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Primary effusion lymphoma (PEL) is a malignant B-cell lymphoma attributable to Kaposi sarcoma-associated herpesvirus (KSHV) infection. PEL is characterized by invasive behavior, showing recurrent effusions in body cavities. The clinical outcome and typical prognosis in patients with PEL are poor and potentially lethal. Clarification of the pathogenesis in PEL is urgently needed in order to develop novel therapies. PEL cells generally lack B-cell surface markers, and we therefore hypothesized that the B-cell transcription factor, PAX5, would be down-regulated in PEL. The expression of PAX5 is detected from the pro-B to the mature B-cell stage and is indispensable for the differentiation of B-cells. PAX5 was silenced in PEL cells via its promoter methylation. Up-regulation of PAX5 induced several genes coding for B-cell surface marker mRNA, but not protein level. PAX5 inhibited cell growth via G1 cell cycle arrest. PAX5 bound to RB and increased its protein expression. RB/E2F-regulated genes were significantly down-regulated in microarray analysis and PCR experiments. To elucidate the in vivo role of PAX5, we examined the restoration of PAX5 in a PEL mouse model. The ascites volume and organ invasions were significantly suppressed by PAX5 restoration. Reduction of PAX5 has played a crucial role in the oncogenesis of PEL, and PAX5 is a tumor suppressor in PEL. Targeting PAX5 could represent a novel therapeutic strategy for patients with PEL., Competing Interests: Declaration of competing interest 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 paper., (Copyright © 2024. Published by Elsevier Inc.)

Viruses normally reprogram the host cell metabolic pathways as well as metabolic sensors to facilitate their persistence. The serine-threonine liver kinase B1 (LKB1) is a master upstream kinase of 5'-AMP-activated protein kinase (AMPK) that senses the energy status and therefore regulates the intracellular metabolic homeostasis. Previous studies showed that AMPK restricts Kaposi's sarcoma-associated herpesvirus (KSHV) lytic replication in endothelial cells during primary infection and promotes primary effusion lymphoma (PEL) cell survival. However, the role of LKB1 in KSHV lytic reactivation and KSHV-associated malignancies is unclear. In this study, we found that LKB1 is phosphorylated or activated in KSHV-positive PEL cells. Mechanistically, KSHV-encoded vCyclin mediated LKB1 activation in PEL cells, as vCyclin knockout ablated, while vCyclin overexpression enhanced LKB1 activation. Furthermore, knockdown of LKB1 inactivated AMPK and induced KSHV reactivation, as indicated by the increased expression of viral lytic genes and the increased virions in supernatants. Accordingly, AMPK inhibition by functional knockdown or a pharmacologic inhibitor, Compound C, promoted KSHV reactivation in PEL cells. Furthermore, inhibition of either LKB1 or AMPKα1 efficiently induced cell death by apoptosis of PEL cells both in vitro and in vivo . Together, these results identify LKB1 as a vulnerable target for PEL, which could be potentially exploited for treating other virus-associated diseases.IMPORTANCEKaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic virus associated with several human cancers, such as primary effusion lymphoma (PEL). Here, we showed that serine-threonine liver kinase B1 (LKB1), upstream of 5 ' AMP-activated protein kinase (AMPK), is activated by KSHV-encoded vCyclin and maintains KSHV latency in PEL cells. Inhibition of either LKB1 or AMPK enhances KSHV lytic replication from latency, which at least partially accounts for PEL cell death by apoptosis. Compound C, a potent AMPK inhibitor, induced KSHV reactivation and efficiently inhibited PEL progression in vivo . Thus, our work revealed that LKB1 is a potential therapeutic target for KSHV-associated cancers., Competing Interests: The authors declare no conflict of interest.

A 67-year-old man underwent renal transplantation in his twenties. He developed refractory pleural effusion, with many large lymphocytes with severe atypia and mitosis in the effusion, indicating malignant lymphoma. He finally died of respiratory failure. An autopsy revealed atypical lymphocytes positive for CD3, CD4, and CD30 and negative for CD8, CD20, PAX5, human herpesvirus (HHV) 8, and Epstein-Barr virus-encoded small RNAs by immunohistochemistry and in situ hybridization. Atypical lymphocytes also had T-cell receptor gene rearrangements Jβ2, Jγ2, and Jδ1 and chromosomal aberrations der(8)t(1;8)(q21;p21), add(13)(q12), add(14)(q32), and add(16)(q12-13). A few atypical lymphocytes were present at other sites. We finally diagnosed this case as monomorphic T-cell post-transplant lymphoproliferative disorder with features of HHV8-negative primary effusion lymphoma. A literature review only identified six cases (four HHV8-negative, two HHV8-positive) of effusion lymphoma of T-cell type, including the present case. Interestingly, about half of HHV8-negative and HHV8-positive cases had a history of renal transplantation in their twenties. All cases showed tumor CD30 expression, whereas CD4 and CD8 expressions were inconsistent. These findings indicated that this lymphoma may be associated with post-transplant lymphoproliferative disorder by renal transplantation at a young age, although further cases need to be analyzed., (© 2024. The Author(s).)

Primary effusion lymphoma (PEL) is an uncommon B-cell lymphoma associated with human herpesvirus 8 and comprises 3-4% of all HIV-related lymphomas. It traditionally presents as a pleural, pericardial, and/or peritoneal effusion, though it can occasionally manifest as an extracavitary or solid mass in the absence of an effusion. The extracavitary or solid variant of primary effusion lymphoma has been reported in the skin, gastrointestinal tract, lung, and lymph nodes. However, very few cases have been reported in the central nervous system. We describe a case of extracavitary or solid variant of primary effusion lymphoma presenting as a brain mass in an HIV-positive man, highlighting the clinicopathologic and immunophenotypic findings of a rare entity.

Primary effusion lymphoma (PEL) is an aggressive neoplasm often diagnosed in immunosuppressed patients demonstrating peritoneal, pleural, or pericardial effusions. This high-grade lymphoma is strongly associated with human herpesvirus 8 (HHV8) infection and most of the lesions also show the presence of Epstein-Barr virus in tumor cells, which lacks CD20 expression and reveals a plasmablastic morphology and phenotype. The extracavitary or solid variant of PEL is even rarer and usually affects the lymph nodes and is currently considered a clinical manifestation of the classic PEL. In the oral cavity, extracavitary PEL is extremely rare and only a few patients have been previously reported, with no detailed clinicopathological description. The recognition of oral extracavitary PEL is even more important given the occurrence of plasmablastic lymphoma in the oral mucosa, which shares many clinical, microscopic, and phenotypic features with PEL, therefore, demanding from pathologists the search for HHV8, especially in immunosuppressed patients, and an appropriate clinical evaluation. In this report, we aim to describe a very rare extracavitary PEL affecting the palate of a 36-year-old patient and to review the literature regarding the extracavitary presentation of this aggressive lymphoma. This report demonstrates the importance of searching for HHV8 infection in oral lymphomas with plasmablastic features., Competing Interests: Declaration of Conflicting InterestsThe authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

The 2022 European Association for Haematopathology/Society for Hematopathology lymphoma workshop session on cavity-based lymphomas included sixty-eight cases in seven sections. The disease entities discussed include primary effusion lymphomas (PEL), extracavitary primary effusion lymphomas and confounding entities (ECPEL), HHV8-negative B-lineage lymphomas-effusion based (EBV-negative, EBV-positive, and plasmablastic types), diffuse large B-cell lymphoma associated with chronic inflammation, fibrin-associated diffuse large B-cell lymphoma (FA-DLBCL), breast implant-associated anaplastic large cell lymphoma (BIA-ALCL), and other lymphomas presenting as an effusion. All entities above are discussed; however, three are delved into greater detail given the challenges with classification: ECPEL, HHV8-negative effusion-based lymphomas, and FA-DLBCL. Cases exemplifying the diagnostic difficulty in differentiating ECPEL from HHV8-positive diffuse large B-cell lymphoma and germinotropic lymphoproliferative disorder were discussed. The more recently recognized effusion-based HHV8-negative large B-cell lymphoma is explored, with several cases submitted raising the question if this subset should be carved out as a specific entity, and if so, what should be the refining diagnostic criteria. Case submissions to the FA-DLBCL section yielded one of the largest case series to date, including classic cases, cases furthering the discussion on disease sites and prognosis, as well as novel concepts to be considered in this entity. The 2022 EA4HP/SH workshop cases allowed for further confirmation of the characteristics of some of the more historically accepted cavity-based lymphomas, as well as further inquiry and debate on relatively new or evolving entities., (© 2023. The Author(s).)

