Defining the role of CAR in lung homeostasis and response to inflammation

Over 5 million people in the UK suffer from asthma or forms of allergic inflammatory lung diseases that affect their ability to breathe properly. These disorders can have a significant impact on quality of life and in the most severe cases can be fatal. Some people with asthma and inflammatory lung diseases can be treated using inhaled steroids, which can help to expand the airways and alleviate breathing problems. However, long-term cures for these disorders or treatments for those who do not respond to steroids are still not available. Moreover, we still do not understand many of the factors that contribute to inflammation in the lungs following responses to allergic triggers. Understanding more about these factors is very important in enabling new drugs to be developed that can help treat these inflammatory lung disorders. We have identified a protein called CAR that is present in the cells that line the lung airways (called epithelial cells). In healthy lungs, CAR is important in helping these cells to form a barrier to protect the lung tissue. However, we have shown that CAR changes it's function when lungs are exposed to factors that drive inflammation, such as house dust mites which can trigger asthma in over 50% of patients. We have shown that CAR actually promotes the recruitment of immune cells to lungs after exposure to house dust mite and if we remove or block CAR in these cells, the inflammation is reduced. Reducing the amount of inflammation is very important as it stops the onset of the clinical features seen in severe asthma patients. CAR is therefore a very good candidate factor for further exploration as a potential key factor controlling inflammation in the lung. Our current project will investigate the way that CAR controls the immune cells that are recruited to the lung after exposure to house dust mites. We will also test whether agents that block the function of CAR can prevent the recruitment of immune cells to the lung. Our studies will characterise a new potential therapeutic target for the treatment of asthma and other inflammatory lung diseases and will potentially provide new avenues for the development of drugs to combat these disorders in future.

Coxsackie virus and adenovirus receptor (CAR), is a member of the immunoglobulin (IgG) superfamily that functions as a cell-cell adhesion molecule through homophillic interaction in trans. CAR was originally identified as a primary receptor for Adenovirus cell binding. However, our work and that of others has demonstrated that CAR also has a key role in stabilising epithelial cell-cell adhesions as both an adherens and tight junction localized protein. As well as forming homodimers in trans, CAR has also been shown to bind in trans to other members of the JAM family proteins, including JAM-L that is expressed on the surface of leukocytes and this facilitates leukocyte transmigration and activation. Our recent studies have shown that CAR levels in epithelial cells control junctional protein dynamics and that this is in part regulated through phosphorylation of the CAR C-terminus at S290/T293 by PKC. Our unpublished data shows that conditional depletion of CAR from the epithelial cells in mouse lungs leads to a dramatic reduction in inflammatory responses to house dust mite. These findings place CAR as a key novel molecule in regulation of the interplay between epithelial and immune cells and thus of potential relevance in the development and progression of lung diseases such as asthma and COPD. Our present study will define how CAR elicits a pro-inflammatory effect in the lung and analyse the pathways and CAR-binding proteins that regulate this process. We will also define how CAR associates with itself across adjacent cell membranes, and determine whether alterations to the underlying matrix (as occurs in asthma) also leads to altered CAR function and disease exacerbation. Data arising from this study will provide new insight into CAR function in the epithelium and potentially enable the development of novel therapeutic approaches in future.

The programme of research outlined here will impact on both academic and industrial sectors. Our data will be shared through talks and publications, as well as publicised widely through King's and MRC websites and social media. Our extensive academic networks, ranging from biophysics to clinical teams, will ensure the team on this project is exposed to a broad scope of potential feedback and new training opportunities. We will also learn from the discussions emerging from the IBIN TTL network (established by the PI), the goal of which seek to define new ways to study cell interactions and communication within tissues. We will also share our data with other basic and clinical research teams within the MRC and Asthma UK Centre to facilitate exchange of ideas and potentially extension of our hypotheses into other model systems as experiments progress.

Our experimental approaches and findings also hold clear potential for the identification of drugs that target CAR in inflammatory lung disease. We will discuss our findings as they emerge with current industrial collaborators and plan to progress any outcomes to pre-clinical stage with additional funding via MRC DPFS applications or similar. Our collaborator in Berlin also has an ongoing programme to develop novel inhibitors to CAR for heart disease. Through our collaboration, we will also be in a position to combine efforts to engage with companies with an interest in pursuing CAR as a target for multiple indications. The importance of more advanced in vitro and ex-vivo assays to analyse cell function is increasingly recognised as being extremely important in drug discovery by the pharmaceutical industry and the methods and reagents we plan to develop in this study will place us at the forefront of emerging technologies leading to the design and validation of new therapies. Given our existing industrial collaborations, we are already very well placed to explore future commercial applications arising from the current proposal. We believe our experience and expertise gained from the current proposal will also place us in an excellent position to engage in other similar imaging development-based studies with other industrial partners in future.