Epstein-Barr virus (EBV) and Kaposi's sarcoma herpesvirus (KSHV), formally designated human herpesvirus 4 (HHV-4) and 8 (HHV-8), respectively, are viruses that can cause a variety of cancers in ...humans. EBV is found in non-Hodgkin and Hodgkin lymphomas, as well as in lymphoproliferative disorders, which occur more commonly but not exclusively in individuals with immunodeficiency. EBV also causes nonlymphoid malignancies such as nasopharyngeal carcinoma. KSHV causes primary effusion lymphomas, multicentric Castleman's disease, and Kaposi's sarcoma. The frequency of lymphoid malignancies related to infection by one of these two herpesviruses is greatly increased in individuals with immunodeficiency, whether primary or acquired, for example, as a consequence of HIV infection and AIDS or in the case of therapeutic immunosuppression for organ transplantation. Our current understanding indicates that EBV and KSHV contribute to lymphomagenesis by affecting genomic stability and by subverting the cellular molecular signaling machinery and metabolism to avoid immune surveillance and enhance tumor cell growth and survival. Understanding the viral associations in specific lymphoproliferative disorders and the molecular mechanisms of viral oncogenesis will lead to better prevention, diagnosis, and treatment strategies for these diseases.
Kaposi sarcoma (KS) gained public attention as an AIDS-defining malignancy; its appearance on the skin was a highly stigmatizing sign of HIV infection during the height of the AIDS epidemic. The ...widespread introduction of effective antiretrovirals to control HIV by restoring immunocompetence reduced the prevalence of AIDS-related KS, although KS does occur in individuals with well-controlled HIV infection. KS also presents in individuals without HIV infection in older men (classic KS), in sub-Saharan Africa (endemic KS) and in transplant recipients (iatrogenic KS). The aetiologic agent of KS is KS herpesvirus (KSHV; also known as human herpesvirus-8), and viral proteins can induce KS-associated cellular changes that enable the virus to evade the host immune system and allow the infected cell to survive and proliferate despite viral infection. Currently, most cases of KS occur in sub-Saharan Africa, where KSHV infection is prevalent owing to transmission by saliva in childhood compounded by the ongoing AIDS epidemic. Treatment for early AIDS-related KS in previously untreated patients should start with the control of HIV with antiretrovirals, which frequently results in KS regression. In advanced-stage KS, chemotherapy with pegylated liposomal doxorubicin or paclitaxel is the most common treatment, although it is seldom curative. In sub-Saharan Africa, KS continues to have a poor prognosis. Newer treatments for KS based on the mechanisms of its pathogenesis are being explored.
•KSHV/HHV8 causes three human diseases: Kaposi Sarcoma, Primary Effusion Lymphoma and the plasma cell variant of Multicentric Castleman Disease.•These occur more commonly in patients with ...immunodeficiency, including AIDS.•KS is heterogeneous, and may represent a spectrum between an angioproliferative disease and a true malignancy.•KSHV encodes viral genes that can cause inflammation, angiogenesis and neoplastic transformation.
Kaposi Sarcoma-associated Herpesvirus (KSHV, HHV8) causes three human malignancies, Kaposi Sarcoma (KS), an endothelial tumor, as well as Primary Effusion Lymphoma (PEL) and the plasma cell variant of Multicentric Castleman's Disease (MCD), two B-cell lymphoproliferative diseases. All three cancers occur primarily in the context of immune deficiency and/or HIV infection, but their pathogenesis differs. KS most likely results from the combined effects of an endotheliotropic virus with angiogenic properties and inflammatory stimuli and thus represents an interesting example of a cancer that arises in an inflammatory context. Viral and cellular angiogenic and inflammatory factors also play an important role in the pathogenesis of MCD. In contrast, PEL represents an autonomously growing malignancy that is, however, still dependent on the continuous presence of KSHV and the action of several KSHV proteins.
