Arsenic trioxide inhibits EBV reactivation and promotes cell death in EBV-positive lymphoma cells

Abstract

BackgroundEpstein-Barr Virus (EBV) is associated with hematopoietic malignancies, such as Burkitt’s lymphoma, post-transplantation lymphoproliferative disorder, and diffuse large B-cell lymphoma. The current approach for EBV-associated lymphoma involves chemotherapy to eradicate cancer cells, however, normal cells may be injured and organ dysfunction may occur with currently employed regimens. This research is focused on employing arsenic trioxide (ATO) as EBV-specific cancer therapy takes advantage of the fact the EBV resides within the malignant cells.Methods and resultsOur research reveals that low ATO inhibits EBV gene expression and genome replication. EBV spontaneous reactivation starts as early as 6 h after re-suspending EBV-positive Mutu cells in RPMI media in the absence of ATO, however this does not occur in Mutu cells cultured with ATO. ATO’s inhibition of EBV spontaneous reactivation is dose dependent. The expression of the EBV immediate early gene Zta and early gene BMRF1 is blocked with low concentrations of ATO (0.5 nM – 2 nM) in EBV latency type I cells and EBV-infected PBMC cells. The combination of ATO and ganciclovir further diminishes EBV gene expression. ATO-mediated reduction of EBV gene expression can be rescued by co-treatment with the proteasome inhibitor MG132, indicating that ATO promotes ubiquitin conjugation and proteasomal degradation of EBV genes. Co-immunoprecipitation assays with antibodies against Zta pulls down more ubiquitin in ATO treated cell lysates. Furthermore, MG132 reverses the inhibitory effect of ATO on anti-IgM-, PMA- and TGF-β-mediated EBV reactivation. Thus, mechanistically ATO’s inhibition of EBV gene expression occurs via the ubiquitin pathway. Moreover, ATO treatment results in increased cell death in EBV-positive cells compared to EBV-negative cells, as demonstrated by both MTT and trypan blue assays. ATO-induced cell death in EBV-positive cells is dose dependent. ATO and ganciclovir in combination further enhances cell death specifically in EBV-positive cells.ConclusionATO-mediated inhibition of EBV lytic gene expression results in cell death selectively in EBV-positive lymphocytes, suggesting that ATO may potentially serve as a drug to treat EBV-related lymphomas in the clinical setting.

Toxic metal exposure as a possible risk factor for COVID-19 and other respiratory infectious diseases

“…Particularly, As 2 O 3 exposure significantly inhibits growth of human T-cell leukemia virus-infected T-cell lines through induction of apoptosis and cell cycle arrest ( Ishitsuka et al, 2000 ). Similarly, As administration was also shown to inhibit Epstein-Barr Virus replication and stimulates cell death in EBV-positive cells ( Yin et al, 2017 ). Due to the observed effects of As exposure on viral agents of leukemia, As and IFN-α is widely used as an antileukemic agent ( Hachiman et al, 2018 ).…”

Arsenicals, the Integrated Stress Response, and Epstein–Barr Virus Lytic Gene Expression

“…Previous investigators have studied the impact of arsenicals on EBV gene expression [ 7 , 8 , 9 , 28 ]. Sides et al reported activation of Zta and BMRF1 RNA expression in epithelial cells [ 8 ].…”

“…Yin et al studied EBV-positive lymphoma cell lines treated with As 2 O 3 at nanomolar concentrations for three days or longer. They reported decreased cell viability and reduced expression of Zta and BMRF1 [ 9 ]. Our results parallel those in the previous reports in some regards but differ in others, possibly reflecting differences in particular cell lines studied or in the duration of drug treatment.…”

“…Arsenic trioxide (As 2 O 3 ) has been reported to activate EBV lytic gene expression in epithelial cells [ 7 , 8 ] but not in Burkitt lymphoma cells [ 9 ]. As 2 O 3 is used in the clinic in the treatment of promyelocytic leukemia [ 10 ].…”

New developments in the pathology of malignant lymphoma: a review of the literature published from May to August 2017

“…They conclude that their results provide evidence for the impact of microenvironment on EBV-carrying DLBCL cells and might have therapeutic implications. Yin et al [ 14 ] are also able to promote cell death in EBV-positive cells in vitro, by using arsenic trioxide (ATO). EBV spontaneous reactivation starts as early as 6 h after re-suspending EBV-positive Mutu cells in RPMI media in the absence of ATO; however, this does not occur in Mutu cells cultured with ATO.…”