Concerted Activity of IgG1 Antibodies and IL-4/IL-25-Dependent Effector Cells Trap Helminth Larvae in the Tissues following Vaccination with Defined Secreted Antigens, Providing Sterile Immunity to Challenge Infection

Over 25% of the world's population are infected with helminth parasites, the majority of which colonise the gastrointestinal tract. However, no vaccine is yet available for human use, and mechanisms of protective immunity remain unclear. In the mouse model of Heligmosomoides polygyrus infection, vaccination with excretory-secretory (HES) antigens from adult parasites elicits sterilising immunity. Notably, three purified HES antigens (VAL-1, -2 and -3) are sufficient for effective vaccination. Protection is fully dependent upon specific IgG1 antibodies, but passive transfer confers only partial immunity to infection, indicating that cellular components are also required. Moreover, immune mice show greater cellular infiltration associated with trapping of larvae in the gut wall prior to their maturation. Intravital imaging of infected intestinal tissue revealed a four-fold increase in extravasation by LysM(+)GFP(+) myeloid cells in vaccinated mice, and the massing of these cells around immature larvae. Mice deficient in FcR gamma chain or C3 complement component remain fully immune, suggesting that in the presence of antibodies that directly neutralise parasite molecules, the myeloid compartment may attack larvae more quickly and effectively. Immunity to challenge infection was compromised in IL-4R alpha- and IL-25-deficient mice, despite levels of specific antibody comparable to immune wild-type controls, while deficiencies in basophils, eosinophils or mast cells or CCR2-dependent inflammatory monocytes did not diminish immunity. Finally, we identify a suite of previously uncharacterised heat-labile vaccine antigens with homologs in human and veterinary parasites that together promote full immunity. Taken together, these data indicate that vaccine-induced immunity to intestinal helminths involves IgG1 antibodies directed against secreted proteins acting in concert with IL-25-dependent Type 2 myeloid effector populations.

The microbiota provides colonisation resistance against intestinal helminths by regulating intestinal physiology

Luc Xavier Marie Lebon

Intestinal helminth infections are a global health problem affecting particularly people living in poor countries. Heavy worm burdens can lead to nutritional and cognitive deficiencies. Type-two immunity plays a crucial role in promoting tissue repair and limiting parasite burdens; however, our knowledge as to how these responses lead to protection is limited. Moreover, intestinal helminths have co-evolved with their mammalian hosts and are well known to possess potent immuno-modulatory capacities allowing them to establish chronic and repeated infections. As adult worms reside in the gut, we asked whether the presence of intestinal bacterial communities (commonly referred to as the microbiota) impacted on worm burdens or type-two immunity following infection. For this purpose, we infected germ-free (living in a sterile environment) or antibiotic-treated mice with the natural murine parasite Heligmosomoides polygyrus bakeri. As worms are normally hatched from faecal cultures and carry their own microbiota, we first developed a system to generate bacteriologically-sterile (axenic) infective larvae. In germ-free and antibiotic-treated mice, the number of adult worms and their egg production were both increased, despite the presence of a slightly increased helminth-elicited type-two immune response, as shown by cytokine and antibody responses. The increased worm fitness in the absence of microbiota was already detected early following infection and was associated with a more anterior distribution of larvae along the intestinal tract. Thus, we hypothesised that the microbiota may impact on parasite burdens through observed alterations of intestinal physiology, such as increased bile acid production and deregulated smooth muscle cell contractility. To test the importance of these factors, we performed experiments in which we treated mice with an excess of bile acids or altered intestinal transit through drug treatment. Both interventions lead to changes in worm infectivity or location, mirroring different observations generated in our germ-free and antibiotic-treated mice. In summary, our data indicate that a complex microbiota affords 'colonisation resistance' against intestinal helminths, in a manner that is independent of the immune response but dependent on changes to intestinal physiology.

