The molecular basis of a novel mechanism of antigen presentation: phosphoantigen-dependent activation of Vgamma9/Vdelta2 T cells by BTN3A molecules.

A particular sub-set of lymphocytes, gammadelta T cells, are found in the circulation but their precise function and how they are controlled has been obscure. A key insight has been made linking activation of gammadelta T cells to a human protein called BTN3A1. The importance of circulating gammadelta T cells is highlighted by the observation that their frequency in the blood can increase from a few percent in most people to up to 30-40% of total lymphocytes in response to infection and cancer. Their function is to home in and kill infected and cancerous cells. Most of the time, cancer cells and those cells in the body which are infected, by for example pathogenic bacteria, can be identified and destroyed by this surveillance mechanism. However, life-threatening cancers and harmful infections do occur. We know that part of the reason for this is a result of the ability of diseased cells to escape detection by the immune system. The realization that cancers and pathogens can modulate the immune response to favour their own growth and replication is a major theme in immunological research. We believe that understanding in detail the mechanism of gammadelta T cell activation and the role played by BTN3A proteins, will help us to intervene in this dynamic between the immune system and harmful diseases.
Another aspect of our study is the realization that inappropriate regulation of cells such as gammadelta T cells can lead to another class of diseases, collectively termed autoimmunity. In this case, the potent killing ability of the immune system is directed against cells of the body in the absence of any overt disease from external sources, such as infection. The immune system appears to be responding to confusing signals and attacks seemingly healthy cells inappropriately. Again, a clear understanding of the mechanism of gammadelta T cell activation may help us to intervene positively and perhaps block inappropriate activation signals, thereby suppressing autoimmune disease progression. The central aim of our proposed work is directed towards these goals.

BTN3A proteins are members of the B7-like group of immunoglobulin receptors, which interact with and modulate the function of T cells. Recent data show that BTN3A1 influences the activation of human gammadelta (gd) T cells by direct or indirect presentation of self/non-self phosphoantigens (pAg), a mechanism relevant to the immune-surveillance of microbial infections and of transformed and stressed tissues. In response to the microbial metabolite HMB-PP and the aminobisphosphonate drug zoledronate, we have evidence that multiple BTN3A molecules are required for gd T cell activation, consistent with recruitment into a multi-protein complex. We have used a number of techniques to identify BTN3A1 interacting molecules and propose to investigate their role in regulating gd T cells. Immunoprecipitation and protein mass spectroscopy were used to identify chaperone molecules potentially required for the formation of the CD277 antibody epitope, the critical determinant of gd T cell activation. Yeast-two hybrid assays were used to identify the cytoskeletal adaptor periplakin. The amino-terminal plakin domain of periplakin, composed of six spectrin repeat alpha-helices and SH3 domain, interacts with a membrane proximal di-leucine motif in the BTN3A1 cytoplasmic tail, a mechanism with similarity to E-cadherin/p120-catenin pathways. Our data suggest a role for BTN3A in epithelial cell homeostasis, linking the cytoplasmic sensing of pAg, metabolic intermediates of the mevalonate pathway, to BTN3A protein folding and glycosylation in the endoplasmic reticulum. We propose to exploit these unpublished observations. We will conduct binding assays, crystal trials and in vitro mutagenesis to explore the interaction between the B30.2 domain of BTN3A proteins and pAg. We will investigate the role of periplakin and related molecules and probe the oligomeric coupling of different BTN3A isoforms in relation to trafficking and T cell activation.

The research we describe could have implications for cancer and autoimmunity patients. Drugs and therapeutics may develop from the research, which could influence T cell recognition of tumours. Patients could benefit in the long-term from drugs and therapeutic strategies that emanate from the research.

Despite major advances in the understanding of disease patho-physiology, current therapeutic options in autoimmunity and cancer have not been universally successful. Long term prognosis is generally still very poor. These conditions affect a large number of people in the general population, either directly or indirectly, representing a substantial burden on health resources. Most cancer drugs for instance are expensive, relatively ineffective and often potentially toxic. Study of the immune response to diseases like cancer suggests that new therapeutic approaches, based particularly on modulation of T cell responses could be effective.
Within the past two years, significant progress has been made in unravelling the complex regulation of gammadelta T cells, a major subset of T lymphocytes. Gammadelta T cells regulate tissue homeostasis and are considered to be major instigators of immune recognition of disease. Understanding how gammadelta T cells are activated and how dysregulated activation contributes to disease is seen as a major focus. The central aim of our proposal is the identification and characterisation of molecular interactions which can be targeted in order to modulate gammadelta T cell function. Modulation of gammadelta T cells is regarded as a tractable approach to treat cancer and autoimmunity.
Exploitation. The development of new therapies for autoimmunity and cancer could be of considerable benefit to the UK heath budget. Cambridge Enterprise is a wholly owned subsidiary of the University of Cambridge that is responsible for commercialisation arrangements for University discoveries. The portfolio companies Horizon Discovery and BlueGnome were both winners in the International Trade category of the 2012 Queen's Award for Enterprise. We have contacted Cambridge Enterprise with a view to the potential commercialisation of our findings. At present, our findings are considered too preliminary for patent protection. However, we are in contact regularly with Cambridge Enterprise to discuss possible ways forward along these lines.
In addition, given the great success achieved by Cambridge institutions in the past, there is considerable interest in the broader community in the Cambridge area in the exploitation of scientific discoveries. Much of this interest is channeled through the Judge Business School, Cambridge, who run popular courses and events aimed at communicating basic science to potential investors and other experts in scientific exploitation. We regularly attend such networking events at the Judge Business School.
Collaboration. Key collaborations with Dr. Matthias Eberl (Cardiff), Dr. Leo James, (MRC-LMB Cambridge) and Prof. Ben Willcox (CRUK, Birmingham) have been established in order to carry out the proposed project. Interaction with these UK based scientists is a key feature of our proposal and is critical to its success. This will be managed through regular face-to-face meetings to discuss progress and strategy.
Dissemination/communication: Our findings will be disseminated to the wider scientific community via publishing in high impact, open access journals and through presentation in international scientific conferences. These functions will be performed by both the PI and the named postdoctoral researcher. We envisage the main bulk of our findings will be published in the final year of the project.
Who might benefit from this research: patients, heath care professionals. It could also have considerable economic benefit.
How might they benefit from the research: this research may lead to novel approaches for treatment in cancer therapy and autoimmunity, where therapeutic options are limited