{"id":{"repo_id":"penn","oai_identifier":"oai:repository.upenn.edu:20.500.14332/28145"},"canonical_url":"https://search.dev.ndltd.org/etd/penn/oai:repository.upenn.edu:20.500.14332/28145","repository":{"repo_id":"penn","name":"University of Pennsylvania","base_url":"https://repository.upenn.edu/server/oai/request"},"display":{"title":"Ebna1-Specific T Cell Responses During Persistent Rhesus Lcv infection and The Development of a Novel Therapeutic Prototype Vaccine for Ebv-Associated Diseases","abstract":"The impact of EBV on human health is substantial, but vaccines that prevent primary EBV infections or treat EBV-associated diseases are not yet available. The Epstein-Barr nuclear antigen 1 (EBNA1) is an important target for vaccination because it is the only protein expressed in all forms of latency and in all EBV-associated malignancies. The overarching goal throughout this dissertation was to determine if EBNA1 is a suitable target for vaccine development. This was addressed in two ways. First, because an improved understanding of EBNA1-specific T cell responses benefits EBV vaccine development, we characterized responses against EBNA1 of the EBV-homologous rhesus lymphocryptovirus (rhLCV) in naturally infected rhesus macaques. We assessed frequency, phenotype, and cytokine production profiles of rhesus (rh)EBNA1-specific T cells by intracellular cytokine staining (ICS) and polychromatic flow cytometry. We found that most naturally infected animals had CD4+ and/or CD8+ T cells against rhEBNA1 and rhBZLF1, an immediate-early lytic phase antigen of rhLCV. Peptide-specific CD8+ T cells showed a more activated effector phenotype, while most peptide-specific CD4+ T cells exhibited a resting central memory phenotype. T cells were highly functional and produced various combinations of the cytokines IFN-γ, IL-2, and TNF-α. The differentiation status and functional profiles of rhEBNA1-specific T cells suggests they are not impaired by chronic exposure to low levels of antigen, and rhEBNA1-specific T cells therefore represent a viable population to target through vaccination. Similarities between our findings and the human response further validate the rhLCV model for studying chronic EBV infection and for pre-clinical vaccine development. We then asked if vaccination could expand functional rhEBNA1-specific T cells during persistent rhLCV infection. To test this, we developed two serologically distinct replication-defective adenoviral vectors that expressed chimeric rhEBNA1 constructs fused to functional and non-functional versions of Herpes Simplex Virus- glycoprotein D (HSV-gD). HSV-gD has been shown to augment T cell responses by inhibiting the immunosuppressive Herpes Virus Entry Mediator (HVEM) pathway during T cell activation. After confirmation of vaccine specificity and antigenicity in vitro, rhesus macaques were vaccinated in a prime-boost regimen, and responses in peripheral blood were measured by ICS and polychromatic flow cytometry. Importantly, we found that vaccination induced the expansion of highly functional rhEBNA1-specific CD8+ and CD4+ T cells in vivo, regardless of HSV-gD binding ability. Vaccination did not increase rhBZLF1-specific T cell responses, thus indicating that rhEBNA1-specific responses were vaccine-driven. Overall, these results serve as important proof-of-principle analyses of a therapeutic EBNA1-based vaccine regimen and demonstrate that EBNA1 is a viable target that warrants exploration in future vaccine studies.","abstract_html":"The impact of EBV on human health is substantial, but vaccines that prevent primary EBV infections or treat EBV-associated diseases are not yet available. The Epstein-Barr nuclear antigen 1 (EBNA1) is an important target for vaccination because it is the only protein expressed in all forms of latency and in all EBV-associated malignancies. The overarching goal throughout this dissertation was to determine if EBNA1 is a suitable target for vaccine development. This was addressed in two ways. First, because an improved understanding of EBNA1-specific T cell responses benefits EBV vaccine development, we characterized responses against EBNA1 of the EBV-homologous rhesus lymphocryptovirus (rhLCV) in naturally infected rhesus macaques. We assessed frequency, phenotype, and cytokine production profiles of rhesus (rh)EBNA1-specific T cells by intracellular cytokine staining (ICS) and polychromatic flow cytometry. We found that most naturally infected animals had CD4+ and/or CD8+ T cells against rhEBNA1 and rhBZLF1, an immediate-early lytic phase antigen of rhLCV. Peptide-specific CD8+ T cells showed a more activated effector phenotype, while most peptide-specific CD4+ T cells