Epstein-Barr virus antibodies in patients with ataxia-telangiectasia and other immunodeficiency diseases

An unusual antibody response to the Epstein—Barr virus (EBV) has been noted in patients with ataxia—telangiectasia. Of a group of 16 such patients 8 were found to have antibodies in their serum to the EBV viral capsid antigen (VCA), and 4 of them also had antibodies to the EBV early antigen (EA); antibodies to the nuclear antigen (EBNA), however, were absent in 3 of the 8. The antibody pattern persisted for more than 2 years in the patients available for follow-up study. In comparison, of 24 patients with various other immunodeficiency syndromes 9 were found to have EBV-VCA antibodies in their serum, but none of the 9 had EA antibodies and 3 lacked EBNA antibodies. Two other groups of subjects, all of whom had EBV-VCA and EBNA antibodies in their serum late after an EBV infection, were also studied; 82 had infectious mononucleosis and 55 were healthy and had no such history. EA antibodies were detected in 45 of the first group during the acute phase of the illness but persisted in only 6 of the 68 who were followed up for more than 2 years, and they were detected in only 7 of the second group.All eight lymphoblastoid cell lines established from the peripheral blood of the four patients with ataxia—telangiectasia who are still available for follow-up study express EBV-VCA, whereas most similar cell lines established from normal individuals express only EBNA. In two of these patients cell-mediated immunity, as assessed from lymphocyte transformation induced by mitogens, was markedly decreased but autologous cell-mediated immune regression of EBV-induced transformation of B (bone-marrow-derived)-lymphocytes was normal. The percentage of T (thymus-derived)-helper cells was greatly decreased in two of the three patients in whom it was measured, and the percentage of T-suppressor cells was greatly increased in one of them, but the percentage of total T-lymphocytes was within normal limits in all three.The possible significance of these findings — in particular, the persistence of EA antibodies and the diminished restriction of expression of EA — in the late development of tumours after an EBV infection in patients with ataxia-telangiectasia deserves careful attention. Finally, the apparent correlation between immunoglobulin deficiency and poor or absent EBNA antibody response warrants further study.     

Identification of multiple Epstein-Barr virus-induced nuclear antigens with sera from patients with rheumatoid arthritis.

By means of the protein immunoblot technique, the Epstein-Barr virus (EBV) nuclear antigen (EBNA) could be identified in a variety of EBV-transformed cell lines with anti-EBNA-positive sera from normal donors. The molecular weight of EBNA expressed in each of the cell lines varied between 70,000 and 75,000 and was dependent upon the strain of infecting virus. In contrast, 15 of 21 sera from patients with rheumatoid arthritis identified antigens in addition to EBNA. The most prominent of these antigens had molecular weights of 110,000 to 115,000 and 92,000. All of the EBV genome-positive cell lines except for QIMR-GOR and cell lines containing the P3HR-1 virus expressed these antigens. The antigens were not present in the EBV genome-negative Ramos and BJAB cell lines, nor were they identified with EBV seronegative sera, indicating that they were EBV related. There was no direct correlation between the presence of antibodies in sera to EBNA, viral capsid antigen or early antigen, and reaction with the 92,000-molecular-weight antigen in immunoblots, indicating that this antigen was distinct from previously described EBV-related antigens.     

激活Raji细胞EB病毒早期抗原植物的研究

以前材料证明Raji细胞EB病毒早期抗原对于诊断鼻咽癌有很高的特异性。我们研究了1693种植物,试验发现52种植物具有激活EB病毒作用。本文报道了这些植物并讨论它们与鼻咽癌关系。     

Biological differences between epstein-barr virus (EBV) strains with regard to lymphocyte transforming ability, superinfection and antigen induction

EB virus (EBV) preparations derived from various producing lymphoblastoid cell lines (LCL) differed in their biological properties, as judged by the following four tests: (1) cord blood lymphocyte (CBL) transformation into EBV-carrying LCL; (2) early antigen (EA) induction in Raji cells; (3) inhibition of Raji cell growth; (4) induction of the EBV-determined nuclear antigen (EBNA) in CBL. B95-8 virus transformed and induced EBNA in CBL but did not induce EA in Raji cells, nor did it inhibit their growth. P3HR-1 virus did not transform CBL, induced no EBNA or EA in CBL, but induced EA in Raji cells and inhibited their growth. EBV isolated from the QIMR-WIL, 833L, F137 and cb-8-7 LCL resembled the B95-8 virus with regard to its biological activity (CBL transformation, EA induction in and growth inhibition of Raji cells). Transformation of CBL as contrasted to EA induction in, and growth inhibition of Raji cells thus appear as mutually exclusive viral functions.     