Background/aim: Primary effusion lymphoma (PEL) is classified as a rare non-Hodgkin's B-cell lymphoma that is caused by Kaposi's sarcoma-associated herpesvirus (KSHV); PEL cells are latently infected with KSHV. PEL is frequently resistant to conventional chemotherapies. Therefore, the development of novel therapeutic agents is urgently required. Nigericin, a H + and K + ionophore, possesses unique pharmacological effects. However, the effects of nigericin on PEL cells remain unknown., Materials and Methods: We examined the cytotoxic effects of the K + ionophores, nigericin, nonactin, and valinomycin, on various B-lymphoma cells including PEL. We also evaluated ionophore-induced changes in signaling pathways involved in KSHV-induced oncogenesis. Moreover, the effects of nigericin on mitochondrial membrane potential and viral reactivation in PEL were analyzed., Results: Although the three tested ionophores inhibited the proliferation of several B-lymphoma cell lines, nigericin inhibited the proliferation of PEL cells compared to KSHV-negative cells. In PEL cells, nigericin disrupted the mitochondrial membrane potential and caused the release of cytochrome c, which triggered caspase-9-mediated apoptosis. Nigericin also induced both an increase in phosphorylated p38 MAPK and proteasomal degradation of β-catenin. Combination treatment of nigericin with the p38 MAPK inhibitor SB203580 potentiated the cytotoxic effects towards PEL cells, compared to either compound alone. Meanwhile, nigericin did not influence viral replication in PEL cells., Conclusion: Nigericin induces apoptosis in PEL cells by mitochondrial dysfunction and down-regulation of Wnt/β-catenin signaling. Thus, nigericin is a novel drug candidate for treating PEL without the risk of de novo KSHV infection., (Copyright © 2023 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Chronic infection with Kaposi's sarcoma-associated herpes virus (KSHV) in B lymphocytes causes primary effusion lymphoma (PEL), the most aggressive form of KSHV-related cancer, which is resistant to conventional chemotherapy. In this study, we report that the BCBL-1 KSHV + PEL cell line does not harbor oncogenic mutations responsible for its aggressive malignancy. Assuming that KSHV viral oncogenes play crucial roles in PEL proliferation, we examined the effect of cyclin-dependent kinase 9 (CDK9) inhibitor FIT-039 on KSHV viral gene expression and KSHV + PEL proliferation. We found that FIT-039 treatment impaired the proliferation of KSHV + PEL cells and the expression of KSHV viral genes in vitro. The effects of FIT-039 treatment on PEL cells were further evaluated in the PEL xenograft model that retains a more physiological environment for the growth of PEL growth and KSHV propagation, and we confirmed that FIT-039 administration drastically inhibited PEL growth in vivo. Our current study indicates that FIT-039 is a potential new anticancer drug targeting KSHV for PEL patients., (© 2023. The Author(s).)

Kaposi's sarcoma-associated herpesvirus (KSHV) causes several malignancies in people living with HIV, including primary effusion lymphoma (PEL). PEL cell lines exhibit oncogene addictions to both viral and cellular genes. Using CRISPR screens, we previously identified cellular oncogene addictions in PEL cell lines, including MCL1. MCL1 is a member of the BCL2 family, which functions to prevent intrinsic apoptosis and has been implicated in several cancers. Despite the overlapping functions of the BCL2 family members, PEL cells are dependent only on MCL1, suggesting that MCL1 may have nonredundant functions. To investigate why PEL cells exhibit selective addiction to MCL1, we inactivated the intrinsic apoptosis pathway by engineering BAX/BAK1 double knockout cells. In this context, PEL cells become resistant to MCL1 knockdown or MCL1 inactivation by the MCL1 inhibitor S63845, indicating that the main function of MCL1 in PEL cells is to prevent BAX/BAK1-mediated apoptosis. The selective requirement to MCL1 is due to MCL1 being expressed in excess over the BCL2 family. Ectopic expression of several BCL2 family proteins, as well as the KSHV BCL2 homolog, significantly decreased basal caspase 3/7 activity and buffered against staurosporine-induced apoptosis. Finally, overexpressed BCL2 family members can functionally substitute for MCL1, when it is inhibited by S63845. Together, our data indicate that the expression levels of the BCL2 family likely explain why PEL tumor cells are highly addicted to MCL1. Importantly, our results suggest that caution should be taken when considering MCL1 inhibitors as a monotherapy regimen for PEL because resistance can develop easily. IMPORTANCE Primary effusion lymphoma (PEL) is caused by Kaposi's sarcoma-associated herpesvirus. We showed previously that PEL cell lines require the antiapoptotic protein MCL1 for survival but not the other BCL2 family proteins. This selective dependence on MCL1 is unexpected as the BCL2 family functions similarly in preventing intrinsic apoptosis. Recently, new roles for MCL1 not shared with the BCL2 family have emerged. Here, we show that noncanonical functions of MCL1 are unlikely essential. Instead, MCL1 functions mainly to prevent apoptosis. The specific requirement to MCL1 is due to MCL1 being expressed in excess over the BCL2 family. Consistent with this model, shifting these expression ratios changes the requirement away from MCL1 and toward the dominant BCL2 family gene. Together, our results indicate that although MCL1 is an attractive chemotherapeutic target to treat PEL, careful consideration must be taken, as resistance to MCL1-specific inhibitors easily develops through BCL2 family overexpression.

Rare cases of human herpesvirus 8 (HHV8)-negative effusion-based large B-cell lymphoma (EB-LBCL) occur in body cavities without antecedent or concurrent solid mass formation. In contrast to HHV8 + primary effusion lymphoma (PEL), EB-LBCL has no known association with HIV or HHV8 infection. However, the small sample sizes of case reports and series worldwide, especially from non-Japanese regions, have precluded diagnostic uniformity. Therefore, we conducted a retrospective, multi-institutional study of 55 cases of EB-LBCL and performed a comprehensive review of an additional 147 cases from the literature to identify distinct clinicopathologic characteristics. In our study, EB-LBCL primarily affected elderly (median age 80 years), immunocompetent patients and manifested as lymphomatous effusion without a solid component. The lymphomatous effusions mostly occurred in the pleural cavity (40/55, 73%), followed by the pericardial cavity (17/55, 31%). EB-LBCL expressed CD20 (53/54, 98%) and PAX5 (23/23, 100%). Most cases (30/36, 83%) were of non-germinal center B-cell subtype per the Hans algorithm. HHV8 infection was absent (0/55, 0%), while Epstein-Barr virus was detected in 6% (3/47). Clinically, some patients were managed with drainage alone (15/34, 44%), while others received rituximab alone (4/34, 12%) or chemotherapy (15/34, 44%). Eventually, 56% (22/39) died with a median overall survival (OS) of 14.9 months. Our findings were similar to those from the literature; however, compared to the non-Japanese cases, the Japanese cases had a significantly higher incidence of pericardial involvement, a higher rate of chemotherapy administration, and longer median OS. Particularly, we have found that Japanese residence, presence of pericardial effusion, and absence of MYC rearrangement are all favorable prognostic factors. Our data suggest that EB-LBCL portends a worse prognosis than previously reported, although select patients may be managed conservatively. Overall, EB-LBCL has distinct clinicopathologic characteristics, necessitating the establishment of separate diagnostic criteria and consensus nomenclature., (© 2022. The Author(s), under exclusive licence to United States & Canadian Academy of Pathology.)

