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J. Virol. doi:10.1128/JVI.02227-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Standardized and highly efficient expansion of Epstein-Barr virus (EBV)-specific CD4+ T cells using virus-like particles

Clinical Cooperation Group, Department of Pediatrics, Munich University of Technology & GSF-Research Centre for Environment and Health, Munich, Germany; German Cancer Research Center, Department of Virus Associated Tumours, Heidelberg, Germany

^* To whom correspondence should be addressed. Email: mautner{at}gsf.de.

	Abstract

EBV-specific T cell lines generated by repeated stimulation with EBV-immortalized lymphoblastoid B cell lines (LCL) have been successfully used to treat EBV-associated post-transplant lymphoproliferative disease (PTLD) in hematopoietic stem cell transplant recipients. However, PTLD in solid organ transplant recipients and other EBV-associated malignancies respond less efficiently to this adoptive T cell therapy. LCL-stimulated T cell preparations are polyclonal and contain CD4+ and CD8+ T cells, but the composition varies greatly between lines. Because T cell lines with higher CD4+ T cell proportions show improved clinical efficacy, we assessed which factors might compromise the expansion of this T cell population. Here we show that spontaneous virus production by LCL and hence presentation of viral antigens varies intra- and interindividually, and is further impaired by acyclovir treatment of LCL. Moreover, stimulation of T cell with LCL grown in media supplemented with FCS caused expansion FCS-reactive CD4+ T cells, whereas human serum from EBV-seropositive donors diminished viral antigen presentation. To overcome these limitations, we used PBMC pulsed with non-transforming virus-like particles (VLP) as antigen-presenting cells. This strategy facilitated the specific and rapid expansion of EBV-specific CD4+ T cells and thus might contribute to the development of standardized protocols for the generation of T cell lines with improved clinical efficacy.