Tumor suppressor p53 is reported to be an attractive immunotherapy target because it is mutated in approximately half of human cancers, resulting in its inactivation and often an accumulation of the protein in the tumor cells. Peptides derived from over-expressed p53 protein are presented by class I MHC molecules and may act as tumor-associated epitopes. Due to the diversity of p53 mutations, immunogenic peptides representing wild-type sequences are preferable as a basis for a broad-spectrum p53-targeting cancer therapeutic. It has been shown that PBLs transduced with a murine TCR isolated from T cell clones specific for the p53 (264–272) epitope, derived from HLA-A2.1 transgenic mice, were capable of lysing p53-expressing tumor cell lines, while p53-negative lines and normal cells were ignored. However, it was recently shown that there was no significant correlation between p53 expression in tumors and recognition by the anti-p53 TCR transduced T cells. Thus, the purpose of this study is to understand the influence of p53 mutations on its presentation by the MHC system. To that end, we have generated several constructs encoding different p53 mutant proteins and have expressed them in HLA-A2+/p53- cells. Expression of the different mutants was assessed by intracellular flow cytometry. Upon co-culture with p53-specific T cells, we measured specific recognition of the p53 expressing cells by means of cytokine secretion as IFNγ and TNFα and the expression of the activation antigen CD69. Our data show that there is an inverse correlation between intracellular expression and recognition by T-cells, and thus we hypothesize that p53 mutant stability can greatly influence its recognition by the immune system. This pattern was redundant in several cell lines tested, and was not associated with any influence on class I MHC levels of expression, suggesting it could represent an intrinsic property of p53 mutant proteins. Thus, these results may have significant therapeutic implications for p53-based immunotherapy of cancer.
