Roles for Treg Expansion and HMGB1 Signaling through the TLR1-2-6 Axis in Determining the Magnitude of the Antigen-Specific Immune Response to MVA85A

et al. (2013) Roles for Treg Expansion and HMGB1 Signaling through the TLR1-2-6 Axis in Determining the Magnitude of the Antigen-Specific Immune Response to MVA85A. PLoS ONE 8(7): e67922. doi:10.1371/journal.pone.0067922 Roles for Treg Expansion and HMGB1 Signaling through the TLR1-2-6 Axis in Determining the Magnitude of the Antigen-Specific Immune Response to MVA85A Magali Matsumiya 0 1 Elena Stylianou 0 1 Kristin Griffiths 0 1 Zoe Lang 0 1 Joel Meyer 0 1 Stephanie A. Harris 0 1 Rosalind Rowland 0 1 Angela M. Minassian 0 1 Ansar A. Pathan 0 1 Helen Fletcher 0 1 Helen McShane 0 1 Riccardo Manganelli, University of Padova, Medical School, Italy 0 Current Address: Centre for Infection , Immunity and Disease Mechanisms , School of Health Sciences and Social Care, Brunel University , Middlesex , United Kingdom 1 The Jenner Institute, University of Oxford , Oxford , United Kingdom A better understanding of the relationships between vaccine, immunogenicity and protection from disease would greatly facilitate vaccine development. Modified vaccinia virus Ankara expressing antigen 85A (MVA85A) is a novel tuberculosis vaccine candidate designed to enhance responses induced by BCG. Antigen-specific interferon- (IFN) production is greatly enhanced by MVA85A, however the variability between healthy individuals is extensive. In this study we have sought to characterize the early changes in gene expression in humans following vaccination with MVA85A and relate these to long-term immunogenicity. Two days post-vaccination, MVA85A induces a strong interferon and inflammatory response. Separating volunteers into high and low responders on the basis of T cell responses to 85A peptides measured during the trial, an expansion of circulating CD4+ CD25+ Foxp3+ cells is seen in low but not high responders. Additionally, high levels of Toll-like Receptor (TLR) 1 on day of vaccination are associated with an increased response to antigen 85A. In a classification model, combined expression levels of TLR1, TICAM2 and CD14 on day of vaccination and CTLA4 and IL2R two days post-vaccination can classify high and low responders with over 80% accuracy. Furthermore, administering MVA85A in mice with anti-TLR2 antibodies may abrogate high responses, and neutralising antibodies to TLRs 1, 2 or 6 or HMGB1 decrease CXCL2 production during in vitro stimulation with MVA85A. HMGB1 is released into the supernatant following atimulation with MVA85A and we propose this signal may be the trigger activating the TLR pathway. This study suggests an important role for an endogenous ligand in innate sensing of MVA and demonstrates the importance of pattern recognition receptors and regulatory T cell responses in determining the magnitude of the antigen specific immune response to vaccination with MVA85A in humans. - Tuberculosis (TB) remains a major global health issue, with an estimated 8.7 million cases and 1.4 million deaths in 2011 [1]. BCG, the only licensed vaccine against TB, shows only partial, variable efficacy against pulmonary TB [24]. Twelve candidate vaccines are currently in clinical trials [5] and results of the first efficacy trial of a new vaccine against Mycobacterium tuberculosis (M.tb), in which Modified Vaccinia virus Ankara expressing antigen 85A (MVA85A) was given to BCG-vaccinated South African infants, have recently been reported [6]. Although MVA85A did not confer improved protection to TB compared to BCG alone in this setting, further analysis of samples collected will provide a valuable opportunity to investigate the immune basis of protection against TB. Efforts to produce T cell inducing vaccines against diseases such as TB, HIV and malaria, have made use of viral vectors as antigen delivery systems to enhance the immune response to the antigen of interest. Vaccine candidates are selected on the basis of safety, efficacy in preclinical animal disease models and the ability of the vaccine to induce the secretion of interferon- (IFN-) by antigen-specific CD4+ and CD8+ T-cells [79]. The secretion of cytokines by stimulated cells, particularly IFN-, remains the primary gauge of vaccineinduced adaptive immune responses in both animals and humans [10]. IFN- is essential for protection against TB and, although it is not a correlate of protection, it correlates well with other Th1 functions and is the cytokine that gives the most robust measure of response to vaccination [11,12]. Therefore it is of paramount importance to understand the mechanisms by which different vaccines induce this immune response, as well as understanding the basis of a protective immune response to the diseases for which the vaccines are being tested. Attenuated poxviruses have been widely used in vaccine development; MVA for example has been tested in clinical trials against malaria, TB, HIV, influenza and a variety of cancers [1318]. There is now interest in determining how MVA initiates the immune response. The induction of apoptosis by MVA is well documented [1921] and recent work has shown that deletion of the anti-apoptotic gene F1L from the genome of MVA leads to increased immune responses in mice, suggesting apoptosis plays an important role in the induction of immunity to MVA [22]. Additionally, Delaloye et al. have shown the importance of Toll-Like Receptors 2 and 6 (TLR2-6), melanoma differentiation-associated gene 5 protein (MDA-5) and the NALP3 inflammasome in immune sensing of MVA [23], although no ligand for this association has yet been characterized. HMGB1, a nuclear protein released during cell death acts as a danger associated molecular pattern (DAMP) and can bind to several TLR receptors. It has been implicated in the immune response to several viruses and has been proposed as the elusive TLR2 ligand during MVA and vaccinia infection. The field of vaccinology has seen, through seminal work using the yellow fever vaccine YF-17D as a model, that systems biology can be a powerful tool in identifying the mechanisms by which an immune response to vaccination develops [24,25]. In this study we describe the innate immune response to MVA85A in BCG-vaccinated volunteers in the UK and use the data to investigate the links between early changes in gene expression and IFN- ELISpot responses measured at the time of the trial. We are not able to postulate on the relevance of this to protection, as correlates of protection remain an elusive goal. However, the ex-vivo IFN- ELISpot is a good measure of vaccine take and correlates with many aspects of Th1 type immunity. It has been used in multiple studies across different diseases to assess vaccine immunogenicity, although it is not a correlate of protection in any of these diseases. In the case of tuberculosis, IFN- is also known to be necessary, though insufficient, for protection. Understanding the mechanisms underlying the immune response to vaccination is an important goal that complements but is separate from studies examining the basis of protective immu (...truncated)