Primary effusion lymphoma (PEL) is an aggressive B-cell non-Hodgkin lymphoma in immunocompromised individuals such as AIDS patients. PEL shows a poor prognosis (median survival time < 6 months) compared with other AIDS-related lymphomas, and is generally resistant to conventional treatments. Novel drugs for PEL treatment are required. Midkine inhibitor (iMDK) was previously found to suppress midkine protein expression. Interestingly, iMDK suppressed cell proliferation in PEL cell lines in a time- and dose-dependent manner, regardless of midkine gene expression. We examined the mechanism of iMDK on PEL. Importantly, iMDK strongly induced cell cycle arrest at the G2/M phase within 12 h of incubation and suppressed the p-CDK1 protein level, which is associated with the cell cycle checkpoint at G2/M, resulting in mitotic catastrophe with observation of multipolar division. After mitotic catastrophe, iMDK-treated PEL showed apoptosis with caspase-3, - 8, and - 9 activation at 24 h incubation. However, iMDK showed no effects on viral protein-activated signaling pathways such as JAK-STAT, PI3K-Akt and NF-κB, and HHV-8/KSHV gene expression in PEL. These results indicate that iMDK is a novel CDK1 inhibitor and a promising lead compound for PEL chemotherapy treatment., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Primary effusion lymphoma (PEL) is associated with human herpesvirus 8 and frequently with Epstein-Barr virus (EBV). We report here a single-center series of 19 human immunodeficiency virus-associated PELs, including 14 EBV+ and 5 EBV- PELs. The objectives were to describe the clinicopathologic features of PELs, with a focus on programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) expression, to search for genetic alterations by targeted deep sequencing analysis, and to compare the features between EBV+ and EBV- cases. All the patients were male, and the median age at diagnosis was 47 years old (interquartile range: 40 to 56 y). Reflecting the terminal B-cell differentiation, immunophenotypic profiles showed low expression levels of B-cell markers, including CD19 (0/19), CD20 (1/19), CD79a (0/19), PAX5 (1/19), BOB1 (3/19), and OCT2 (4/19), contrasting with a common expression of CD38 (10/19), CD138 (7/19), and IRF4/MUM1 (18/19). We observed a frequent aberrant expression of T-cell markers, especially CD3 (10/19), and less frequently CD2 (2/19), CD4 (3/19), CD5 (1/19), and CD8 (0/19). Only 2 cases were PD-L1 positive on tumor cells and none PD-1 positive. With respect to immune cells, 3 samples tested positive for PD-L1 and 5 for PD-1. Our 36-gene lymphopanel revealed 7 distinct variants in 5/10 PELs, with either a single or 2 mutations per sample: B2M (n=2), CD58 (n=1), EP300 (n=1), TNFAIP3 (n=1), ARID1A (n=1), and TP53 (n=1). Finally, we did not observe any major clinical, pathologic, or immunohistochemical differences between EBV+ and EBV- PELs and the outcome was similar (2-y overall survival probability of 61.9% [95% confidence interval, 31.2-82.1] vs. 60.0% [95% confidence interval, 12.6-88.2], respectively, P=0.62)., Competing Interests: Conflicts of Interest and Source of Funding: Supported by institutional grants from INSERM and the Institut Carnot CALYM. 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.)

Lymphomas remain a leading cause of morbidity and mortality for HIV-positive patients. The most common lymphomas include diffuse large B-cell lymphoma, Burkitt lymphoma, primary effusion lymphoma, plasmablastic lymphoma and Hodgkin lymphoma. Appropriate approach is determined by lymphoma stage, performans status, comorbidities, histological subtype, status of the HIV disease and immunosuppression. Treatment outcomes have improved due to chemotherapy modalities and effective antiretroviral therapy. This review summarizes epidemiology, pathogenesis, pathology, and current treatment landscape in HIV associated lymphoma., (Copyright © 2021 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.)

Viral infection has been implicated in the pathogenesis of a plethora of human diseases. Although antiviral therapies effectively confront the viral spread and infection, how to completely eradicate the viral genome from infected cells remains a challenge. In this study, we demonstrated the reversible switching of primary cells between normal and malignant states by an oncogenic virus Kaposi's sarcoma-associated herpesvirus (KSHV) and CRISPR/Cas9-mediated targeting of a major viral latent protein. Primary cells can be transformed into malignant status by infection of KSHV, while elimination of the KSHV genome from latent KSHV-infected cells reverses KSHV-transformed primary cells back to a "normal state" by CRISPR/Cas-mediated knockout of viral major latent gene LANA. As a proof of concept, we demonstrated efficient elimination of KSHV episome in KSHV-associated primary effusion lymphoma cells resulting in the induction of apoptosis by liposome-encapsulated CRISPR/Cas9 ribonucleoprotein complexes (Lipo/Cas9-LANAsgRNA). Our work illustrates CRISPR/Cas as a promising technology for eliminating oncogenic viruses from persistently infected cells by taking advantage of the genetic differences between viral and cellular genomes. Compared to traditional antiviral therapy, our study offer an approach for antagonizing human oncogenic virus-related cancers by directly targeting as well as clearing viral genomes., (© 2021 Wiley Periodicals LLC.)

Statins are inhibitors of the mevalonate pathway that besides being cholesterol lowering agents, display anti-cancer properties. This is because cholesterol is an essential component of cell membranes but also because the mevalonate pathway controls protein farnesylation and geranylation, processes essential for the activity of GTPase family proteins. In this study, we found that Lovastatin exerted a dose- and time-dependent cytotoxic effect against PEL cells, an aggressive B cell lymphoma strictly associated with the gammaherpesvirus KSHV and characterized by a poor response to conventional chemotherapies. At molecular level, Lovastatin by dephosphorylating STAT3, induced ERK1/2 activation that inhibited autophagy and phosphorylated p53ser15 that in turn maintained ERK1/2 activated and up-regulated p21. However, p21 played a pro-survival role in this setting, as its inhibition by UC2288 further reduced cell survival in PEL cells undergoing Lovastatin treatment. In conclusion, this study suggests that Lovastatin may represent a valid therapeutic alternative against PEL cells, especially if used in combination with p21 inhibitors., (© 2021 The Authors. IUBMB Life published by Wiley Periodicals LLC on behalf of International Union of Biochemistry and Molecular Biology.)

We present a rare case of primary effusion lymphoma (PEL) in a 75 year old, HIV-negative male patient with multiple comorbidities. Imaging studies revealed a massive right pleural effusion and a significant lung collapse with multiple plural soft tissue nodules. Immediate thoracentesis was performed. Cytologic evaluation of the pleural fluid showed abnormally large cell with increased nuclear to cytoplasmic ratio, irregular nuclear contours and prominent nucleoli, with phenotypic expression of HHV-8, CD138, CD30, and MUM1 markers and negative staining for epithelial and mesothelial markers. PEL is a rare and aggressive large B-cell lymphoma often affecting immunocompromised adults and is mostly associated with human herpes virus 8/Kaposi sarcoma-associated herpes virus (HHV-8/KSHV). However, cases in immunocompetent elderly patients have been reported. The cytomorphologic features of PEL overlaps with those of aggressive lymphomas such as diffuse large B-cell lymphoma, plasmablastic lymphoma, and anaplastic large-cell lymphoma. Also, mesothelioma, metastatic carcinoma or melanoma should be considered in the differential diagnosis. Hence, PEL should be kept in mind in the diagnostic algorithm of cytological evaluation of serosal fluid not only in HIV positive patients but also HIV-negative elderly patients. In this report, we aim to highlight the cytologic and immunohistochemical staining pattern of this rare entity to increase awareness of this entity among cytopathologists., (© 2021 Wiley Periodicals LLC.)

Background: Primary effusion lymphoma (PEL) is an aggressive B cell non-Hodgkin lymphoma that develops especially in AIDS patients and immunocompromised patients infected with human herpes virus-8 (HHV-8)/Kaposi's sarcoma-associated herpesvirus (KSHV). PEL has a poor prognosis in patients despite conventional chemotherapeutic treatment, and a safe and efficient therapy is required., Purpose: To examine the effects on PEL of cucurbitacin B (CuB), a triterpene found in plants of the Cucurbitaceae family that has several anti-cancer activities., Study Design: We evaluated the anti-cancer activities of CuB in vitro and in vivo., Methods: Cell proliferation of PEL cell lines was measured by MTT assay. Cleaved caspases and signaling transduction associated proteins were analyzed by western blotting. Wright and Giemsa staining and immunofluorescence staining were carried out to observe cell morphology. Cell cycles were analyzed by flow cytometry. RT-PCR was performed to detect viral gene expressions. A xenograft mouse model was employed to evaluate the anti-cancer activity of CuB in vivo., Results: CuB inhibited cell proliferation of PEL cell lines (BCBL-1, BC-1, GTO and TY-1) in a dose-dependent manner (0-50 nM) and induced apoptosis of BCBL-1 cells via caspase activation in a dose- and time-dependent manner. In addition, CuB caused cell-shape disruption by inducing actin aggregation and suppressing the p-cofilin level, resulting in BCBL-1 cell arrest at the G2/M phase. In contrast, CuB showed almost no suppression of p-STAT3 and p-Akt activation, which were constitutively activated by KSHV-derived proteins. Furthermore, CuB (0.5 mg/kg) via intraperitoneal injection significantly (p < 0.05) suppressed solid tumor growth in the xenograft mouse model., Conclusion: This study suggests that CuB is a promising agent for PEL treatment., (Copyright © 2021. Published by Elsevier GmbH.)