An Epstein-Barr virus isolated from a lymphoblastoid cell line has a 16-kilobase-pair deletion which includes gp350 and the Epstein-Barr virus nuclear antigen 3A

Epstein-Barr virus (EBV) is associated with human cancers, including nasopharyngeal carcinoma, Burkitt's lymphoma, gastric carcinoma and, somewhat controversially, breast carcinoma. EBV infects and efficiently transforms human primary B lymphocytes in vitro. A number of EBV-encoded genes are critical for EBV-mediated transformation of human B lymphocytes. In this study we show that an EBV-infected lymphoblastoid cell line obtained from the spontaneous outgrowth of B cells from a leukemia patient contains a deletion, which involves a region of approximately 16 kbp. This deletion encodes major EBV genes involved in both infection and transformation of human primary B lymphocytes and includes the glycoprotein gp350, the entire open reading frame of EBNA3A, and the amino-terminal region of EBNA3B. A fusion protein created by this deletion, which lies between the BMRF1 early antigen and the EBNA3B latent antigen, is truncated immediately downstream of the junction 21 amino acids into the region of the EBNA3B sequence, which is out of frame with respect to the EBNA3B protein sequence, and indicates that EBNA3B is not expressed. The fusion is from EBV coordinate 80299 within the BMRF1 sequence to coordinate 90998 in the EBNA3B sequence. Additionally, we have shown that there is no detectable induction in viral replication observed when SNU-265 is treated with phorbol esters, and no transformants were detected when supernatant is used to infect primary B lymphocytes after 8 weeks in culture. Therefore, we have identified an EBV genome with a major deletion in critical genes involved in mediating EBV infection and the transformation of human primary B lymphocytes that is incompetent for replication of this naturally occurring EBV isolate.     

Epstein-barr virus (EBV)-associated antibodies in a case of Burkitt's lymphoma during 14 years of sustained remission

Summary A Burkitt's lymphoma (BL) patient who sustained remission for more than 14 years after chemotherapy was monitored by means of serial serum samplings. The sera were titrated for antibodies against the Epstein-Barr virus (EBV)-associated cell membrane antigen (MA), viral capsid antigens (VCA), early antigen complex (EA R/D), and nuclear antigen (EBNA), and also for reactivity in the antibody-dependent cellular cytotoxicity (ADCC) test. The initial serological profile corresponded to that of most BL patients with active disease. During remission, it changed to resemble that of normal persons with persistent, latent EBV infection, at least qualitatively. The prognostic and biological implications of the titer levels and their changes are discussed.Abbreviations ADCC antibody-dependent cellular cytotoxicity - BCG Bacillus Calmette-Guérin - BL Burkitt's lymphoma - D diffuse component of EA - EA Epstein-Barr virus-induced early antigens - EBNA Epstein-Barr virus-specific nuclear antigen - EBV Epstein-Barr virus - KCC Kenya Cancer Council Registry - MA EBV-associated cell membrane antigen complex - NK natural killer - R restricted component of EA - VCA EB viral capsid antigen complex     

Detection of Epstein-Barr virus-associated antigen in fine needle aspiration smears from cervical lymph nodes in the diagnosis of nasopharyngeal carcinoma

Epstein-Barr virus (EBV) has consistently been shown to be associated with undifferentiated nasopharyngeal carcinoma, and an EBV-associated nuclear antigen (EBNA) has been detected in the cells of nasopharyngeal carcinoma. A study on the applicability of EBNA detection in fine needle aspiration (FNA) smears from cervical lymph nodes in the diagnosis of metastatic nasopharyngeal carcinoma was performed. All 11 cases (100%) with metastatic nasopharyngeal carcinoma showed EBNA-positive tumor cells, characterized by bright, granulated nuclear fluorescence. Three (50%) of six cases with other metastatic head and neck carcinomas also showed EBNA-positive tumor cells. These findings suggest that the presence of EBNA-positive tumor cells in FNA smears from cervical lymph nodes is not specific for metastatic nasopharyngeal carcinoma. On the other hand, a negative result in the presence of tumor cells may help to exclude it. A larger study is required to verify these preliminary findings.     

