Immunosuppressive FK506 treatment leads to more frequent EBV-associated lymphoproliferative disease in humanized mice

Post-transplant lymphoproliferative disorder (PTLD) is a potentially fatal complication after organ transplantation frequently associated with the Epstein-Barr virus (EBV). Immunosuppressive treatment is thought to allow the expansion of EBV-infected B cells, which often express all eight oncogenic EBV latent proteins. Here, we assessed whether HLA-A2 transgenic humanized NSG mice treated with the immunosuppressant FK506 could be used to model EBV-PTLD. We found that FK506 treatment of EBV-infected mice led to an elevated viral burden, more frequent tumor formation and diminished EBV-induced T cell responses, indicative of reduced EBV-specific immune control. EBV latency III and lymphoproliferation-associated cellular transcripts were up-regulated in B cells from immunosuppressed animals, akin to the viral and host gene expression pattern found in EBV-PTLD. Utilizing an unbiased gene expression profiling approach, we identified genes differentially expressed in B cells of EBV-infected animals with and without FK506 treatment. Upon investigating the most promising candidates, we validated sCD30 as a marker of uncontrolled EBV proliferation in both humanized mice and in pediatric patients with EBV-PTLD. High levels of sCD30 have been previously associated with EBV-PTLD in patients. As such, we believe that humanized mice can indeed model aspects of EBV-PTLD development and may prove useful for the safety assessment of immunomodulatory therapies. Author summary Transplant recipients are medicated with potent immunosuppressive drugs, like FK506, to prevent graft rejection by the host's adaptive immune system. Such treatments can lead to the emergence of post-transplant lymphoproliferative disorders (PTLD) driven by the Epstein-Barr virus (EBV), a ubiquitous human oncovirus that is usually kept under control by T cells. Here, we aimed to establish a model of human EBV-PTLD. To this end, we investigated immunodeficient mice harboring human immune system components reconstituted from human hematopoietic progenitor cells, termed humanized mice. This model enables both the infection of human B cells with EBV and the examination of effects of immunosuppressive compounds on lymphocytes in vivo. We found that EBV-associated lymphoproliferations in humanized mice express characteristic viral and human genes observed in EBV-PTLD patients and found similarities in the profiles of serum proteins known for their association with the disease. As such, we believe that EBV-infected humanized mice treated with the immunosuppressive drug FK506 can be used to model specific aspects of EBV-PTLD. Conversely, similar models may prove useful in the preclinical risk-assessment of novel compounds in relation to EBV-associated lymphoproliferation and our study may serve as a template of how one could approach such investigations.