Abstract Kaposi’s sarcoma (KS) is a cancer affecting skin and internal organs for which the Kaposi’s sarcoma associated herpesvirus (KSHV) is a necessary cause. Previous work has pursued KS diagnosis ...by quantifying KSHV DNA in skin biopsies using a point-of-care (POC) device which performs quantitative loop-mediated isothermal amplification (LAMP). These previous studies revealed that extracting DNA from patient biopsies was the rate limiting step in an otherwise rapid process. In this study, a simplified, POC-compatible alkaline DNA extraction, ColdSHOT, was optimized for 0.75 mm human skin punch biopsies. The optimized ColdSHOT extraction consistently produced 40,000+ copies of DNA per 5 µl reaction from 3 mg samples—a yield comparable to standard spin column extractions—within 1 h without significant equipment. The DNA yield was estimated sufficient for KSHV detection from KS-positive patient biopsies, and the LAMP assay was not affected by non-target tissue in the unpurified samples. Furthermore, the yields achieved via ColdSHOT were robust to sample storage in phosphate-buffered saline (PBS) or Tris-EDTA (TE) buffer prior to DNA extraction, and the DNA sample was stable after extraction. The results presented in this study indicate that the ColdSHOT DNA extraction could be implemented to simplify and accelerate the LAMP-based diagnosis of Kaposi’s sarcoma using submillimeter biopsy samples.
Protein-protein interactions featuring intricate binding epitopes remain challenging targets for synthetic inhibitors. Interactions of NEMO, a scaffolding protein central to NF-κB signaling, ...exemplify this challenge. Various regulators are known to interact with different coiled coil regions of NEMO, but the topological complexity of this protein has limited inhibitor design. We undertook a comprehensive effort to block the interaction between vFLIP, a Kaposi's sarcoma herpesviral oncoprotein, and NEMO using small molecule screening and rational design. Our efforts reveal that a tertiary protein structure mimic of NEMO is necessary for potent inhibition. The rationally designed mimic engages vFLIP directly causing complex disruption, protein degradation and suppression of NF-κB signaling in primary effusion lymphoma (PEL). NEMO mimic treatment induces cell death and delays tumor growth in a PEL xenograft model. Our studies with this inhibitor reveal the critical nexus of signaling complex stability in the regulation of NF-κB by a viral oncoprotein.
Classical Hodgkin lymphoma (cHL) is characterized by sparsely distributed Hodgkin and Reed-Sternberg (HRS) cells amid reactive host background, complicating the acquisition of neoplastic DNA without ...extensive background contamination. We overcame this limitation by using flow-sorted HRS and intratumor T cells and optimized low-input exome sequencing of 10 patient samples to reveal alterations in genes involved in antigen presentation, chromosome integrity, transcriptional regulation, and ubiquitination. β-2-microglobulin (B2M) is the most commonly altered gene in HRS cells, with 7 of 10 cases having inactivating mutations that lead to loss of major histocompatibility complex class I (MHC-I) expression. Enforced wild-type B2M expression in a cHL cell line restored MHC-I expression. In an extended cohort of 145 patients, the absence of B2M protein in the HRS cells was associated with lower stage of disease, younger age at diagnosis, and better overall and progression-free survival. B2M-deficient cases encompassed most of the nodular sclerosis subtype cases and only a minority of mixed cellularity cases, suggesting that B2M deficiency determines the tumor microenvironment and may define a major subset of cHL that has more uniform clinical and morphologic features. In addition, we report previously unknown genetic alterations that may render selected patients sensitive to specific targeted therapies.
•We show feasibility of whole-exome sequencing on purified primary HRS cells and report recurrent genetic alterations characterizing cHL.•B2M is the most frequently mutated gene in cHL, strongly associated with nodular sclerosis subtype, younger age, and better overall survival.
Hsp90 is a molecular chaperone that protects proteins, including oncogenic signaling complexes, from proteolytic degradation. PU-H71 is a next-generation Hsp90 inhibitor that preferentially targets ...the functionally distinct pool of Hsp90 present in tumor cells. Tumors that are driven by the MYC oncoprotein may be particularly sensitive to PU-H71 due to the essential role of Hsp90 in the epichaperome, which maintains the malignant phenotype in the setting of MYC. Burkitt lymphoma (BL) is an aggressive B-cell lymphoma characterized by MYC dysregulation. In this study, we evaluated Hsp90 as a potential therapeutic target in BL. We found that primary BL tumors overexpress Hsp90 and that Hsp90 inhibition has antitumor activity
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, including potent activity in a patient-derived xenograft model of BL. To evaluate the targets of PU-H71 in BL, we performed high-affinity capture followed by proteomic analysis using mass spectrometry. We found that Hsp90 inhibition targets multiple components of PI3K/AKT/mTOR signaling, highlighting the importance of this pathway in BL. Finally, we found that the anti-lymphoma activity of PU-H71 is synergistic with dual PI3K/mTOR inhibition
and
Overall, this work provides support for Hsp90 as a therapeutic target in BL and suggests the potential for combination therapy with PU-H71 and inhibitors of PI3K/mTOR.