Amplification of Epstein-Barr virus (EBV) DNA by superinfection with a strain of EBV derived from nasopharyngeal carcinoma.

Epstein-Barr virus (EBV) from a nasopharyngeal carcinoma (NPC) hybrid cell line (NPC-KT) lacking defective viral DNA molecules superinfected Raji cells and induced EBV early antigens (EA), as did virus from P3HR-1 cells, which contained defective molecules. The EBV polypeptides induced by NPC-KT appeared to be identical to those induced by P3HR-1 virus. The ability of NPC-KT virus to induce EA was enhanced more than 10-fold by treatment of superinfected cells with dimethyl sulfoxide; however, dimethyl sulfoxide treatment did not enhance superinfection by P3HR-1 virus. After infection, DNA synthesis of both the superinfecting NPC-KT virus and the resident Raji viral genome was induced. In addition to amplified Raji EBV episomal DNA, a fused terminal fragment of NPC-KT viral DNA was detected. The detection of fused terminal DNA fragments suggests that the superinfecting virion DNA either circularizes or polymerizes after superinfection and is possibly amplified through circular or concatenated replicative intermediates.     

Linkage between STAT regulation and Epstein-Barr virus gene expression in tumors

Epstein-Barr virus (EBV) latency gene expression in lymphoblastoid cell lines is regulated by EBNA2. However, the factors regulating viral expression in EBV-associated tumors that do not express EBNA2 are poorly understood. In EBV-associated tumors, EBNA1 and frequently LMP1 are synthesized. We found that an alternative latent membrane protein 1 (LMP1) promoter, L1-TR, located within the terminal repeats is active in both nasopharyngeal carcinoma and Hodgkin's disease tissues. Examination of the L1-TR and the standard ED-L1 LMP1 promoters in electrophoretic mobility shift assays revealed that both promoters contain functional STAT binding sites. Further, both LMP1 promoters responded in reporter assays to activation of JAK-STAT signaling. Cotransfection of JAK1 or v-Src or treatment of cells with the cytokine interleukin-6 upregulated expression from ED-L1 and L1-TR reporter plasmids. Cotransfection of a dominant negative STAT3 beta revealed that STAT3 is likely to be the biologically relevant STAT for EBNA1 Qp and LMP1 L1-TR promoter regulation. In contrast, LMP1 expression from ED-L1 was not abrogated by STAT3 beta, indicating that the two LMP1 promoters are regulated by different STAT family members. Taken together with the previous demonstration of JAK-STAT activation of Qp driven EBNA1 expression, this places two of the EBV genes most commonly expressed in tumors under the control of the same signal transduction pathway. Immunohistochemical analyses of nasopharyngeal carcinoma tumors revealed that STAT3, STAT5, and STAT1 are constitutively activated in these tumors while STAT3 is constitutively activated in the malignant cells of Hodgkin's disease. We hypothesize that chronic or aberrant STAT activation may be both a necessary and predisposing event for EBV-driven tumorigenesis in immunocompetent individuals.     

A familial syndrome of susceptibility to chronic active Epstein-Barr virus infection.

In two members of a family (daughter and father) active Epstein-Barr virus (EBV) infections persisted over periods of 4 and 3 years respectively (possibly 10 years in the father). Both had persistent splenomegaly and occasional bouts of unexplained fever but lived otherwise normal lives. The other members of the family (mother and son) were healthy. The titres of antibody to the EBV viral capsid antigen (VCA) and early antigen (EA) were extremely high in the daughter''s blood, whereas the titres of antibody to the Epstein-Barr nuclear antigen were low in the daughter''s blood and undetectable in the father''s. Target cells of the EBV infection that were obtained from the daughter''s blood were established in culture with great difficulty and showed increased expression of VCA and EA. Other immunologic investigations in the two patients revealed that the ratio of helper to suppressor T lymphocytes was inverted, natural killer-cell activity was abnormally low, lymphocyte responses to certain mitogens were depressed and there was a serum factor blocking mitogen-induced transformation. The possibility that the patients'' unusual susceptibility to EBV infection represented an inherited syndrome (perhaps X-linked) is discussed.     

