Maribavir Inhibits Epstein-Barr Virus Transcription in Addition to Viral DNA Replication

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A lthough a number of antiviral drugs are effective inhibitors of Epstein-Barr virus (EBV) replication and are used empirically, none is of proven effectiveness for treatment of EBV infection (1, 2). Maribavir (MBV), which is in late-stage clinical trials for use against human cytomegalovirus (HCMV) infection in allogeneic stem cell and bone-marrow transplant recipients (3,4), is of special interest because it is also a potent inhibitor of EBV replication (5-7). In stem-cell and organ transplant recipients, EBV infection poses the hazard of generating B-cell lymphomas that are ultimately fatal. While no drugs are currently approved for treatment of EBV disease, several that inhibit EBV are available, and these can be divided into two main classes: those that target the viral DNA polymerase and those that function independently of it (8-12). Acyclic nucleoside and phosphonated nucleotide analogs, as well as pyrophosphate analogs, all target the viral polymerase.A new class of HCMV inhibitors, benzimidazole compounds, with more specific antiviral properties and fewer adverse side effects, blocked HCMV DNA maturation and encapsidation processes and led to the design of 1-H-␤-L-ribofuranoside-2-isopropylamino-5,6-dichlorobenzimidazole (maribavir [MBV]) (13)(14)(15)(16)(17)(18)(19)(20). Unlike its parent compound, which inhibits HCMV replication but not EBV replication, MBV inhibits both (7,21). Inhibitory effects of MBV are produced mainly through inhibition of the HCMV and EBV protein kinases (PK) (21-24). Previous phase 3 studies with a dosage of 100 mg twice a day (BID) did not have sufficient activity to prevent HCMV disease, but the safety profile and data from case studies suggested that higher doses would be clinically active (3). MBV is now in new phase 2 trials at doses of 400, 800, and 1,200 mg BID (3).Maribavir selectively inhibits the HCMV protein kinase, UL97, determined by direct inhibition of kinase activity in vitro and by genetic mapping of the MBV-resistant phenotype (21). MBV also inhibits the EBV protein kinase (BGLF4), resulting in inhibition of phosphorylation of the EBV DNA processivity factor BMRF1, but does not seem to act directly on the EBV kinase in vitro (7,24).We have recently found that MBV also inhibits expression of multiple EBV transcripts, in contrast to acyclovir (ACV), which has little effect on EBV RNAs. Thus, MBV has a unique dual effect on viral DNA transcription as well as replication (25). In this study, we find that the inhibitory profile of MBV transcripts is similar to that produced by mutant EBV in which PK expression and activity have been knocked out (26). Thus, the results suggest that MBV largely affects EBV transcript levels through inhibition of BGLF4.To determine if the profile of viral transcripts produced by MBV is mediated by the viral kinase, we utilized BGLF4 knockout (KO) (dBGLF4/NeoST) and revertant (dBGLF4/NeoSt/R) viruses constructed and characterized by Murata et al. (26). 293 cells maintaining wild-type (WT), BGLF4 knockout, and revertant EBV genomes (2...

supporting

confidence: 55%

supporting

confidence: 50%

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Maribavir Inhibits Epstein-Barr Virus Transcription through the EBV Protein Kinase

Whitehurst

Sanders

Law

et al. 2013

Self Cite

“…Although genome-wide analyses agree that the expression of late lytic genes lags behind that of early lytic cycle genes (45), it has also been shown that the requirement for EBV DNA replication for their expression is not universal. While the promoters for some late genes are dependent on EBV genome replication for their expression (1), several late lytic cycle genes can be activated in the absence of viral genome synthesis (9,30) and some are expressed in the presence of acyclovir in Akata cells undergoing lytic cycle (42). Furthermore, the viral BcRF1 gene product acts as a TATA binding protein for some late genes (13), which may affect the timing of their expression.…”

Section: Discussionmentioning

confidence: 99%

Genome-Wide Analyses of Zta Binding to the Epstein-Barr Virus Genome Reveals Interactions in both Early and Late Lytic Cycles and an Epigenetic Switch Leading to an Altered Binding Profile

Ramasubramanyan

Kanhere

Osborn

et al. 2012

cThe Epstein-Barr virus (EBV) genome sustains substantial epigenetic modification involving chromatin remodelling and DNA methylation during lytic replication. Zta (ZEBRA, BZLF1), a key regulator of the EBV lytic cycle, is a transcription and replication factor, binding to Zta response elements (ZREs) in target promoters and EBV lytic origins of replication. In vitro, Zta binding is modulated by DNA methylation; a subset of CpG-containing Zta binding sites (CpG ZREs) is bound only in a DNA methylation-dependent manner. The question of how the dynamic epigenetic environment impacts Zta interaction during the EBV lytic cycle is unknown. To address this, we used chromatin immunoprecipitation coupled with next-generation sequencing (ChIP-Seq) to identify Zta binding sites across the EBV genome before and after viral DNA replication. Replication did not alter the association of Zta across many regions of the EBV genome, but a striking reduction in Zta binding occurred at some loci that contain CpG ZREs. Separating Zta-bound DNA into methylated and nonmethylated fractions, we found that promoters that contain CpG ZREs were enriched in the methylated fraction but that Zta binding to promoters lacking CpG ZREs was not reduced. We hypothesize that the loss of DNA methylation on the EBV genome during the lytic cycle causes the reduced binding to CpG ZREs; this may act as a lytic cycle epigenetic switch. However, the epigenetic changes associated with the replicated EBV genome do not affect the interaction of Zta with many loci that are rich in non-CpG ZREs; this leads to sustained binding at these regions.

“…In this study, we show that Pin1 interacts with EBV DNA polymerase BALF5 and modulates productive viral DNA replica-tion. Because there is a very limited number of anti-EBV drugs developed or being developed to date, including acyclic nucleoside analogs, such as acyclovir, and kinase inhibitors, such as maribavir (26,27), a search for an effective molecular target has been needed. Pin1 may be a potential target for development of novel antiviral drugs.…”

Section: T He Epstein-barr Virus (Ebv) Is a Human Gammaherpesvirus Thmentioning

confidence: 99%

Pin1 Interacts with the Epstein-Barr Virus DNA Polymerase Catalytic Subunit and Regulates Viral DNA Replication

Narita

Murata

Ryo

et al. 2013

Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1) protein is known as a regulator which recognizes phosphorylated Ser/Thr-Pro motifs and increases the rate of cis and trans amide isomer interconversion, thereby altering the conformation of its substrates. We found that Pin1 knockdown using short hairpin RNA (shRNA) technology resulted in strong suppression of productive Epstein-Barr virus (EBV) DNA replication. We further identified the EBV DNA polymerase catalytic subunit, BALF5, as a Pin1 substrate in glutathione S-transferase (GST) pulldown and immunoprecipitation assays. Lambda protein phosphatase treatment abolished the binding of BALF5 to Pin1, and mutation analysis of BALF5 revealed that replacement of the Thr178 residue by Ala (BALF5 T178A) disrupted the interaction with Pin1. To further test the effects of Pin1 in the context of virus infection, we constructed a BALF5-deficient recombinant virus. Exogenous supply of wild-type BALF5 in HEK293 cells with knockout recombinant EBV allowed efficient synthesis of viral genome DNA, but BALF5 T178A could not provide support as efficiently as wild-type BALF5. In conclusion, we found that EBV DNA polymerase BALF5 subunit interacts with Pin1 through BALF5 Thr178 in a phosphorylation-dependent manner. Pin1 might modulate EBV DNA polymerase conformation for efficient, productive viral DNA replication.