Fungal suppression by human gut Vdelta2 T-cells in mucosal immunity and inflammation

Background: Recent rises in many human chronic diseases have been linked with changes in gut bacteria, but we know very little about how human health is influenced by the fungus that lives in our intestine. Children with inflammatory bowel disease (IBD) have increased amounts of fungus in the gut, and patients with genetic problems in anti-fungal immunity suffer from severe intestinal inflammation. Recent work by Dr. Neil McCarthy found that human gut tissue can be protected against fungus by a cell type called 'Vdelta2' which is common in humans but missing in mice. Vdelta2 cells are activated by an immune system protein called BTN which detects a bacterial chemical called HMB-PP. We think this could be an important new type of anti-fungal immune response that may not be working properly in patients with IBD.

Hypothesis: Vdelta2 cells prevent fungus growth to protect against human gut inflammation.

Aim 1. Identify how Vdelta2 cells restrict growth of Candida fungus. We will add the fungus Candida to human intestinal tissue and investigate how Vdelta2 cells stop fungal growth. Lots of scientific studies have been done on Candida, so this is a good fungus to use when setting-up our tests. We will add chemicals to block different types of Vdelta2 cell response and observe the effect on Candida growth. These experiments will tell us how Vdelta2 cells stop Candida so that we can later test how well this works with real gut fungus.

Aim 2. Determine how Vdelta2 cells restrict growth of real human gut fungi. We will use 'dirty' gut tissue biopsies to test whether growth of real intestinal fungus can be blocked by Vdelta2 cells. Fungus experts will help us to identify the species that we collect from these experiments. This will tell us how Vdelta2 cells stop the growth of different types of fungus in the healthy human gut, so that we can next test if this immune response works differently in IBD.

Aim 3. Test if Vdelta2 cells can restrict fungal growth in inflamed human gut. We will test whether Vdelta2 cells can control fungus in gut tissue from IBD patients, and whether inflammation changes how the fungus grows. We will also identify the fungal species to see if these differ between healthy and inflamed gut. Other experiments will test whether fungus grows more easily in tissue from IBD patients being treated with drugs that block Vdelta2 cell responses. This will give us important information about how Vdelta2 control of fungi works differently in patients with gut inflammation and how this might be changed by IBD therapies. We will then be in a good position to look for other factors that change the ability of Vdelta2 cells to limit fungal growth in the gut.

Aim 4. Examine whether fungi can escape Vdelta2 responses by forming 'biofilms'. Gut fungi can team-up with certain types of bacteria to form thick organic layers called 'biofilms' that are difficult for the immune system to break down. We will therefore test if fungus can 'escape' Vdelta2 cell responses by forming biofilms with gut bacteria. These experiments will also be used to extend our research into other areas where fungal biofilms often cause major problems in human patients (medical implants, tissue transplants, and on plastic tubing used in hospitals).

Aim 5. Test if Vdelta2 cells can prevent fungal growth in BTN-deficient patients. Vdelta2 cells are activated by the immune system protein BTN, so we will test whether Vdelta2 cells can still control fungal growth in blood from volunteers with genetic changes in BTN proteins. These experiments will generate important new information about how BTN proteins work in human patients so that we design better treatments in future.

New treatments: Several drugs have already been produced that can strongly activate Vdelta2 T-cells in human tissues, so it is highly likely that Vdelta2 cell-targeted therapies could also be developed to increase patient protection against fungi and reduce inflammation in IBD.

Background: Gut dysbiosis is now recognized as a major driver of the rapid increase in many chronic diseases over the last century, but the influence of commensal fungi on human tissue inflammation is largely unknown. The applicant, Dr Neil McCarthy recently observed that human gut tissue can be protected against fungal outgrowth by using the bacterial metabolite HMB-PP to activate Vdelta2 T-cells (which are absent in rodents). Since host responses to gut fungi are strongly implicated in inflammatory bowel disease (IBD), we will determine whether this novel mechanism of anti-fungal immunity in human mucosal tissue is disrupted in IBD.

Hypothesis: Bacterial activation of Vdelta2 T-cells restricts fungus growth and protects against human gut inflammation.