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H1 linker histones are the most abundant chromatin-binding proteins
. In vitro studies indicate that their association with chromatin determines nucleosome spacing and enables arrays of nucleosomes ...to fold into more compact chromatin structures. However, the in vivo roles of H1 are poorly understood
. Here we show that the local density of H1 controls the balance of repressive and active chromatin domains by promoting genomic compaction. We generated a conditional triple-H1-knockout mouse strain and depleted H1 in haematopoietic cells. H1 depletion in T cells leads to de-repression of T cell activation genes, a process that mimics normal T cell activation. Comparison of chromatin structure in normal and H1-depleted CD8
T cells reveals that H1-mediated chromatin compaction occurs primarily in regions of the genome containing higher than average levels of H1: the chromosome conformation capture (Hi-C) B compartment and regions of the Hi-C A compartment marked by PRC2. Reduction of H1 stoichiometry leads to decreased H3K27 methylation, increased H3K36 methylation, B-to-A-compartment shifting and an increase in interaction frequency between compartments. In vitro, H1 promotes PRC2-mediated H3K27 methylation and inhibits NSD2-mediated H3K36 methylation. Mechanistically, H1 mediates these opposite effects by promoting physical compaction of the chromatin substrate. Our results establish H1 as a critical regulator of gene silencing through localized control of chromatin compaction, 3D genome organization and the epigenetic landscape.
KSHV is the causative agent of Kaposi sarcoma (KS), a spindle-shaped endothelial cell neoplasm accompanied by an inflammatory infiltrate. To evaluate the role of KSHV vFLIP in the pathogenesis of KS, ...we constructed mice with inducible expression of vFLIP in endothelial cells. Abnormal cells with endothelial marker expression and fusiform appearance were observed in several tissues reminiscent of the spindle cells found in KS. Serum cytokines displayed a profound perturbation similar to that described in KSHV inflammatory cytokine syndrome (KICS), a recently described clinical condition characterized by elevated IL6 and IL10. An increased myeloid component with suppressive immune phenotype was found, which may contribute to functional changes in the microenvironment and cellular heterogeneity as observed in KS. These mice represent the first in vivo demonstration that vFLIP is capable of inducing vascular abnormalities and changes in host microenvironment with important implications for understanding the pathogenesis and treating KSHV-associated diseases.
Epstein-Barr virus (EBV) persistently infects 95% of adults worldwide and is associated with multiple human lymphomas that express characteristic EBV latency programs used by the virus to navigate ...the B-cell compartment. Upon primary infection, the EBV latency III program, comprised of six Epstein-Barr Nuclear Antigens (EBNA) and two Latent Membrane Protein (LMP) antigens, drives infected B-cells into germinal center (GC). By incompletely understood mechanisms, GC microenvironmental cues trigger the EBV genome to switch to the latency II program, comprised of EBNA1, LMP1 and LMP2A and observed in GC-derived Hodgkin lymphoma. To gain insights into pathways and epigenetic mechanisms that control EBV latency reprogramming as EBV-infected B-cells encounter microenvironmental cues, we characterized GC cytokine effects on EBV latency protein expression and on the EBV epigenome. We confirmed and extended prior studies highlighting GC cytokine effects in support of the latency II transition. The T-follicular helper cytokine interleukin 21 (IL-21), which is a major regulator of GC responses, and to a lesser extent IL-4 and IL-10, hyper-induced LMP1 expression, while repressing EBNA expression. However, follicular dendritic cell cytokines including IL-15 and IL-27 downmodulate EBNA but not LMP1 expression. CRISPR editing highlighted that STAT3 and STAT5 were necessary for cytokine mediated EBNA silencing via epigenetic effects at the EBV genomic C promoter. By contrast, STAT3 was instead necessary for LMP1 promoter epigenetic remodeling, including gain of activating histone chromatin marks and loss of repressive polycomb repressive complex silencing marks. Thus, EBV has evolved to coopt STAT signaling to oppositely regulate the epigenetic status of key viral genomic promoters in response to GC cytokine cues.
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