Human immune responses to synthetic peptides from the Epstein-Barr nuclear antigen

Humans infected with Epstein-Barr virus (EBV), the causative agent of infectious mononucleosis, develop antibodies against a nuclear antigen (EBNA) that is present in virally transformed B lymphocytes. The EBNA protein contains a unique glycine-alanine repeating sequence. We have synthesized peptides corresponding to various regions of the EBNA molecule within and near this sequence. Rabbit antibodies against the peptides within the sequence reacted directly with the EBNA protein, as detected by Western blotting. The sera of individuals with antibodies against Epstein-Barr virus contained abundant antibodies also reactive with one or several of the synthetic peptides within the sequence. Moreover, human antibodies against these simple peptides were induced specifically early in the course of infectious mononucleosis. When compared with normal controls, antibody levels to the glycine-alanine peptides were significantly higher in patients with rheumatoid arthritis and progressive systemic sclerosis, but not in patients with two other autoimmune diseases. These results document that i) antibodies against the peptides detect the EBNA protein, ii) humans infected with EBV produce high titers of antibodies reactive with these synthetic antigens, and iii) antibody titers against the peptides are abnormally elevated in certain autoimmune diseases.     

Variations of Epstein-Barr Virus Nuclear Antigen 1 in Epstein-Barr Virus-Associated Gastric Carcinomas from Guangzhou,Southern China

Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) is the only viral protein consistently expressed in all EBV-associated malignancies, and play a critical role in the onset, progression, and/or maintenance of these tumors. Based on the signature changes at amino acid residue 487, EBNA1 is classified into five distinct subtypes: P-ala, P-thr, V-leu, V-val and V-pro. In the present study, the sequence variations of EBNA1 in EBV-associated gastric carcinoma (EBVaGC) and throat washing (TW) samples of healthy EBV carriers in Guangzhou, southern China, where nasopharyngeal carcinoma (NPC) is endemic, were analyzed by PCR and DNA sequencing. V-val subtype was the most predominant (53.6%, 15/28) in EBVaGC, followed by P-ala (42.9%, 12/28) and V-leu (32.1%, 9/28) subtypes. In TWs of healthy EBV carriers, V-val subtype was also predominant (85.7%, 18/21). The sequence variations of EBNA1 in EBVaGC were similar to those in TW of healthy EBV carriers (p>0.05), suggesting that the EBV strains in EBVaGC might originate from the viral strains prevalent within the background population. The predominance of V-val subtype in EBVaGC in Guangzhou was similar to that in EBVaGC in northern China and Japan, but was different from that in EBVaGC in America, suggesting that the variations of EBNA1 in EBVaGC represent geographic-associated polymorphisms rather than tumor-specific mutations. In addition, the EBNA1 variations in EBVaGC in gastric remnant carcinoma were also determined. V-leu subtype was detected in all 4 (100%) cases, although 2 cases occurred as mixed infection with P-ala subtype. This is different from the predominant V-val subtype in EBVaGC in conventional gastric carcinoma, suggesting that V-leu might be a subtype that adapts particularly well to the microenvironment within the gastric stump and enters the remnant gastric mucosa epithelia easily. This, to our best knowledge, is the first investigation of EBNA1 polymorphisms in EBVaGC from endemic area of NPC.     

Proteomic profiling of EBNA1-host protein interactions in latent and lytic Epstein-Barr virus infections

The Epstein-Barr nuclear antigen 1 (EBNA1) protein of Epstein-Barr virus (EBV) is expressed in both latent and lytic modes of EBV infection and contributes to EBV-associated cancers. Using a proteomics approach, we profiled EBNA1-host protein interactions in nasopharyngeal and gastric carcinoma cells in the context of latent and lytic EBV infection. We identified several interactions that occur in both modes of infection, including a previously unreported interaction with nucleophosmin and RNA-mediated interactions with several heterogeneous ribonucleoproteins (hnRNPs) and La protein.     

Epstein-Barr viral antigens used in the diagnosis of nasopharyngeal carcinoma

The major antigen complexes of Epstein-Barr virus (EBV) include the latent infectious proteins, early antigens, membrane antigens and viral capsid antigens. The various polypeptides within each antigen complex have been identified and isolated through gene-cloning technology. These polypeptides are exploited to be used as serological markers for the diagnosis of nasopharyngeal carcinoma (NPC) through enzyme-linked immunosorbent assay. This paper reviews the recent studies on the profile of antibodies in patients with NPC towards these EBV polypeptides of each antigen complex. The sensitivity and specificity of each polypeptide when used as serological markers to NPC patients' sera are summarized.     