Method: Human blood and healthy/inflamed gut tissue will be treated with antibiotics to ablate bacteria and promote fungal growth (either added Candida or endogenous fungi) with/without Vdelta2 T-cell activation by HMB-PP. Key mediators of fungal suppression will be identified using blocking antibodies, recombinant cytokines, and therapeutic reagents that modify Vdelta2 T-cell function in vivo. We will also assess the fungal-protective effects of biofilm formation and genetic deficiencies in the BTN3A proteins that regulate Vdelta2 T-cell responses (patients identified by East London Genes & Health project). Fungal isolates will be identified by MALDI-TOF.

Outputs: We will determine how Vdelta2 T-cells suppress intestinal fungi, the range of fungal species restricted by this mechanism, and how gut inflammation and IBD therapies interact with this process. Assays in patients with BTN3A mutations will generate crucial new data on how these proteins regulate Vdelta2 T-cell function and inform the design of novel therapies. Several Vdelta2-targeted drugs have already been developed that potently activate these cells in vivo and could potentially be used to increase patient protection against fungi and treat IBD.

Invasive fungal diseases kill more people every year than either tuberculosis or malaria, but there are currently no vaccines against fungi and the available therapies are highly toxic to human patients. Drug-resistant species such as Candida auris are now beginning to emerge in UK hospitals and have proven highly transmissible, lead to significantly prolonged hospital stay, and confer up to 70% risk of mortality. This research is therefore closely aligned with MRC's leadership of a cross-council initiative to tackle anti-microbial resistance by exploring how host immunity can be boosted to fight infection. Who will benefit from this research?

Patients with inflammatory bowel disease (IBD);
Gut fungi have recently been identified as a major component of IBD pathology and been implicated in several failed antibody drug trials. This research will shed new light on the immune mechanisms that protect human gut tissue against fungal infection and inflammation, and will inform the design of more effective therapeutic strategies in future. Synthetic phosphoantigens have already been developed that potently activate primate Vdelta2+ T-cells in vivo, hence it is highly likely that this project will assist pharma in developing new methods of boosting human anti-fungal immunity in patients with a variety of disorders including IBD.

Patients with epithelial barrier disorders;
Blood Vdelta2 T-cells are readily recruited to the human gut but also populate other epithelial barrier sites at high risk of fungal infection. This research will therefore be directly relevant for a variety of different patient types who suffer severe/recurrent fungal pathology due to epithelial barrier disruption (including lung, skin, nasal, genital and oral pathologies). Defining novel Vdelta2-dependent mechanisms of anti-fungal immunity in human gut, and any new drug treatments stemming from this work, will therefore impact on a wide range of patient groups beyond the immediate field of study in IBD.

Elderly and immunosuppressed patients;
Rates of severe fungal infection are set to increase rapidly in years ahead due to declining patient resistance. This work will inform efforts to identify methods of enhancing or preserving host immunity to fungal pathogens, which will be essential to offset the effects of population ageing and widening use of immunosuppressive drugs and antibiotics. Candida albicans is already the fourth most common cause of nosocomial bloodstream infections and confers >50% risk of mortality among elderly in-patients. Novel treatments and diagnostics arising from this research could thus lead to reduced rates of invasive fungemia and better outcomes in high-risk groups (patients with cancer or undergoing invasive surgery, HIV-infected individuals etc).

Clinicians and healthcare professionals;
Biofilm formation on catheters and central lines incurs significant patient morbidity and high rates of technique failure across a wide range of clinical settings. Fungal line infections also impose a major financial burden on the NHS by extending hospital stay as well as conferring elevated risk of sepsis and patient mortality. Fungal dissemination from infected lines to remote organs can also lead to severe pathology frequently affecting the eyes, kidneys and heart. Novel strategies for disrupting fungal biofilms will therefore improve clinical care for the increasing number of susceptible patients receiving immunosuppression, medical implants and various tissue/organ transplants etc.

Overall: The MRC has already highlighted the need to tackle the emerging public health threat posed by fungal infections and associated inflammatory disorders in the years ahead. Defining novel mechanisms of anti-fungal immunity in human tissues that can be targeted for therapy will be key to these efforts, ultimately leading to improved patient outcomes, reduced length of hospital stay, and major societal and economic benefits across the UK.