Background: Primary effusion lymphoma is a rare, aggressive large B-cell lymphoma strictly linked to infection by Human Herpes virus 8/Kaposi sarcoma-associated herpes virus. In its classic form, it is characterized by body cavities neoplastic effusions without detectable tumor masses. It often occurs in immunocompromised patients, such as HIV-positive individuals. Primary effusion lymphoma may affect HIV-negative elderly patients from Human Herpes virus 8 endemic regions. So far, rare cases have been reported in transplanted patients. The purpose of our systematic review is to improve our understanding of this type of aggressive lymphoma in the setting of transplantation, focusing on epidemiology, clinical presentation, pathological features, differential diagnosis, treatment and outcome. The role of assessing the viral serological status in donors and recipients is also discussed., Methods: We performed a systematic review adhering to the PRISMA guidelines. The literature search was conducted on PubMed/MEDLINE, Web of Science, Scopus, EMBASE and Cochrane Library, using the search terms "primary effusion lymphoma" and "post-transplant"., Results: Our search identified 13 cases of post-transplant primary effusion lymphoma, predominantly in solid organ transplant recipients (6 kidney, 3 heart, 2 liver and 1 intestine), with only one case after allogenic bone marrow transplantation. Long-term immunosuppression is important in post-transplant primary effusion lymphoma commonly developing several years after transplantation. Kaposi Sarcoma occurred in association with lymphoma in 4 cases of solid organ recipients. The lymphoma showed the classical presentation with body cavity effusions in absence of tumor masses in 10 cases; 2 cases presented as solid masses, lacking effusions and one case as effusions associated with multiple organ involvement. Primary effusion lymphoma occurring in the setting of transplantation was more often Epstein Barr-virus negative. The prognosis was poor. In addition to chemotherapy, reduction of immunosuppressive treatment, was generally attempted., Conclusions: Primary effusion lymphoma is a rare, but often fatal post-transplant complication. Its rarity and the difficulty in achieving the diagnosis may lead to miss this complication. Clinicians should suspect primary effusion lymphoma in transplanted patients, presenting generally with unexplained body cavity effusions, although rare cases with solid masses are described.

Primary effusion lymphoma (PEL), caused by Kaposi's sarcoma-associated herpesvirus (KSHV), presents as a lymphomatous effusion in body cavities and has a poor prognosis. The anti-malaria drug, artesunate, possesses anti-neoplastic potential. Therefore, we aimed to investigate its effect on KSHV-infected PEL cell lines. Artesunate inhibited cell growth and viability of PEL cells, but its effect on peripheral blood mononuclear cells was less pronounced. Artesunate induced G1 phase arrest by downregulating cyclin D1/D2, CDK2/6 and c-Myc. Artesunate increased reactive oxygen species and DNA damage, but did not affect the expression of latent and lytic genes of KSHV. It exhibited cytotoxicity through caspase-dependent and -independent pathways and reduced Bcl-xL, survivin, XIAP and c-IAP1/2 levels. Furthermore, artesunate suppressed NF-κB and AP-1 by inhibiting IκB kinase and IκBα phosphorylation as well as JunB expression. Finally, artesunate treatment attenuated PEL development in mice. Our data support that artesunate is a potential drug for PEL treatment.

Lymphomas presenting in effusions could either be primary or secondary, with very limited data from Taiwan., Methods: We retrospectively reviewed effusion lymphomas from our archives in a tertiary center from July 2011 to June 2019., Results: We identified 59 specimens from 43 patients, including 7 cases with primary effusion lymphoma (PEL) and 36, secondary effusion involvement. Half of the secondary cases presented concurrently with effusion lymphoma, while the remaining half-experienced effusion lymphoma during disease progression. All patients with PELs were males with a median age of 77 and presented with massive pleural effusion. None was HIV-related. Two (29%) PEL cases were positive for human herpes virus 8 (HHV8). The only case with plasmablastic phenotype in the PEL group was positive for both HHV8 and EBV. Four patients died shortly after diagnosis; while the remaining three were alive at the last follow-up (two at 13 months and one at 99 months). Of the secondary cases, diffuse large B-cell lymphoma/high grade B-cell lymphoma was the most common (n = 16, 44%), followed by mantle cell lymphoma (n = 5, 14%). Only 8 cases (22%) were T-cell neoplasms. Prognosis for patients with secondary effusion involvement was dismal, with 1- and 2-year overall survival rates at 17% and 8%, respectively., Conclusion: We found a wide cytopathological spectrum of effusion lymphoma in Taiwan. Most of our PEL cases were distinct from that defined in the World Health Organization scheme by a B-cell phenotype, HHV8-negativity, and absence of immunodeficiency. As compared to PEL cases, the prognosis of those with secondary involvement was extremely poor., (© 2020 Wiley Periodicals LLC.)

Primary effusion lymphoma (PEL) is an aggressive B cell lymphoma that is etiologically linked to Kaposi's sarcoma-associated herpesvirus (KSHV). Despite standard multi-chemotherapy treatment, PEL continues to cause high mortality. Thus, new strategies to control PEL are needed urgently. Here, we show that a phosphodegron motif within the KSHV protein, latency-associated nuclear antigen (LANA), specifically interacts with E3 ubiquitin ligase FBW7, thereby competitively inhibiting the binding of the anti-apoptotic protein MCL-1 to FBW7. Consequently, LANA-FBW7 interaction enhances the stability of MCL-1 by preventing its proteasome-mediated degradation, which inhibits caspase-3-mediated apoptosis in PEL cells. Importantly, MCL-1 inhibitors markedly suppress colony formation on soft agar and tumor growth of KSHV+PEL/BCBL-1 in a xenograft mouse model. These results strongly support the conclusion that high levels of MCL-1 expression enable the oncogenesis of PEL cells and thus, MCL-1 could be a potential drug target for KSHV-associated PEL. This work also unravels a mechanism by which an oncogenic virus perturbs a key component of the ubiquitination pathway to induce tumorigenesis., Competing Interests: The authors have declared that no competing interests exist.

