Isolation of Epstein-Barr virus (EBV)-specific cytotoxic T lymphocytes that lyse Reed-Sternberg cells: Implications for immune-mediated therapy of EBV⁺ Hodgkin's disease

A subset of Hodgkin's disease (HD) patients have detectable Epstein- Barr virus (EBV) genomes in the malignant Reed-Sternberg (R-S) cells, R-S cells express only a limited set of latent EBV proteins, but only LMP1 and LMP2 can potentially elicit a CD8⁺ cytotoxic T-lymphocyte (CTL) response. We have evaluated if either of these proteins could be used as targets for specific adoptive T-cell therapy for EBV-positive (EBV⁺) HD. The success of this strategy requires that R-S cells are susceptible to lysis by CD8⁺ CTL, and that CTL specific for LMP1 and LMP2 can be detected and potentially amplified in HD patients. Antigen presentation and CTL sensitivity was evaluated with an in vitro maintained, phenotypically representative R-S cell line, HDLM-2. The R-S cells were able to process and present viral proteins, and to be efficiently lysed by specific CTL in a Class I-restricted manner. Since CTL responses to LMP1 and LMP2 do not represent the dominant responses to EBV, we examined if CTL clones specific for these proteins could be isolated despite the presence of weak of nondetectable responses in polyclonal T-cell lines. LMP-specific clones were generated from individuals either by cloning from the polyclonal EBV-reactive T-cell lines or by direct stimulation of peripheral blood mononuclear cells (PBMC) with cells expressing LMP1 or LMP2 as the only EBV protein. Our ability to isolate CTL specific for LMP proteins from individuals with HD and the sensitivity of R- S cells for CTL-mediated lysis suggest that the pursuit of specific adoptive immunotherapy represents a viable strategy for the subset of HD patients with EBV⁺ tumors.