Exosomes are small membrane-bound vesicles of endosomal origin secreted by many cell types including tumour cells. Their potential use in cancer immunotherapy has been suggested by the discovery that tumour-derived exosomes can be a source of tumour associated antigens (T AA) and protect against tumour challenge in mice. The initial excitement has been tempered more recently however, by reports showing that exosomes can have inhibitory effects on T cells and natural killer (NK) cells. Heat-shock proteins are endogenous danger signals that activate the immune system; their expression is increased by heat-shock stress or v-irradiation. In this thesis, heat-shock stress and v-irradiation are investigated as mechanisms for increasing the immunogenicity of human and murine melanoma-derived exosomes. Exosomes derived from untreated or irradiated human melanoma cells did not activate DC, although heat-shock exosomes induced a small increase in expression of MHC-II, CD40 and inflammatory cytokines. Although exosome- induced maturation was far from optimal compared to LPS, exosomes did not inhibit the LPS-induced maturation of DC and could therefore be used to load DC with TAA for immunotherapy. The use of exosomes for priming a naive T cell response was investigated in a human in vitro system. Control, irradiated and heat-shock exosomes induced similar levels of cytotoxicity against melanoma cell lines suggesting that they were equally effective in generating an immune response. Murine melanoma-derived exosomes were investigated as anti-cancer vaccines in vivo in a mouse model. Neither control nor stressed exosomes, +/- DC were able to protect mice against subsequent tumour challenge. However, Treg depletion in vivo mediated rejection of tumour challenge; vaccination with exosomes inhibited this immune response, although heat-shock exosomes appeared least inhibitory, suggesting they may be more immunogenic than control or irradiated exosomes. Finally, the immunogenicity of exosomes might be increased by upregulating expression of TAA or immune adjuvants. NKG7 is a tetraspanin that is similar to CD63, a protein found enriched in exosomes. EGFP was successfully targeted into exosomes by linking it to the protein NKG7. EGFP-expressing exosomes induced strong maturation of DC, indicating increased immunogenicity. Therefore, the immunogenicity of exosomes can be enhanced by using a targeting mechanism to increase their expression of immune adjuvants.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:578643 |
| Date | January 2007 |
| Creators | Ilett, Elizabeth Jane |
| Publisher | University of Leeds |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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