Pomalidomide (Pom) is an immunomodulatory drug that has efficacy against Kaposi's sarcoma, a tumor caused by Kaposi's sarcoma-associated herpesvirus (KSHV). Pom also induces direct cytotoxicity in primary effusion lymphoma (PEL), a B-cell malignancy caused by KSHV, in part through downregulation of IRF4, cMyc, and CK1α as a result of its interaction with cereblon, a cellular E3 ubiquitin ligase. Additionally, Pom can reverse KSHV-induced downregulation of MHCI and co-stimulatory immune surface molecules ICAM-1 and B7-2 on PELs. Here, we show for the first time that Pom-induced increases in ICAM-1 and B7-2 on PEL cells lead to an increase in both T-cell activation and NK-mediated cytotoxicity against PEL. The increase in T-cell activation can be prevented by blocking ICAM-1 and/or B7-2 on the PEL cell surface, suggesting that both ICAM-1 and B7-2 are important for T-cell co-stimulation by PELs. To gain mechanistic insights into Pom's effects on surface markers, we generated Pom-resistant (PomR) PEL cells, which showed about 90% reduction in cereblon protein level and only minimal changes in IRF4 and cMyc upon Pom treatment. Pom no longer upregulated ICAM-1 and B7-2 on the surface of PomR cells, nor did it increase T-cell and NK-cell activation. Cereblon-knockout cells behaved similarly to the pomR cells upon Pom-treatment, suggesting that Pom's interaction with cereblon is necessary for these effects. Further mechanistic studies revealed PI3K signaling pathway as being important for Pom-induced increases in these molecules. These observations provide a rationale for the study of Pom as therapy in treating PEL and other KSHV-associated tumors., Competing Interests: I have read the journal's policy and the authors of this manuscript have the following competing interests: This research was funded in part by Cooperative Research and Development Agreements (CRADAs) between Celgene Corporation (now Bristol Myers Squibb Co.) and the National Cancer Institute. RY reports receiving drug for clinical trials from Merck Co. and from EMD-Serano through CRADAs with the NCI. Two co-authors (DAD and RY) are co-inventors on US Patent 10,001,483 entitled "Methods for the treatment of Kaposi's sarcoma or KSHV-induced lymphoma using immunomodulatory compounds, and uses of biomarkers". It is our understanding that foreign patents have also been filed for this invention. An immediate family member of RY is a co-inventor on patents related to internalization of target receptors, on KSHV viral IL-6, and on the use of calreticulin and calreticulin fragments to inhibit angiogenesis. These inventions were all made as part of duties as a full-time employees of the US government under 45 Code of Federal Regulations Part 7. All rights, title, and interest to these patents have been or should by law be assigned to the U.S. Department of Health and Human Services. The government conveys a portion of the royalties it receives to its employee-inventors under the Federal Technology Transfer Act of 1986 (P.L. 99-502).

Background: Primary effusion lymphoma (PEL) is a very rare non-Hodgkin lymphoma caused by human herpesvirus-8 (HHV8) that grows in liquid phase within body cavities. The diagnosis of PEL is based on cytology but requires confirmatory ancillary tests. PEL occurs mainly in association with HIV infection. This study describes 9 cases of PEL in HIV-negative patients and compares their characteristics with 10 HIV-associated cases of PEL diagnosed at a single institution in Italy between 1995 and 2019., Methods: Clinical records were reviewed for demographic data, comorbidities, laboratory abnormalities, and outcome. PEL samples were evaluated for cytomorphology, immunophenotype, immunoglobulin (IG)/T cell receptor (TR) rearrangements, and HHV8 and Epstein-Barr virus (EBV) viral loads in effusion supernatants., Results: HIV-unrelated PEL occurred in 8 elderly patients (7 men, 1 woman) and 1 young adult with primary antibody deficiency. Cytology revealed HHV8-positive lymphoma cells lacking B/T cell antigens and exhibiting 2 cell patterns (polymorphous or monotonous). IG was clonally rearranged in all cases; aberrant TRG occurred in 2 cases. Effusion supernatants had more than 10 6 HHV8 DNA copies per mL and variable loads of EBV DNA. Compared with HIV-associated PEL, the HIV-negative cohort was characterized by older age, less frequent association with Kaposi sarcoma and/or multicentric Castleman disease, comparable but less abnormal laboratory parameters, and a nonsignificant survival benefit. PEL cases with low apoptosis were associated with better prognosis., Conclusion: To the best of our knowledge, our case series of HIV-unrelated PEL is the largest thus far, expands the spectrum of cytological findings, and supports the need for a multidisciplinary approach in the diagnostic workup., (© 2020 American Cancer Society.)

Kaposi's sarcoma-associated herpesvirus (KSHV, HHV-8) is a gamma herpesvirus associated with several human malignancies. Transposable elements (TEs) are ubiquitous in eukaryotic genomes, occupying about 45% of the human genome. TEs have been linked with a variety of disorders and malignancies, though the precise nature of their contribution to many of them has yet to be elucidated. Global transcriptome analysis for differentially expressed TEs in KSHV-associated primary effusion lymphoma (PEL) cells (BCBL1 and BC3) revealed large number of differentially expressed TEs. These differentially expressed TEs include LTR transposons, long interspersed nuclear elements (LINEs), and short interspersed nuclear elements (SINEs). Further analysis of LINE-1 (L1) elements revealed expression upregulation, hypo-methylation, and transition into open chromatin in PEL. In agreement with high L1 expression, PEL cells express ORF1 protein and possess high reverse transcriptase (RT)-activity. Interestingly, inhibition of this RT-activity suppressed PEL cell growth. Collectively, we identified high expression of TEs, and specifically of L1 as a critical component in the proliferation of PEL cells. This observation is relevant for the treatment of KSHV-associated malignancies since they often develop in AIDS patients that are treated with RT inhibitors with potent inhibition for both HIV and L1 RT activity.

Primary effusion lymphoma (PEL) has a very poor prognosis. To evaluate the contributions of enhancers/promoters interactions to PEL cell growth and survival, here we produce H3K27ac HiChIP datasets in PEL cells. This allows us to generate the PEL enhancer connectome, which links enhancers and promoters in PEL genome-wide. We identify more than 8000 genomic interactions in each PEL cell line. By incorporating HiChIP data with H3K27ac ChIP-seq data, we identify interactions between enhancers/enhancers, enhancers/promoters, and promoters/promoters. HiChIP further links PEL super-enhancers to PEL dependency factors MYC, IRF4, MCL1, CCND2, MDM2, and CFLAR. CRISPR knock out of MEF2C and IRF4 significantly reduces MYC and IRF4 super-enhancer H3K27ac signal. Knock out also reduces MYC and IRF4 expression. CRISPRi perturbation of these super-enhancers by tethering transcription repressors to enhancers significantly reduces target gene expression and reduces PEL cell growth. These data provide insights into PEL molecular pathogenesis.

Primary effusion lymphoma is a rare, clinically aggressive large B-cell neoplasm universally associated with human herpesvirus 8 that occurs in the setting of immune compromise. It is classically described as a lymphomatous effusion occurring within body cavities. Recently, however, solid tumor masses, and rarely an intravascular form, have been described. We report a case of a cutaneous intravascular primary effusion lymphoma occurring within ectatic vascular spaces of a Kaposi sarcoma skin lesion in a human immunodeficiency virus-positive adult. Human herpesvirus 8 immunohistochemistry was positive in the nuclei of the Kaposi sarcoma spindled cells as well as within large intravascular plasmacytoid cells. This unusual case highlights the importance of careful assessment of the nature of human herpesvirus 8-positive staining cells in an otherwise typical Kaposi sarcoma. A careful search for dual pathology in immune-compromised patients as well as the importance of histologic assessment of skin lesions in human immunodeficiency virus-positive patients is also highlighted.

Primary effusion lymphoma (PEL) is an aggressive malignancy with poor prognosis even under chemotherapy. Kaposi sarcoma-associated herpesvirus (KSHV), one of the human oncogenic viruses, is the principal causative agent. Currently, there is no specific treatment for PEL; therefore, developing new therapies is of great importance. Sphingolipid metabolism plays an important role in determining the fate of tumor cells. Our previous studies have demonstrated that there is a correlation between sphingolipid metabolism and KSHV+ tumor cell survival. To further develop sphingolipid metabolism-targeted therapy, after screening a series of newly synthesized ceramide analogs, here, we have identified compounds with effective anti-PEL activity. These compounds induce significant PEL apoptosis, cell-cycle arrest, and intracellular ceramide production through regulation of ceramide synthesizing or ceramide metabolizing enzymes and dramatically suppress tumor progression without visible toxicity in vivo. These new compounds also increase viral lytic gene expression in PEL cells. Our comparative transcriptomic analysis revealed their mechanisms of action for inducing PEL cell death and identified a subset of novel cellular genes, including AURKA and CDCA3, controlled by sphingolipid metabolism, and required for PEL survival with functional validation. These data provide the framework for the development of promising sphingolipid-based therapies against this virus-associated malignancy., (© 2020 by The American Society of Hematology.)