Abnormal anti-Epstein Barr virus antibodies in carriers of the X-linked lymphoproliferative syndrome and in females at risk

The asymptomatic hemizygous female carriers of the X-linked lymphoproliferative syndrome (XLP) have abnormal antibody responses to EBV. This suggests partial expression of the defect that leads to EBV-provoked life-threatening diseases in their affected sons. EBV specific antibodies were measured in 65 serum samples of 12 obligate carrier females and seven of their daughters (females at risk) during periods ranging from 1 to 5 yr. Abnormal qualitative antiviral capsid antigen (VCA) IgG titers were nearly fourfold higher than normal controls, two carriers had persistent IgM anti-VCA antibody, two-thirds had persistent IgA anti-VCA antibody, and half of the women had titers to early antigen (EA). Five of seven females exhibited a similar persistent pattern. In contrast, none of the unaffected family members nor 23 normal controls expressed IgA or IgM titers to VCA even with high exposure to the virus, and anti-EA was detected in only one control. Therefore, these findings may prove useful for detecting carriers of the syndrome. Abnormal anti-EBV titers similar to the carrier pattern have been reported in patients and other immunosuppressed individuals, and are indicative of active viral infection.     

Epstein-Barr virus gene expression in malignant lymphomas induced by experimental virus infection of cottontop tamarins.

Inoculation of cottontop tamarins with a large dose of Epstein-Barr virus (EBV) leads to the induction of multiple EBV genome-positive lymphomas. These tumors have been characterized as oligoclonal or monoclonal large-cell malignant lymphomas that closely resemble the EBV genome-positive B-cell lymphomas that arise in human allograft recipients. The expression of latent and lytic EBV-encoded proteins was investigated in these virus-induced tamarin lymphomas and in derived cell lines. The tamarin tumors were found to express EBV nuclear antigen 1 (EBNA 1), EBNA 2, EBNA leader protein, and the latent membrane protein (LMP) as determined both by immunohistochemical staining and by immunoblotting. However, within the limits of the immunoblotting assays, no expression of the EBNA 3a protein family could be detected. Assays for lytic-cycle proteins by using both polyclonal human sera and monoclonal antibodies against viral capsid antigen, early antigen, and membrane antigen (gp340/220) showed minimal, if any, expression of these antigens in the lymphoma biopsies. In contrast, the cell lines derived from these lymphomas, even in early passage, expressed abundant levels of the lytic-cycle antigens and also expressed the EBNA 3a protein as well as EBNA 1, EBNA 2, EBNA leader protein, and LMP. This finding suggests that the virus-lymphoma cell interaction, in particular the switch to lytic cycle, is subject to some form of host control in vivo. The expression of EBNA 2 and LMP in these tamarin lymphomas strengthens their resemblance to posttransplant lymphomas in humans, since these human tumors are also EBNA 2 and LMP positive (L. S. Young, C. Alfieri, K. Hennessy, H. Evans, C. O'Hara, K. Anderson, A. Rickinson, E. Kieff, and J. I. Cohen, submitted for publication). Since both proteins are known to be important effector molecules of virus-induced B-cell growth transformation in vitro, their expression in these lymphomas constitutes the best evidence for a direct oncogenic role for EBV in vivo.     

Cellular immunity in infectious mononucleosis. II. Specific reactivity to Epstein-Barr Virus antigens and correlation with clinical and hematologic parameters.

Infectious mononucleosis (IM) patients, Epstein-Barr virus (EBV)-seropositive and seronegative healthy donors, and patients with other viral infections were tested for lymphocyte blastogenesis (LB) with phytohemagglutinin and six EBV (virus concentrate, culture supernatant, and soluble [S] antigen) or control antigens. Fluorescent antibodies to EBV viral capsid antigen of IgG, IgM, IgA specificities, to nuclear antigen (EBNA), and heterophile antibodies were also assayed. These were correlated with clinical parameters (fever, pharyngitis, adenopathy, hepatitis, splenomegaly, atypical lymphocytes, and total mononuclear cell counts). EBV viral and S antigen-induced LB was significantly greater in seropositive donors. IM patients had antigenspecific LB below that of seropositive donors initially and low responses for the acute phase of illness when clinical symptoms were present and antibody titers were maximal. Specific LB rose to a peak at 3.5 to 9 weeks when the patients had recovered, most laboratory findings had returned to normal, and antibodies had declined. At peak, specific LB in IM patients exceeded that of seropositive donors, but later declined. These results demonstrate specific cell-mediated immunity (CMI) to EBV, and indicate that this develops slowly in IM and contrasts with the evolution of the clinical events and humoral immunity. This correlation supports the hypothesis that CMI is the mechanism of terminating lymphoproliferation in IM.     

