Aspergillus and chronic lung allograft dysfunction: an immunophenotyping approach to optimise therapies

Lung transplantation is an established treatment for patients with end stage lung disease. Survival following lung transplantation is the worst out of all the solid organ transplant groups with a median survival of only 6.5 years. The two most common causes of death after the first-year post transplant are infection and chronic lung allograft dysfunction (CLAD). Aspergillus fumigatus is a mould that can cause a wide spectrum of disease from asymptomatic colonisation to life-threatening infection. Invasive aspergillosis (IA) the most severe form of infection, predominantly affects immunocompromised patients and is a particular problem after lung transplant.
Colonisation with Aspergillus has been linked to the development of CLAD in a number of studies, however a recent international multi-center study with large numbers of lung transplant recipients refuted this. We recently studied Aspergillus in our patients and contrasted colonisation and infection in patients with and without CLAD. We discovered that Aspergillus colonisation was more frequent in patients without CLAD (32.7% vs 13.7%), whereas Aspergillus infection was more frequent in patients with CLAD (18.9% vs 11.3%) and was associated with high mortality. This supports the hypothesis that colonisation may not be linked to CLAD.
Despite no clear evidence that tells us when Aspergillus colonisation is causing inflammation that may lead to the development of CLAD, clinicians treat Aspergillus aggressively due to the potential risk of CLAD. Treatment for Aspergillus colonisation and infection requires long term therapy with tri-azole drugs. These are associated with significant side effects and long term toxicity. For example, pancreatitis, hepatotoxicity and squamous cell carcinomas. Additionally, there is increasing evidence that long term use is selecting out strains of Aspergillus that are resistant to tri-azole drugs. It is important to establish whether colonisation does lead to CLAD in order to avoid unnecessary side effects of tri-azole drugs.
We propose a cross-sectional study examining blood and bronchoalveolar lavage fluid (lung washings) from patients who have had a lung transplant. Patients will be grouped according to whether they have/do not have CLAD, and according to their Aspergillus status (no aspergillus/colonised/infected). We will undertake multi-parameter immune profiling to identify specific immunological and cytokine signatures associated with Aspergillus in combination with comprehensive clinical phenotype and physiological data to establish whether these biological signatures could act as markers for severity or offer prognostic value in identification of patients likely to progress to more severe disease/CLAD. We will run Aspergillus T-cell specific mechanistic studies in parallel which will complement the multiparameter immune profiling and provide further insights into the potential mechanisms that Aspergillosis may be associated with CLAD following lung transplant.
Finally, we will start to develop a lung transplant biobank framework. The initial phase will involve the two largest lung transplant centres (Harefield & Newcastle). Data generated through this MRC-CARP award will be used for a further grant application and will form the basis for ongoing study to further understand how specific risk factors for CLAD (Aspergillosis, Gastroesophageal reflux, bacterial & viral infection, episodes of acute rejection and the development of donor specific antibodies) may contribute to the final common pathway of graft destruction/CLAD.
CLAD shares similar features with other fibrotic and alveolar degradation processes seen in chronic respiratory diseases, such as asthma, chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis, and thus provides a unique window into pathobiology common to other chronic respiratory diseases and may provide valuable insights into novel strategies with wider therapeutic utility.

Technical Summary
Colonisation or infection with the major mould pathogen Aspergillus fumigatus occurs in up to 50% of lung transplant recipients, leading to accelerated chronic lung allograft dysfunction (CLAD) and increased mortality. Better understanding is required of when the presence of Aspergillus in the airway is promoting pulmonary inflammation, and how this relates to the development of CLAD. This is clinically important, because it is currently unclear whether patients with co-existing airway Aspergillus and CLAD require either antifungal therapy, immunosuppression, or both. In particular, as CLAD may be multifactorial, better assays to identify when this is driven by Aspergillus are needed.
In order to better understand the interplay between Aspergillus, pulmonary inflammation and CLAD we will undertake a cross-sectional comparative study of lung transplant patients developing CLAD in comparison to control lung transplant recipients without CLAD, comparing subgroups with pulmonary aspergillosis, Aspergillus colonisation and no Aspergillus. We will adopt the recently developed and validated multi-parameter immunophenotyping approach developed by Prof. Hayday in order to identify immunological signatures associated with Aspergillus-driven pulmonary inflammation. To underpin these immunophenotyping studies we will also undertake Aspergillus antigen-specific T cell immunophenotyping, in order to identify where a particular inflammatory pathway is being driven by Aspergillus. Together, these studies will allow us to identify which patients with Aspergillus are developing lung inflammation, and what particular immune signatures are particularly associated with CLAD. This will allow better identification of when Aspergillus is driving CLAD, allowing us to target those individuals who clearly need to be treated with antifungals. It will also provide greater insight into what immunogical pathways may require immunotherapeutic targeting in this context.