Solid organ transplantation has become a leading therapy for children with a variety of end stage organ diseases. However, the immunosuppression required for prevention of graft rejection places transplant recipients at increased risk of serious opportunistic viral infections that can have deleterious effects on graft and patient survival. In pediatric transplant recipients, Epstein Barr virus (EBV)-associated complications are of particular concern. The clinical manifestations of EBV infection are complex and can range from asymptomatic viremia to aggressive B cell lymphomas associated with post-transplant lymphoproliferative disorder (PTLD). A major question in transplantation has been to understand why some children can control EBV infection while others develop serious life-threatening complications. Thus, new strategies are needed to develop more personalized approaches for diagnosis and treatment of pediatric transplant recipients infected with EBV. We hypothesize that EBV infection shapes the post-transplant innate and adaptive immune response in children and that these changes can be exploited to identify unique immune-based signatures that promote functional EBV immunity and long-term graft survival. To test this hypothesis we propose to utilize extra samples collected as part of CTOT-C-06, ?Biomarkers for Post-Transplant Lymphoproliferative Disorders in Children? (PI:Esquivel) and to analyze existing data from CTOT-C-02, ?Immune Development in Pediatric Transplantation? (PI:Kirk). We have generated strong preliminary data indicating that: 1) EBV viremia is common in the first year post-transplant in pediatric allograft recipients; 2) NK cells and T cells are increased in lesions of EBV+ PTLD lymphomas compared to EBV- B cell lymphomas; 3) a specific subset of NK cells that express NKG2A+ is capable of responding to, and killing, EBV-infected cells; 4) clonally expanded EBV- specific T cells that utilize TCR with shared sequences within the antigen-binding portion can be identified in EBV-infected individuals; and 5) patients with EBV+ B cell lymphomas secrete immunomodulatory cytokines that can influence the host immune response. In the first Aim we mine data previously obtained in CTOT-C- 02 to investigate NK cell phenotypes and T cell signatures in the context of viral infection and clinical outcome. We determine the relationship between NKG2A+ NK cells, viral load, and control of EBV infection and utilize mass cytometry (cytometry time of flight, CyTOF) to obtain a complete phenotype of EBV-reactive NKG2A+ cells. In the second Aim we use single cell assays to link T cell receptor usage and effector phenotype to reveal the signature of EBV-specific T cells associated with immune protection from EBV.
In Aim 3 we address the enigma of why many patients who have had immunosuppression halted as a first line response to the diagnosis of EBV+PTLD maintain their allograft without returning to immunosuppression. We use CyTOF to determine whether recovery from EBV+ PTLD in the absence of immunosuppression favors the development of IL-10-producing B regulatory cells. Together, these studies will provide novel mechanistic insights into the immune response to EBV and will facilitate improvements in diagnosis, management and treatment of EBV disease in pediatric transplant recipients.

Public Health Relevance

Solid organ transplantation has become an effective therapeutic option for children with end stage organ disease. However, the immunosuppression required for prevention of immune-mediated rejection of the graft also places children at increased risk of infection and malignancy. Epstein Barr virus (EBV) is a common virus in the human population that is linked to serious complications, including cancer, in pediatric transplant recipients. Effective diagnosis and medical management of EBV-related disease is hampered by our lack of understanding of the immune mechanisms that determine the outcome of EBV infection. In this proposal we will use cutting edge approaches to characterize on a single cell level, the immune response to EBV in children whose immune system is controlling the virus compared to those that are not. These studies will provide new opportunities to improve detection, diagnosis and management of EBV disease and cancer in pediatric transplant recipients.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project--Cooperative Agreements (U01)
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Special Emphasis Panel (ZAI1)
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Hayes, Deborah
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Stanford University
Schools of Medicine
United States
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