Characterization of the Variability of Epstein-Barr Virus Genes in Nasopharyngeal Biopsies: Potential Predictors for Carcinoma Progression

Epstein-Barr virus (EBV) infection is a significant factor in the pathogenesis of nasopharyngeal carcinoma, especially in the undifferentiated carcinoma of nasopharyngeal type (UCNT, World Health Organization type III), which is the dominant histopathological type in high-risk areas. The major EBV oncogene is latent membrane protein 1 (LMP1). LMP1 gene shows variability with different tumorigenic and immunogenic potentials. EBV nuclear antigen 1 (EBNA1) regulates progression of EBV-related tumors; however, the influence of EBNA1 sequence variability on tumor pathogenesis is controversial. The aims of this study were to characterize polymorphisms of EBV genes in non-endemic nasopharyngeal carcinoma biopsies and to investigate potential sequence patterns that correlate with the clinical presentation of nasopharyngeal carcinoma. In total, 116 tumor biopsies of undifferentiated carcinoma of nasopharyngeal type (UCNT), collected from 2008 to 2014, were evaluated in this study. The genes EBNA2, LMP1, and EBNA1 were amplified using nested-PCR. EBNA2 genotyping was performed by visualization of PCR products using gel electrophoresis. Investigation of LMP1 and EBNA1 included sequence, phylogenetic, and statistical analyses. The presence of EBV DNA was significantly distributed between TNM stages. LMP1 variability showed six variants, with the detection of the first China1 and North Carolina variants in European nasopharyngeal carcinoma biopsies. Newly discovered variants Srb1 and Srb2 were UCNT-specific LMP1 polymorphisms. The B95-8 and North Carolina variants are possible predictors for favorable TNM stages. In contrast, deletions in LMP1 are possible risk factors for the most disfavorable TNM stage, independent of EBNA2 or EBNA1 variability. A newly discovered EBNA1 subvariant, P-thr-sv-5, could be a potential diagnostic marker, as it represented a UCNT-specific EBNA1 subvariant. A particular combination of EBNA2, LMP1, and EBNA1 polymorphisms, type 1/Med/P-thr was identified as a possible risk factor for TNM stage IVB or progression to the N3 stage.     

Dual stimulation of Epstein-Barr Virus (EBV)-specific CD4+- and CD8+-T-cell responses by a chimeric antigen construct: potential therapeutic vaccine for EBV-positive nasopharyngeal carcinoma

Virus-associated malignancies are potential targets for immunotherapeutic vaccines aiming to stimulate T-cell responses against viral antigens expressed in tumor cells. Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma, a high-incidence tumor in southern China, expresses a limited set of EBV proteins, including the nuclear antigen EBNA1, an abundant source of HLA class II-restricted CD4(+) T-cell epitopes, and the latent membrane protein LMP2, a source of subdominant CD8(+) T-cell epitopes presented by HLA class I alleles common in the Chinese population. We used appropriately modified gene sequences from a Chinese EBV strain to generate a modified vaccinia virus Ankara recombinant, MVA-EL, expressing the CD4 epitope-rich C-terminal domain of EBNA1 fused to full-length LMP2. The endogenously expressed fusion protein EL is efficiently processed via the HLA class I pathway, and MVA-EL-infected dendritic cells selectively reactivate LMP2-specific CD8(+) memory T-cell responses from immune donors in vitro. Surprisingly, endogenously expressed EL also directly accesses the HLA class II presentation pathway and, unlike endogenously expressed EBNA1 itself, efficiently reactivates CD4(+) memory T-cell responses in vitro. This unscheduled access to the HLA class II pathway is coincident with EL-mediated redirection of the EBNA1 domain from its native nuclear location to dense cytoplasmic patches. Given its immunogenicity to both CD4(+) and CD8(+) T cells, MVA-EL has potential as a therapeutic vaccine in the context of nasopharyngeal carcinoma.