exhibited a resting central memory phenotype. T cells were highly functional and produced various combinations of the cytokines IFN-γ, IL-2, and TNF-α. The differentiation status and functional profiles of rhEBNA1-specific T cells suggests they are not impaired by chronic exposure to low levels of antigen, and rhEBNA1-specific T cells therefore represent a viable population to target through vaccination. Similarities between our findings and the human response further validate the rhLCV model for studying chronic EBV infection and for pre-clinical vaccine development. We then asked if vaccination could expand functional rhEBNA1-specific T cells during persistent rhLCV infection. To test this, we developed two serologically distinct replication-defective adenoviral vectors that expressed chimeric rhEBNA1 constructs fused to functional and non-functional versions of Herpes Simplex Virus- glycoprotein D (HSV-gD). HSV-gD has been shown to augment T cell responses by inhibiting the immunosuppressive Herpes Virus Entry Mediator (HVEM) pathway during T cell activation. After confirmation of vaccine specificity and antigenicity in vitro, rhesus macaques were vaccinated in a prime-boost regimen, and responses in peripheral blood were measured by ICS and polychromatic flow cytometry. Importantly, we found that vaccination induced the expansion of highly functional rhEBNA1-specific CD8+ and CD4+ T cells in vivo, regardless of HSV-gD binding ability. Vaccination did not increase rhBZLF1-specific T cell responses, thus indicating that rhEBNA1-specific responses were vaccine-driven. Overall, these results serve as important proof-of-principle analyses of a therapeutic EBNA1-based vaccine regimen and demonstrate that EBNA1 is a viable target that warrants exploration in future vaccine studies.","abstract_has_math":false,"creators":["Leskowitz, Rachel Mandy"],"institution":null,"degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":["Hildegund C. Ertl"],"committee_chairs":[],"committee_members":[],"year":2014,"date_issued":"2014-01-01","date_published":"2014-01-01","updated_at":"2026-07-24T03:46:44Z","subjects":[],"languages":["en"],"rights":["Rachel Mandy Leskowitz"],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://repository.upenn.edu/handle/20.500.14332/28145","outbound_label":"Repository record","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Hildegund C. 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The Epstein-Barr nuclear antigen 1 (EBNA1) is an important target for vaccination because it is the only protein expressed in all forms of latency and in all EBV-associated malignancies. The overarching goal throughout this dissertation was to determine if EBNA1 is a suitable target for vaccine development. This was addressed in two ways. First, because an improved understanding of EBNA1-specific T cell responses benefits EBV vaccine development, we characterized responses against EBNA1 of the EBV-homologous rhesus lymphocryptovirus (rhLCV) in naturally infected rhesus macaques. We assessed frequency, phenotype, and cytokine production profiles of rhesus (rh)EBNA1-specific T cells by intracellular cytokine staining (ICS) and polychromatic flow cytometry. We found that most naturally infected animals had CD4+ and/or CD8+ T cells against rhEBNA1 and rhBZLF1, an immediate-early lytic phase antigen of rhLCV. Peptide-specific CD8+ T cells showed a more activated effector phenotype, while most peptide-specific CD4+ T cells exhibited a resting central memory phenotype. T cells were highly functional and produced various combinations of the cytokines IFN-γ, IL-2, and TNF-α. The differentiation status and functional profiles of rhEBNA1-specific T cells suggests they are not impaired by chronic exposure to low levels of antigen, and rhEBNA1-specific T cells therefore represent a viable population to target through vaccination. Similarities between our findings and the human response further validate the rhLCV model for studying chronic EBV infection and for pre-clinical vaccine development. We then asked if vaccination could expand functional rhEBNA1-specific T cells during persistent rhLCV infection. To test this, we developed two serologically distinct replication-defective adenoviral vectors that expressed chimeric rhEBNA1 constructs fused to functional and non-functional versions of Herpes Simplex Virus- glycoprotein D (HSV-gD). HSV-gD has been shown to augment T cell responses by inhibiting the immunosuppressive Herpes Virus Entry Mediator (HVEM) pathway during T cell activation. After confirmation of vaccine specificity and antigenicity in vitro, rhesus macaques were vaccinated in a prime-boost regimen, and responses in peripheral blood were measured by ICS and polychromatic flow cytometry. Importantly, we found that vaccination induced the expansion of highly functional rhEBNA1-specific CD8+ and CD4+ T cells in vivo, regardless of HSV-gD binding ability. Vaccination did not increase rhBZLF1-specific T cell responses, thus indicating that rhEBNA1-specific responses were vaccine-driven. Overall, these results serve as important proof-of-principle analyses of a therapeutic EBNA1-based vaccine regimen and demonstrate that EBNA1 is a viable target that warrants exploration in future vaccine studies."