Development of a dendritic cell-based vaccine to enhance CD4+ T-cell responses to a breast tumour antigen

With 570,000 new cases in the world each year, breast cancer remains the commonest malignancy in women and comprises 18% of all female cancer. The failure of current therapies for the treatment of women with metastatic breast cancer has culminated in the emergence of immunotherapy as a potential means of eradicating residual disease following surgery.;The study sought to develop a dendritic cell-based vaccine which was capable of generating CD4+ T-cell proliferative responses against a known breast tumour antigen. p53 was selected as the antigen to be manipulated due to increasing evidence of T-cell responses to both mutant and wild type forms in breast cancer patients. As it is favourable to pulse dendritic cells with nucleic acids rather than recombinant protein or peptides it was necessary to target endogenous p53 to the MHC class II pathway to enhance presentation of p53 to CD4+ T-cells. cDNA encoding the signal peptide motif, YTPL, present in the cytoplasmic tail of HLA-DM, known to target HLA-DM to MHC class II compartments, was appended to the 3' terminus of p53. Experiments assessed the ability of YTPL to target p53 to the lysosomal compartment in transfected HeLa cells. Control constructs encoding p53 targeted to other cellular compartments were analysed. In order to transfect DC with nucleic acids, experiments were also performed to establish optimal transfection conditions utilising non-viral methods.;This study has successfully targeted p53 to the lysosomal compartment of HeLa cells by virtue of YTPL and has investigated the use of many non-viral techniques to transfect DC. Further work is necessary to determine which construct encoding p53 provides optimal access to the MHC class II pathway and to improve transfection efficiencies of dendritic cells. Thus, this study has paved the way for a dendritic cell-based vaccine for breast cancer.