Introduction: Human herpes virus 8-related lymphoproliferative disorders are a complex and heterogeneous group of entities and some of them are eminently diagnosed by cytopathology. In a routine laboratory, these lesions account for less than 1% of the effusion fluids samples. However, they represent up to 30% of all the lymphoma diagnosis from effusion cytological samples and their consideration in the diagnostic flow chart is mandatory, especially in human immunodeficiency virus-positive patients., Methods: A retrospective series of cytological specimens from cavity effusions (n = 605) were analysed. Five human herpes virus 8-related lymphoproliferative processes were recruited. A combination of morphological criteria (enhanced with May-Grünwald Giemsa staining), cell block-based immunocytochemistry and flow cytometry were undertaken for final characterisation., Results: The identification of malignant cells may be difficult. Some specimens are particularly rich, easily leading to suspect a lymphoproliferative process, whereas in other cases, the presence of abundant reactive mesothelial cells, histiocytes, neutrophils, small reactive T and B lymphocytes may obscure the neoplastic process. The biological behaviour may be very heterogeneous and a standardised therapy for these cases is still lacking, although some patients may benefit from antiretroviral therapy in a human immunodeficiency virus setting., Conclusions: The present case series highlights some characteristic findings of these entities to reaffirm useful cytopathological diagnostic criteria, stressing the crucial role of the appropriate technical processing of effusion fluids to obtain the best performances., (© 2020 John Wiley & Sons Ltd.)

Primary Effusion Lymphoma (PEL) is a B-cell lymphoma associated with Kaposi's sarcoma herpesvirus (KSHV) infection. However, the mechanism of oncogenesis of PEL is still unclear. Studies have shown that the cellular transcriptional coactivator p300 regulates the interaction between host and virus, which plays a vital role in viral replication. In this study, we investigated the role of p300 in BCBL1 cells during the KSHV life cycle. We found that p300 knockout resulted in an overall increase for the early lytic genes and changed the expression of genes associated with tumor development, proliferation, and the immune response in the KSHV infected B cells. However, knockout of p300 significantly inhibited the expression of the immediate-early gene RTA and the late lytic gene K8 after KSHV lytic activation. Additionally, the intracellular KSHV genome copy number and the virion production were reduced. These results demonstrated that p300 plays a crucial role in suppressing KSHV viral replication in BCBL1. Furthermore, we observed that the growth of BCBL1 was inhibited by knockout of p300, which confirmed our findings that p300 promotes cell proliferation. This study further provided evidence that p300 plays an important role in the pathogenesis of BCBL1, which might lead to the oncogenesis of PEL caused by KSHV infection., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Primary effusion lymphoma (PEL) is defined as a HHV-8-associated large B-cell lymphoma, which favors HIV-infected young adults, typically presenting as a serous (pleural, pericardial, or peritoneal) effusion with no identifiable tumor mass. Uncommon instances of lymphoid proliferations with the same morphology, immunophenotype, and molecular features as PEL, but occurring as a solid tumor mass without serous cavities involvement, have been termed extracavitary (or solid) variant of PEL. We hereby report the exceptional case of a HIV-associated extracavitary PEL primarily localized to the skin and exhibiting a panniculitis-like presentation. Primary cutaneous presentation of extracavitary PEL is exceedingly uncommon, with only 6 cases previously described in the literature. In light of its atypical immunophenotype, the differential diagnosis in case of skin involvement by extracavitary PEL is challenging: demonstration of HHV-8 infection in neoplastic cells is of pivotal importance. Our case is further atypical in that the lymphoid proliferation underwent complete and protracted regression solely by establishment of highly active antiretroviral therapy.

The current literature credits Keech and Creech with the first report of breast implant-associated anaplastic large cell lymphoma (BIA-ALCL) in 1997. Here we discuss what we consider is the first ever description of BIA-ALCL, a recently recognized entity by the WHO. We unearthed the description of a patient that was diagnosed with primary effusion lymphoma (PEL), surrounding a breast implant in 1996. In light of the current state of knowledge, we evaluated the evidence presented in 1996 and consider that BIA-ALCL is a more appropriate diagnosis rather than PEL. We base our proposal on the following features: 1). clinical presentation of effusion around a breast implant, 2). occurring in an HIV negative patient, 3). absence of EBV co-infection, and 4). a historically questionable demonstration of HHV8. In effect we further support that HHV8 is not related with BIA-ALCL based on the following facts: 1). An extensive review of the literature did not disclose a similar case in the next 24 years, 2). Use of state of the art HHV8 by immunohistochemistry did not disclose any positive case among 30 randomly tested cases. We believe this matter is of importance because in the current WHO, the assertion that PEL is a possible complication of breast implants may lead to a diagnosis with poor prognosis and susceptible of morbidity related with aggressive therapy, in contrast with BIA-ALCL that can be cured with surgery alone., (Copyright © 2020. Published by Elsevier Inc.)

Primary effusion lymphoma (PEL) is a subtype of non-Hodgkin lymphoma associated with infection by Kaposi sarcoma-associated herpes virus (KSHV). PEL is an aggressive disease with extremely poor prognosis when treated with conventional chemotherapy. Narciclasine, a natural product present in Amaryllidaceae family of flowering plants including daffodils, belongs to a class of molecules termed 'isocarbostyril alkaloid'. We have found that narciclasine displays preferential cytotoxicity towards PEL at low nanomolar concentrations and is approximately 10 and 100-fold more potent than its structural analogs lycoricidine and lycorine, respectively. Narciclasine arrested cell-cycle progression at the G 1 phase and induced apoptosis in PEL, which is accompanied by activation of caspase-3/7, cleavage of PARP and increase in the surface expression of Annexin-V. Although narciclasine treatment resulted in a marked decrease in the expression of MYC and its direct target genes,time-course experiments revealed that MYC is not a direct target of narciclasine. Narciclasine treatment neither induces the expression of KSHV-RTA/ORF50 nor the production of infectious KSHV virions in PEL. Finally, narciclasine provides dramatic survival advantages to mice in two distinct mouse xenograft models of PEL. In conclusion, our results suggest that narciclasine could be a promising agent for the treatment of PEL.

Plasmablastic neoplasms encompass several entities including plasmablastic lymphoma, plasmablastic plasmacytoma/multiple myeloma, primary effusion lymphoma and its extracavitary variant, anaplastic lymphoma kinase-positive large B-cell lymphoma, and Kaposi sarcoma-associated herpesvirus/human herpesvirus 8 (HHV8)-positive diffuse large B-cell lymphoma, not otherwise specified. Morphologically, the tumor cells are large with eccentrically located nuclei, prominent nucleoli, and basophilic/amphophilic cytoplasm. Immunophenotypically, the tumor cells express plasma cell-related antigens including CD38, CD138, interferon regulatory factor-4 (IRF4)/MUM1, PR domain zinc finger protein-1 (PRDM1), and/or X-box binding protein-1 (XBP1), with frequent loss of CD20. These tumors are diagnostically challenging for general pathologists due to their overlapping morphology and immunophenotype, and due to their rarity, and particularly so when clinical and radiologic information is insufficient. We also discuss HHV8-negative effusion-based lymphoma due to its overlapping features with primary effusion lymphoma. In this review, we focus on the useful diagnostic markers and pertinent molecular findings in these distinct entities and propose a practical diagnostic algorithm using anaplastic lymphoma kinase, HHV8, in situ hybridization for Epstein-Barr virus-encoded small RNA, immunoglobulin M, light chain stains, and clinicoradiologic criteria to avoid misdiagnosis. At the molecular level, MYC protein overexpression with or without MYC rearrangement and PRDM1-inactivating mutations or deletions are noted in a subset of such tumors, especially in plasmablastic lymphoma. Prognosis in these entities is dismal with conventional CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) chemotherapy. Therefore, novel target therapies, such as anti-CD30 agents, and/or immune blockade therapy, are potential treatment options in the future.

Primary effusion lymphoma (PEL) is a rare form of aggressive B-cell lymphoma characterized by a malignant serous effusion involving body cavities. It usually associated with human herpes virus-8 (HHV-8) and coexpression of Epstein-Barr virus and mostly affects patients with HIV. We report a rare case of cutaneous PEL with an unusual intravascular presentation, combined with Kaposi sarcoma involving the skin, lung, and gastrointestinal tract. The molecular genetic analysis of the sarcoma and lymphoma components, using next-generation sequencing was performed. The patient was a 67-year-old man who presented with multiple cutaneous tumors and mass in the left lung. He died 17 hours after the admission to the hospital. At autopsy, in addition to the cutaneous lesions, tumors in the left lung and gastrointestinal mucosa were detected, and no effusions in the body cavities were seen. The biopsy from the cutaneous lesions, pulmonary, and intestinal tumors revealed histological and immunohistochemical features of Kaposi sarcoma. In addition, the skin biopsy specimens contained a diffuse infiltrate composed of large pleomorphic cells, with focal intravascular growth that were negative for pan B-cell markers, weakly positive for CD38 and CD138 but expressed CD3, HHV-8, and Epstein-Barr virus. Molecular genetic studies in this specimen revealed monoclonal rearrangements of the IgH gene. The diagnosis of PEL, solid variant, was made. Next-generation sequencing analysis of the tumorous and normal tissue detected a pathogenic germline mutation of the FAM175A gene and somatic mutations in BRCA2 and RAD51B (in both sarcoma and lymphoma specimens), and INPP4B and RICTOR (in lymphoma specimen only).

