Mature dendritic cells pulsed with freeze–thaw cell lysates define an effective in vitro vaccine designed to elicit EBV-specific CD4+ and CD8+ T lymphocyte responses

Abstract

Abstract Immunotherapy trials targeting the induction of tumor-reactive T-cell responses in cancer patients appear to hold significant promise. Because nonmutated lineage-specific antigens and mutated idiotypic antigens may be coexpressed by tumor cells, the use of autologous tumor material to promote the broadest range of antitumor T-cell specificities has significant clinical potential in cancer vaccination trials. As a model for vaccination in the cancer setting, we chose to analyze the promotion of T-cell responses against Epstein-Barr virus (EBV)-transformed B-lymphoblastoid cell line (B-LCL)–derived antigens in vitro. A series of bulk antigenic formats (freeze–thaw lysate, trifluoroacetic acid lysate, extracted membranes, affinity-purified MHC class I– and class II–presented peptides, acid-eluted peptides) prepared from EBV B-LCLs were tested for their ability to stimulate EBV B-LCL–reactive CD4+ and CD8+ T lymphocytes in vitro when pulsed onto autologous dendritic cells (DCs). DC presentation of freeze–thaw lysate material derived from (either autologous or allogeneic) EBV B-LCLs with an Mr of 10 kd or larger stimulated optimal anti-EBV B-LCL responsiveness from freshly isolated CD4+ and CD8+ peripheral blood T cells. These in vivo “memory” T-cell responses were observed only in EBV-seropositive donors. CD4+ T-cell responses to lysate-pulsed DCs were Th1 type (ie, strong interferon-γ and weak interleukin-5 responses). While CD8+ T-cell responses were also observed in interferon-γ Elispot assays and in cytotoxicity assays, these responses were of low frequency unless the DC stimulators were induced to “mature” after being fed with tumor lysates. Optimal-length, naturally processed, and MHC class I– or class II–presented tumor peptides were comparatively poorly immunogenic in this model system.