Doherty Institute - Theses
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ItemUsing dendritic cell receptors to enhance immunity( 2017)Dendritic cells (DCs) are the most potent initiators of immune responses, being highly specialised for the uptake and presentation of antigens (Ag) to activate T cells. Their priming potential can be harnessed to generate stronger immune responses by targeting Ag to DCs via monoclonal antibodies (mAbs) specific for DC-expressed surface receptors. This thesis builds upon the concept of targeting DCs in two main ways: firstly, by investigating a novel method of targeting adjuvant to DCs, and secondly, by investigating how DC-targeting constructs can be used to prime and boost responses. It was considered whether not only Ag, but also adjuvants could be targeted to DCs to improve their efficacy. A recent finding that the DC receptor DEC-205 can bind to and mediate the immunostimulatory effects of CpG oligonucleotide (ODN) adjuvants led to the hypothesis that CpG ODNs could be targeted to DCs via DEC-205 in order to enhance their potency. The interaction between DEC-205 and CpG ODNs was further characterised to determine the molecular properties of ODNs required for binding. This information was then used to enhance the DEC-205 binding capacity of a particular CpG ODN that normally only weakly binds DEC-205. Enhanced DEC-205 binding was found to significantly improve the stimulatory capacity of this ODN, demonstrating that targeting adjuvant to DCs could be a viable method to improve adjuvant potency. Another receptor, CD14, has also been reported to bind CpG ODNs, so the potential for CD14 to act in synergy with DEC-205 was investigated. However, CD14 was not observed to mediate the uptake or stimulatory effects of CpG ODNs. The identification of natural ligands of DEC-205 is critical for understanding its physiological function. Although ODNs are synthetic molecules, their binding to DEC-205 may signify that DEC-205 is capable of binding other types of DNA that structurally resemble ODNs. A panel of biological DNA samples was screened for DEC-205 binding. While none of the DNA samples were observed to bind DEC-205, some DNA samples were found to bind another receptor, RAGE, suggesting a role for RAGE as a detector of both pathogenic and self-DNA. Most vaccines must be administered more than once, or “boosted”, to achieve optimal efficacy, and DC-targeted vaccines should be no exception. However, our data suggested that simply administering the same DC-targeting construct twice does not effectively boost the response. This was due to interference from the primary antibody response, which can cross-react with and neutralise a subsequently administered boosting construct. To overcome this issue, the efficacy of various heterologous prime-boost strategies designed to reduce the reactivity of the primary response against the boosting construct was assessed. Ultimately, a combination of anti-Clec9A and anti-XCR1 targeting constructs was found to induce the least cross-reactivity and strongest response after boosting. These findings contribute to the development of better adjuvants and immunisation strategies that optimise the efficacy of DC-targeted vaccines. More broadly, they also highlight the value of understanding the underlying biological mechanisms that drive immune responses, which can then be applied to the rational design of more effective vaccines.