HIV infection is associated with an increased risk for developing B-cell lymphoproliferative disorders. The spectrum of disease differs in HIV-infected versus HIV-uninfected persons, with aggressive B-cell non-Hodgkin lymphomas constituting a higher proportion of all lymphoproliferative disorders in the HIV-positive population. Although antiretroviral therapy (ART) has significantly changed the landscape of lymphomas arising in HIV-infected persons, population growth and aging are reflected in the steady increase in non-AIDS-defining cancers. In the ART era, outcomes for HIV-infected lymphoma patients are similar to those of HIV-negative patients. This article reviews the diagnostic features and summarizes current biologic understanding of HIV-associated lymphomas., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Genome-wide CRISPR/Cas9 screens represent a powerful approach to studying mechanisms of drug action and resistance. Cereblon modulating agents (CMs) have recently emerged as candidates for therapeutic intervention in primary effusion lymphoma (PEL), a highly aggressive cancer caused by Kaposi's sarcoma-associated herpesvirus. CMs bind to cereblon (CRBN), the substrate receptor of the cullin-RING type E3 ubiquitin ligase CRL4 CRBN , and thereby trigger the acquisition and proteasomal degradation of neosubstrates. Downstream mechanisms of CM toxicity are incompletely understood, however. To identify novel CM effectors and mechanisms of CM resistance, we performed positive selection CRISPR screens using 3 CMs with increasing toxicity in PEL: lenalidomide (LEN), pomalidomide (POM), and CC-122. Results identified several novel modulators of the activity of CRL4 CRBN The number of genes whose inactivation confers resistance decreases with increasing CM efficacy. Only inactivation of CRBN conferred complete resistance to CC-122. Inactivation of the E2 ubiquitin conjugating enzyme UBE2G1 also conferred robust resistance against LEN and POM. Inactivation of additional genes, including the Nedd8-specific protease SENP8, conferred resistance to only LEN. SENP8 inactivation indirectly increased levels of unneddylated CUL4A/B, which limits CRL4 CRBN activity in a dominant negative manner. Accordingly, sensitivity of SENP8-inactivated cells to LEN is restored by overexpression of CRBN. In sum, our screens identify several novel players in CRL4 CRBN function and define pathways to CM resistance in PEL. These results provide rationale for increasing CM efficacy on patient relapse from a less-efficient CM. Identified genes could finally be developed as biomarkers to predict CM efficacy in PEL and other cancers., (© 2019 by The American Society of Hematology.)

Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma and is associated with two B cell malignancies, primary effusion lymphoma (PEL) and the plasmablastic variant of multicentric Castleman's disease. On several adherent cell types, EphA2 functions as a cellular receptor for the gH/gL glycoprotein complex of KSHV. KSHV gH/gL also has previously been found to interact weakly with other members of the Eph family of receptor tyrosine kinases (Ephs), and other A-type Ephs have been shown to be able to compensate for the absence of EphA2 using overexpression systems. However, whether these interactions are of functional consequence at endogenous protein levels has remained unclear so far. Here, we demonstrate for the first time that endogenously expressed EphA7 in BJAB B cells is critical for the cell-to-cell transmission of KSHV from producer iSLK cells to BJAB target cells. The BJAB lymphoblastoid cell line often serves as a model for B cell infection and expresses only low levels of all Eph family receptors other than EphA7. Endogenous EphA7 could be precipitated from the cellular lysate of BJAB cells using recombinant gH/gL, and knockout of EphA7 significantly reduced transmission of KSHV into BJAB target cells. Knockout of EphA5, the second most expressed A-type Eph in BJAB cells, had a similar, although less pronounced, effect on KSHV infection. Receptor function of EphA7 was conserved for cell-free infection by the related rhesus monkey rhadinovirus (RRV), which is relatively even more dependent on EphA7 for infection of BJAB cells. IMPORTANCE Infection of B cells is relevant for two KSHV-associated malignancies, the plasmablastic variant of multicentric Castleman's disease and PEL. Therefore, elucidating the process of B cell infection is important for the understanding of KSHV pathogenesis. While the high-affinity receptor for the gH/gL glycoprotein complex, EphA2, has been shown to function as an entry receptor for various types of adherent cells, the gH/gL complex can also interact with other Eph receptor tyrosine kinases with lower avidity. We analyzed the Eph interactions required for infection of BJAB cells, a model for B cell infection by KSHV. We identified EphA7 as the principal Eph receptor for infection of BJAB cells by KSHV and the related rhesus monkey rhadinovirus. While two analyzed PEL cell lines exhibited high EphA2 and low EphA7 expression, a third PEL cell line, BCBL-1, showed high EphA7 and low EphA2 expression, indicating a possible relevance for KSHV pathology., (Copyright © 2019 American Society for Microbiology.)

Primary effusion lymphoma (PEL) is an aggressive HIV-associated lymphoma with a relatively poor prognosis in the era of effective HIV therapy. Kaposi sarcoma herpesvirus (KSHV) is the etiologic agent, and ∼80% of tumors are coinfected with Epstein-Barr virus (EBV). A better understanding of how KSHV-related immune dysregulation contributes to the natural history of PEL will improve outcomes. Twenty patients with PEL diagnosed between 2000 and 2013, including 19 treated with modified infusional etoposide, vincristine, and doxorubicin with cyclophosphamide and prednisone (EPOCH), were identified. We compared their clinical, virologic, and immunologic features vs 20 patients with HIV-associated diffuse large B-cell lymphoma and 19 patients with symptomatic interleukin (IL)-6 related KSHV-associated multicentric Castleman disease. Survival analyses of treated patients with PEL were then performed to identify prognostic factors and cancer-specific mortality. Compared with HIV-associated diffuse large B-cell lymphoma, PEL was associated with significant hypoalbuminemia ( P < .0027), thrombocytopenia ( P = .0045), and elevated IL-10 levels ( P < .0001). There were no significant differences in these parameters between PEL and KSHV-associated multicentric Castleman disease. Median overall survival in treated patients with PEL was 22 months, with a plateau in survival noted after 2 years. Three-year cancer-specific survival was 47%. EBV-positive tumor status was associated with improved survival (hazard ratio, 0.27; P = .038), and elevated IL-6 level was associated with inferior survival (hazard ratio, 6.1; P = .024). Our analysis shows that IL-6 and IL-10 levels contribute to the natural history of PEL. Inflammatory cytokines and tumor EBV status are the strongest prognostic factors. Pathogenesis-directed first-line regimens are needed to improve overall survival in PEL., (© 2019 by The American Society of Hematology.)