EPFL2017

Protein engineering approaches for direct antigen targeting in CD8+ T cell inducing vaccines

Vasiliki Panagiotou

Effective vaccine design and public vaccination programs have led to the eradication of several diseases and protected millions of people from deadly infections. However, intracellular infections remain a challenge and their eradication requires the activation of cytolytic mechanisms. CD8+ T cells promote effector mechanisms for infected cell killing subsequent to their priming by antigen-presenting cells. Their activation occurs upon detection of intracellular antigen in a pathway that many current subunit vaccine technologies are trying to approach by cross-presentation. Advances in CD8+ T cell-inducing subunit vaccines include direct targeting of antigens to cross-presenting cells and are studied in this thesis. Antigen delivery to the appropriate cell type for cross-presentation to CD8+ T cells can occur with fusion antibodies. Our laboratory has previously demonstrated that antigens bearing an erythrocyte-binding antibody domain (TER119) target the liver. We used this technology to fuse TER119 and ovalbumin (OVA) antigen with the BBOX domain of the high mobility group box 1 (HMGB1) danger signal and co-administer it with CpG-B adjuvant in a hepatic vaccine context. In liver vaccines, activation of local cytotoxic responses represents an important challenge. Erythrocyte-mediated OVA delivery with adjuvants resulted in cytotoxic CD8+ T lymphocyte (CTL) activation with memory formation in the liver and protection was provided after infection with Listeria monocytogenes. Another method for antigen targeting is through a cross-presenting dendritic cell (DC) subset, called CD8+ DCs. Recent studies in mice showed successful vaccination against OVA fused to XCL1 ligand, which binds CD8+ DCs. To attract CD8+ DCs to the site of injection for increased antigen uptake, we co-injected XCL1-OVA with a fusion protein consisting of XCL1 and the extracellular matrix (ECM) binding domain of placenta growth factor-2 (PlGF-2123-144), called XCL1-PlGF. XCL1-PlGF binds extracellular matrix at the injection site and attracts CD8+ DCs locally. In the context of prophylactic and therapeutic vaccination, the combination of two fusion proteins (XCL1-OVA and XCL1-PlGF) enhanced cytotoxicity and prolonged survival against B16-OVA melanoma. Finally, antigen transport via biomaterial delivery platforms was evaluated as an alternative method to deliver antigens to immunologically relevant sites. Our laboratory and others have shown that nanoscale carriers promote CTL responses in vaccination. Here we present a cationic micelle vaccination platform in which OVA and adjuvant (CpG-B and/or MPLA) loading is mediated by non-covalent molecular encapsulation and adsorption. Following a prime and double boost vaccination, CTL responses were raised in the spleen and lymph nodes of vaccinated mice, along with antigen-specific antibody production. These findings highlight the advantages of the micelle carrier platform in vaccine applications. Overall, this thesis presents novel techniques to target antigens to the appropriate cell compartments, facilitating CTL activation in vaccination either by direct targeting of antigen fused to antibodies and chemokines, or antigen delivery with polymeric vehicles. These platforms for optimized delivery seek to improve current approaches and impact the design of new strategies in CD8+ T cell inductive vaccination.

EPFL2016

Enteric helminth-induced type I interferon signaling protects against pulmonary virus infection through interaction with the microbiota

Eva Sabrina Gollwitzer, Nicola Harris, Benjamin John Marsland

BACKGROUND: Helminth parasites have been reported to have beneficial immunomodulatory effects in patients with allergic and autoimmune conditions and detrimental consequences in patients with tuberculosis and some viral infections. Their role in coinfection with respiratory viruses is not clear. OBJECTIVE: Here we investigated the effects of strictly enteric helminth infection with Heligmosomoides polygyrus on respiratory syncytial virus (RSV) infection in a mouse model. METHODS: A murine helminth/RSV coinfection model was developed. Mice were infected by means of oral gavage with 200 stage 3 H polygyrus larvae. Ten days later, mice were infected intranasally with either RSV or UV-inactivated RSV. RESULTS: H polygyrus-infected mice showed significantly less disease and pulmonary inflammation after RSV infection associated with reduced viral load. Adaptive immune responses, including TH2 responses, were not essential because protection against RSV was maintained in Rag1-/- and Il4rα-/- mice. Importantly, H polygyrus infection upregulated expression of type I interferons and interferon-stimulated genes in both the duodenum and lung, and its protective effects were lost in both Ifnar1-/- and germ-free mice, revealing essential roles for type I interferon signaling and microbiota in H polygyrus-induced protection against RSV. CONCLUSION: These data demonstrate that a strictly enteric helminth infection can have remote protective antiviral effects in the lung through induction of a microbiota-dependent type I interferon response.