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Doctor of Philosophy (PhD)"]},{"key":"dc:format.mimetype","label":"Dc Format Mimetype","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Ebna1-Specific T Cell Responses During Persistent Rhesus Lcv infection and The Development of a Novel Therapeutic Prototype Vaccine for Ebv-Associated Diseases"]}]}],"canonical_facts":{"dc:contributor.advisor":["Hildegund C. Ertl"],"dc:creator":["Leskowitz, Rachel Mandy"],"dc:date":["2023-05-17T12:50:26.000"],"dc:date.accessioned":["2023-05-22T16:30:56Z"],"dc:date.available":["2001-01-01T00:00:00Z"],"dc:date.issued":["2014-01-01"],"dc:description.abstract":["The impact of EBV on human health is substantial, but vaccines that prevent primary EBV infections or treat EBV-associated diseases are not yet available. The Epstein-Barr nuclear antigen 1 (EBNA1) is an important target for vaccination because it is the only protein expressed in all forms of latency and in all EBV-associated malignancies. The overarching goal throughout this dissertation was to determine if EBNA1 is a suitable target for vaccine development. This was addressed in two ways. First, because an improved understanding of EBNA1-specific T cell responses benefits EBV vaccine development, we characterized responses against EBNA1 of the EBV-homologous rhesus lymphocryptovirus (rhLCV) in naturally infected rhesus macaques. We assessed frequency, phenotype, and cytokine production profiles of rhesus (rh)EBNA1-specific T cells by intracellular cytokine staining (ICS) and polychromatic flow cytometry. We found that most naturally infected animals had CD4+ and/or CD8+ T cells against rhEBNA1 and rhBZLF1, an immediate-early lytic phase antigen of rhLCV. Peptide-specific CD8+ T cells showed a more activated effector phenotype, while most peptide-specific CD4+ T cells exhibited a resting central memory phenotype. T cells were highly functional and produced various combinations of the cytokines IFN-γ, IL-2, and TNF-α. The differentiation status and functional profiles of rhEBNA1-specific T cells suggests they are not impaired by chronic exposure to low levels of antigen, and rhEBNA1-specific T cells therefore represent a viable population to target through vaccination. Similarities between our findings and the human response further validate the rhLCV model for studying chronic EBV infection and for pre-clinical vaccine development. We then asked if vaccination could expand functional rhEBNA1-specific T cells during persistent rhLCV infection. To test this, we developed two serologically distinct replication-defective adenoviral vectors that expressed chimeric rhEBNA1 constructs fused to functional and non-functional versions of Herpes Simplex Virus- glycoprotein D (HSV-gD). HSV-gD has been shown to augment T cell responses by inhibiting the immunosuppressive Herpes Virus Entry Mediator (HVEM) pathway during T cell activation. After confirmation of vaccine specificity and antigenicity in vitro, rhesus macaques were vaccinated in a prime-boost regimen, and responses in peripheral blood were measured by ICS and polychromatic flow cytometry. Importantly, we found that vaccination induced the expansion of highly functional rhEBNA1-specific CD8+ and CD4+ T cells in vivo, regardless of HSV-gD binding ability. Vaccination did not increase rhBZLF1-specific T cell responses, thus indicating that rhEBNA1-specific responses were vaccine-driven. Overall, these results serve as important proof-of-principle analyses of a therapeutic EBNA1-based vaccine regimen and demonstrate that EBNA1 is a viable target that warrants exploration in future vaccine studies."],"dc:description.degree":["Doctor of Philosophy (PhD)"],"dc:format.mimetype":["application/pdf"],"dc:identifier.uri":["https://repository.upenn.edu/handle/20.500.14332/28145"],"dc:language":["en"],"dc:rights":["Rachel Mandy Leskowitz"],"dc:title":["Ebna1-Specific T Cell Responses During Persistent Rhesus Lcv infection and The Development of a Novel Therapeutic Prototype Vaccine for Ebv-Associated Diseases"],"dc:type":["Dissertation/Thesis"]},"updated_at":"2026-07-24T03:46:44Z"}