Primary effusion lymphoma (PEL) is caused by Kaposi's sarcoma-associated herpesvirus (KSHV). PEL has a highly active mTOR pathway, which makes mTOR a potential therapeutic target. MLN0128 is an ATP-competitive inhibitor of mTOR that has entered clinical trials for solid tumors. Our results demonstrated that MLN0128 has a greater effect on inhibiting proliferation than the allosteric mTOR inhibitor rapamycin. MLN0128 has ∼30 nM 50% inhibitory concentration (IC 50 ) across several PEL cell lines, including PEL that is resistant to conventional chemotherapy. MLN0128 induced apoptosis in PEL, whereas rapamycin induced G 1 arrest, consistent with a different mechanism of action. MLN0128 inhibited phosphorylation of mTOR complex 1 and 2 targets, while rapamycin only partially inhibited mTOR complex 1 targets. PEL xenograft mouse models treated with MLN0128 showed reduced effusion volumes in comparison to the vehicle-treated group. Rapamycin-resistant (RR) clones with an IC 50 for rapamycin 10 times higher than the parental IC 50 emerged consistently after rapamycin exposure as a result of transcriptional adaptation. MLN0128 was nevertheless capable of inducing apoptosis in these RR clones. Our results suggest that MLN0128 might offer a new approach to the treatment of chemotherapy-resistant PEL. IMPORTANCE Primary effusion lymphoma (PEL) is an aggressive and incurable malignancy, which is usually characterized by lymphomatous effusions in body cavities without tumor masses. PEL has no established treatment and a poor prognosis, with a median survival time shorter than 6 months. PEL usually develops in the context of immunosuppression, such as HIV infection or post-organ transplantation. The optimal treatment for PEL has not been established, as PEL is generally resistant to traditional chemotherapy. The molecular drivers for PEL are still unknown; however, PEL displays a constitutively active mammalian target of rapamycin (mTOR) pathway, which is critical for metabolic and cell survival mechanisms. Therefore, the evaluation of novel agents targeting the mTOR pathway could be clinically relevant for the treatment of PEL., (Copyright © 2019 Caro-Vegas et al.)

Primary effusion lymphoma (PEL) is a rare mature B-cell non-Hodgkin's lymphoma arising in body cavities and presenting with effusions. It has been described predominantly in patients with impaired immunity from the acquired immunodeficiency syndrome and is associated with the Human Herpesvirus-8 (HHV-8). Seldom has PEL been diagnosed in persons negative for the human immunodeficiency virus (HIV), and in such cases it has occurred primarily in the setting of posttransplant immunosuppression. We report an instructive case of a Caribbean-American HIV-negative orthotopic heart transplant recipient with a history of HHV-8-associated Kaposi's sarcoma who developed HHV-8 viremia and PEL of the pleural space early in the posttransplant course. This case highlights the importance of considering PEL in the differential diagnosis of a new pleural effusion in a transplant recipient at risk for HHV-8-associated disease., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Primary effusion lymphoma (PEL) is a rare B-cell malignancy that most often occurs in immunocompromised patients, such as HIV-infected individuals and patients receiving organ transplantation. The main characteristic of PEL is neoplastic effusions in body cavities without detectable tumor masses. The onset of the disease is associated with latent infection of human herpes virus 8/Kaposi sarcoma-associated herpes virus, and the normal counterpart of tumor cells is B cells with plasmablastic differentiation. A condition of immunodeficiency and a usual absence of CD20 expression lead to the expectation of the lack of efficacy of anti-CD20 monoclonal antibody; clinical outcomes of the disease remain extremely poor, with an overall survival at 1 year of ∼30%. Although recent progress in antiretroviral therapy has improved outcomes of HIV-infected patients, its benefit is still limited in patients with PEL. Furthermore, the usual high expression of programmed death ligand 1 in tumor cells, one of the most important immune-checkpoint molecules, results in the immune escape of tumor cells from the host immune defense, which could be the underlying mechanism of poor treatment efficacy. Molecular-targeted therapies for the activating pathways in PEL, including NF-κB, JAK/STAT, and phosphatidylinositol 3-kinase/AKT, have emerged to treat this intractable disease. A combination of immunological recovery from immune deficiency, overcoming the immune escape, and the development of more effective drugs will be vital for improving the outcomes of PEL patients in the future., (© 2018 by The American Society of Hematology.)

Primary effusion lymphoma (PEL) is a rare type of extranodal lymphoma, typically of a B-cell origin, which presents as lymphomatous effusion with no nodal enlargement or tumor masses. The development PEL is universally associated with human herpes virus-8 (HHV-8) infection. Cases of HHV-8-negative primary lymphomatous effusion have recently been reported and referred to as HHV-8-unrelated PEL-like lymphoma. Some cases of this disease have been reported in iatrogenic immunocompromised patients. The mechanisms responsible for the inhibitory effects of the discontinuation of immunosuppressants other than methotrexate (MTX) against the disease, which have been demonstrated for MTX-associated lymphoproliferative disorders, have not yet been elucidated. We describe a case of PEL-like lymphoma that developed in the course of antisynthetase syndrome and was treated with tacrolimus. A single dose of systemic chemotherapy did not improve lymphomatous effusion, whereas the discontinuation of tacrolimus resulted in the long-term remission of this disease.

Effusion-based lymphoma (EBL) is a rare but distinct entity of large B-cell lymphoma in effusion without association with human herpes virus-8 (HHV-8). Spontaneous regression after pleurocentesis has been observed; but to our knowledge, there are no reports on the morphological and molecular features of subsequent aspirations in regressing cases. Here, we report the case of a 92-year-old male with chronic obstructive pulmonary disease, who presented with right pleural effusion. He had no human immunodeficiency virus, hepatitis B virus, or hepatitis C virus infection, and CT scans revealed no mass lesion. The first pleural effusion aspiration cytology revealed large lymphoma cells with vesicular nuclei, irregular nuclear contours, and prominent nucleoli, consistent with EBL. The second aspiration cytology showed a few slightly enlarged lymphocytes in a background of small lymphocytes. Immunohistochemical study on cell block of the second aspiration revealed equal amounts of CD3-positive and CD20-positive cells. All these cells on the section tested negative for HHV-8 through immunohistochemistry and Epstein-Barr virus by in situ hybridization. Our initial impression was EBL in regression. However, flow cytometric immunophenotyping showed monotypic light chain expression of the gated B-cells. B-cell receptor gene rearrangement study showed a clonal result. Furthermore, fluorescence in situ hybridization revealed rearrangement of IGH gene. The diagnosis of the second aspiration was EBL with morphological regression but retained clonal genetic features. The patient passed away one month after diagnosis without chemotherapy. This case illustrated the importance of ancillary studies in confirming the clonal nature of a morphologically regressing EBL., (© 2018 Wiley Periodicals, Inc.)

Kaposi's sarcoma-associated herpesvirus (KSHV, HHV-8) is a gammaherpesvirus associated with several human malignancies. DNA methylation at CpG dinucleotides is an epigenetic mark dysregulated in many cancer types and in KSHV-infected cells. Several previous studies have analyzed in detail the CpG methylation of the KSHV episomal genomes, but little is known about the impact of KSHV on the human genome. Our knowledge of cellular CpG methylation in the context of KSHV infection is currently limited to four hypermethylated human gene promoters. Therefore, we undertook a comprehensive CpG methylation analysis of the human methylome in KSHV-infected cells and KSHV-associated primary effusion lymphoma (PEL). We performed Infinium HumanMethylation450K and MethylationEpic BeadChip arrays and identified panels of hyper- and hypomethylated cellular promoters in KSHV-infected cells. We combined our genome-wide methylation analysis with high-throughput RNA sequencing (RNA-seq) to add functional outcomes to the virally induced methylation changes. We were able to correlate many downregulated genes with promoter hypermethylation and upregulated genes with hypomethylation. In addition, we show that treating the cells with a demethylating agent leads to reexpression of these downregulated genes, indicating that, indeed, DNA methylation plays a role in the repression of these human genes. Comparison between de novo infection and PEL suggests that the virus induces initial hypermethylation followed by a slow increase in genome-wide hypomethylation. This study extends our understanding of the relationship between epigenetic changes induced by KSHV infection and tumorigenesis. IMPORTANCE In cancer cells, certain promoters become aberrantly methylated, contributing to the phenotype of the tumor. KSHV infection seems to modify cellular CpG methylation, but only a few methylated promoters have been identified in KSHV-infected cells. Here, we investigated the CpG methylation of the human genome in KSHV-associated primary effusion lymphoma (PEL) and KSHV-infected cells. We have identified many hyper- and hypomethylated gene promoters and correlated their methylation with cellular gene expression. These differentially methylated cellular promoters can distinguish KSHV-positive cells from uninfected cells and may serve as the foundation for the use of these differentially methylated regions as potential biomarkers for KSHV-associated malignancies. Drugs that reverse these cancerous methylation patterns have the potential to inhibit tumor growth. Here, we show that treating PEL cells with a demethylating drug (5-aza-2'-deoxycytidine) led to inhibition of cell growth, raising the possibility of testing this drug for the treatment of PEL., (Copyright © 2018 American Society for Microbiology.)