represent an important tool for future studies of this recently described human herpesvirus that may be involved in the pathogenesis of Kaposi's sarcoma and some AIDS-related non-Hodgkin's lymphomas.
Primary effusion lymphomas (PELs) associated with infection by the Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) have constitutive nuclear factor (NF)-kappaB activity that is essential for ...their survival, but the source of this activity is unknown. We report that viral FADD-like interleukin-1-beta-converting enzyme FLICE/caspase 8-inhibitory protein (FLIP) activates NF-kappaB more potently than cellular FLIP in B cells and that it is largely responsible for NF-kappaB activation in latently infected PEL cells. Elimination of vFLIP production in PEL cells by RNA interference results in significantly decreased NF-kappaB activity, down-regulation of essential NF-kappaB-regulated cellular prosurvival factors, induction of apoptosis, and enhanced sensitivity to external apoptotic stimuli. vFLIP is the first virally encoded gene shown to be essential for the survival of naturally infected tumor cells.
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