Alvacc: A human in vitro alveolus model to study aerogenic TB vaccine binding and translocation
Led by Dr Diane Lee (University of Surrey, UK), with Prof Mark Chambers (University of Surrey, UK) and Prof Rajko Reljic (SGUL, UK)
Project Aims
Delivering tuberculosis (TB) vaccines directly to the lung by aerosol can result in efficient and better protection by directly targeting immune cells resident in the lung, as shown for BCG vaccine in non-human primates. However, optimising these processes is not trivial. Recent work has shown that protection conferred by BCG can be enhanced by subsequently administering another vaccine to the lung consisting of a harmless bacterial spore coated with proteins found in TB, called "Spore-FP1". The efficacy of Spore-FP1 stems from inclusion of the TB protein, HBHA. As the lung has evolved to normally exclude particles like Spore-FP1, HBHA is believed to improve attachment of the vaccine to the cells lining the alveoli of the lung followed by efficient transport of the vaccine across the alveolar barrier. In turn, this gives Spore-FP1 access to immune cells underlying the alveoli. However, direct experimental evidence for this is lacking. If we could show this was the case, better understanding these processes would allow us to design vaccines for delivery to the lung that are both safe and highly efficient at generating an immune response locally in the lung itself. However, these transient processes are difficult to study in animal models. We address this challenge in this project by studying how Spore-FP1 and BCG vaccines interact with and cross a human laboratory tissue model of the alveolus recently developed by the applicant. The use of fresh human cells from an ethically approved and verified source avoids contradictory results previously encountered using cell lines and any anomalies which might arise from using cells from other species. As well as understanding how HBHA contributes to the performance of Spore-FP1, our new lung model will be available to the scientific community for further studies of vaccine binding and translocation, helping reduce the number of animal experiments.
Project Outcomes
Aerosolised vaccine delivery is an option for inducing localised immune memory and rapid effector responses to respiratory pathogens. Enhanced efficacy of aerosol BCG against M. tuberculosis (Mtb) challenge in NHP has been reported but why aerosol vaccination is so effective is not fully understood. We used a human alveolus in vitro model to study TB vaccine interaction with the airway epithelial-endothelial interface, mimicking an important aspect of respiratory vaccine delivery.
Spore-FP1 (spores of Bacillis subtilis coated with TB antigens) were recently developed at SGUL as a novel TB vaccine. Intranasal delivery of Spore-FP1 significantly enhances the protection induced by BCG vaccination in a mouse aerosol Mtb infection model. It is hypothesised that inclusion of the Mtb Heparin-Binding Haemagglutinin Adhesin (HBHA) antigen allows the vaccine to mimic Mtb’s capacity to bind to lung epithelial cells and gain access to submucosal tissues inducing protective immunity in the lungs. Our aim was to assess the suitability of the HuAlv model to test this hypothesis by studying the uptake, translocation and immunostimulation of Spore-FP1 vaccine, gaining insights into how this, and potentially other aerosolised vaccine candidates may reach submucosal immune cells.
By studying the translocation of Spore-FP1 and comparing with both individual components and referencing to MTb, our aim was to generate proof of principle data that could be used as a foundation to further key studies to improve our understanding of the protective immunity arising through the interaction of aerosolised vaccines with the mucosa. The hope is that improved understanding would expedite the development of vaccines administered to the lower respiratory tract, where transient interactions are difficult to follow in vivo. Using the HuAlv model in this way could lead to rational design of aerosol vaccines with 3Rs benefit.
The HuAlv bilayer model was successfully transferred from the UOS to SGUL, with integral tight junctions demonstrated by both Dextran Blue (DB2000) transport resistance and transepithelial electrical resistance (TEER). Although we were able to demonstrate that Spore FP-1 was associated with AECs, we were not able to quantify Spore-FP1 in the basolateral supernatant of bilayer cultures grown on suspended permeable inserts nor in the basolateral supernatant of AEC monolayers. It is not clear whether this is due to a genuine lack of translocation/transcytosis of Spore-FP1, or insufficient sensitivity of the assays we developed to quantify spores. We were able to demonstrate translocation of live Mtb across bilayer cultures, suggesting live mycobacteria at least do cross the epithelial-endothelial bilayer of the model.
Despite the lack of apparent translocation of the Spore-FP1 vaccine, we did find it caused a time-dependent increase in important activatory and chemoattractant cytokines following administration, which may suggest that immunostimulation, not translocation, may be key to the efficacy of this particular vaccine.
Finally, as studies are exploring the role of non-human primate (NHP) cell-free lung lavage as a mucosal adjuvant of inhaled vaccines, we added filtered lavage to the bilayer model and found it had no detrimental effect upon bilayer integrity (as determined by TEER).
