Many tumour cells express high levels of proteins which are present in normal cells at lower levels. Such proteins might be targets for immunotherapy if lymphocytes with specificity for these proteins are present in the repertoire and can discriminate between cells expressing high or low levels of the target protein. The first step in exploring the possibility of targeting immunotherapy to normal self proteins is therefore to determine whether lymphocytes specific for molecules overexpressed in tumours exist and can be activated to recognize specifically cells expressing high levels of the proteins. It was possible to stimulate murine cyclin D1 or mdm2 specific cytotoxic T lymphocytes (CTL) by in vivo priming of mice with recombinant vaccinia virus expressing the proteins. The in vitro conditions used for restimulation of in vivo primed CTL were found to have a profound effect on the magnitude of the CTL response observed in vitro. In addition, in vitro priming of CTL from naïve spleens with peptide pulsed dendritic cells identified a mdm2 derived peptide, mdp441, which could stimulate CTL capable of recognising endogenously processed mdm2. These peptide specific CTL were of high avidity in contrast to most of the peptide specific CTL stimulated with other self peptides. The mdp441 peptide was not recognised by mdm2 specific CTL generated by in vivo immunisation with mdm2 protein. The results show that tolerance to self proteins is not absolute and that it is possible to stimulate CTL to recognize endogenously processed self protein either by in vivo immunisation with recombinant vaccinia virus expressing the self protein or by in vitro priming of naive responder cells with peptide pulsed antigen presenting cells. These results have implications for immunotherapy of human cancers
