Illuminating childhood respiratory infections: from viral diseases to vaccine delivery

Lead Research Organisation: Queen's University of Belfast
Department Name: Medicine Dentistry and Biomedical Sci

Abstract

Chickenpox and shingles, severe diarrhoea, cervical cancer, meningitis - a diverse assortment of very different diseases which affect young and old; rich and poor; people who live in developed and developing parts of the world. However, what unites this conglomeration of conditions is that in the last couple of years several major pharmaceutical companies have released new vaccines to fight these diseases. ?Vaccines are back?, but in reality they never went away, rather they ?kept their heads down? and delivered. Few products have done more to alleviate disease than vaccines. Moreover, vaccines continue to provide a steady source of income for the companies as other manufacturers who rely on selling low cost generic drugs find them harder to replicate.

The aim of this project is to understand how respiratory viruses cause disease by using and extending an animal model in which monkeys are infected with the virus that causes measles. However, not just any measles virus, rather we plan to use a genetically modified virus which glows bright green when it infects a cell. This allows us to light up disease in a hitherto impossible manner. When you think of measles in a child you probably think of red spots - when you think of measles in our monkey model ? think green! Green spots, green tonsils and green patches along the windpipe and in the lungs. As well as seeing these illuminated manifestations of disease by eye high powered microscopes can be used to identify individually infected cells with supreme levels of sensitivity. Since this animal model has been so beneficial for measles research we plan to use it to study how two other respiratory viruses establish infections by making them ?genetically green?.

It is essential to understand viral diseases comprehensively. This will help break down some of the barriers which get in the way of developing new vaccines. Already we have noticed that the tonsils are targeted in the very early of a measles virus infection. Therefore, with the help of industrial scientists, we will remove all of the water from vaccine and turn it into a solid which quickly melts on contact with saliva. We plan to take these vaccine-melts and place them directly onto the tonsils of the animal. Vaccinating in this way is much more natural than using a syringe to inject a respiratory virus into the arm. This may allow us to make better vaccines.

Technical Summary

We propose a multidisciplinary, integrated programme of work which aims to alleviate bottlenecks which hamper translation of fundamental knowledge into vaccine development. The work is based on our recently published measles/non-human primate pathogenesis model and we plan to identify common pathways of transmission for respiratory viruses. This information will be used to evaluate new routes of vaccine administration and the approach will be piloted by delivering lyophilised measles virus (MV) vaccine directly to the tonsils of the macaque.

The pleomorphic nature of paramyxovirus virions allows them to be engineered to carry extra genetic material which is stably maintained over many passages. Reverse genetics has allowed additional transcription units to be added before, between and after paramyxoviral genes. Open reading frames encoding reporter proteins such as enhanced green fluorescent protein (EGFP) have been used extensively and a significant amount of EGFP is expressed when cells are infected. Fluorescence allows infected cells to be visualised at unprecedented levels of sensitivity both in vitro and in vivo. As EGFP makes the entire cytoplasm of the cell fluorescent it is possible to visualise fine processes which interconnect cells. Therefore these viruses are perfectly suited for the study of virus transmission and pathogenesis.

The MV model will be extended to two other paramyxoviruses, human respiratory syncytial virus (hRSV) and human metapneumovirus (hMPV), which are associated with significant levels of morbidity and mortality. It has been difficult to develop vaccines against these viruses and to move forward it is essential to underpin future product development with a comprehensive understanding of the infections. Complementary whole animal, ex vivo and in vitro assays will be developed. Experimental observations will be cross-validated with pathological samples from naturally infected patients using a panel of human tissues from children infected with wild-type MV, a range of hMPV and hRSV clinical isolates from the UK and MV isolates from Africa. Currently circulating clinical isolates will be used to validate both in vivo and ex vivo assays.

This multidimensional model of disease provides an ideal platform to understand general pathogenesis of respiratory viruses. If we are able to elicit a solid immune response by tonsillar delivery tissue we will follow up this application with another in which we undertake challenge experiments. This is important as it has been recognised by MRC that, ?here are still many gaps in knowledge about how protective immune responses are generated and regulated in mucosal tissues?.

Publications

10 25 50
 
Description DELNI Strengthening the All-Ireland Research Base
Amount £1,550,000 (GBP)
Organisation Government of Northern Ireland 
Department Department for Employment and Learning Northern Ireland (DELNI)
Sector Public
Country United Kingdom
Start 01/2009 
End 12/2011
 
Description Foundation for NIH Grand Challenges in Global Health
Amount £1,100,000 (GBP)
Organisation Bill and Melinda Gates Foundation 
Sector Charity/Non Profit
Country United States
Start 04/2010 
End 06/2014
 
Description MRC Capacity Building Area Studentship
Amount £59,863 (GBP)
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 09/2010 
End 08/2013
 
Title Development of a multidimensional model of disease 
Description We have engineered a recombinant wild-type measles virus which was isolated in Sudan and represents a currently circulating clinically relevant virus. This virus has been tested in the macaque model and has proven to be highly pathogenic, proving our original hypothesis. This will allow us to leverage the techniques developed into others arenas of viral pathogenesis. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2009 
Provided To Others? Yes  
Impact Colleagues in the Center for Biologics Evaluation and Research (CBER), which is part of the Food and Drug Administration (FDA), are very interested in partering with us in developing an equivalent macaque-based, multidimensional model of disease for mumps. We are currently writing an NIH application to secure funding for these studies. 
 
Title Pathological samples from vaccinated and virus-infected macaques 
Description We have assembled a collection of formalin-fixed tissue blocks, frozen tissues and RNA samples from our monkey studies. This has been done this with a view to facilitating other scientists who may be interested in studying how a vaccine viruse behaves in a natural host compared to the wild-type clinical infection. 
Type Of Material Biological samples 
Year Produced 2009 
Provided To Others? Yes  
Impact We have already provided samples to our collaborator in University of W?rzburg collaboration. We plan to make reference to this collection of samples in the first publication which reports the generation of these new recombinant measles viruses. This has now been extended to colleagues in Johns Hopkins University (Baltimore, MD, USA) 
 
Description Duke University and Paul Ehrlich Institut 
Organisation Duke-NUS Graduate Medical School
Department Microbiology Duke-NUS
Country United States 
Sector Academic/University 
PI Contribution We are providing the recombinant measles, mumps and respiratory syncytial clinical isolates and will develop a high content screen to look for host factors which restrict or promote paramyxovirus infection.
Collaborator Contribution Providing access to high content screening platform of siRNAs against all known and putative human genes.
Impact mutlidisciplinary (virology, cell biology and automation technologies).
Start Year 2011
 
Description Duke University and Paul Ehrlich Institut 
Organisation Paul Ehrlich Institute
Department Veterinary Medicine
Country Germany 
Sector Academic/University 
PI Contribution We are providing the recombinant measles, mumps and respiratory syncytial clinical isolates and will develop a high content screen to look for host factors which restrict or promote paramyxovirus infection.
Collaborator Contribution Providing access to high content screening platform of siRNAs against all known and putative human genes.
Impact mutlidisciplinary (virology, cell biology and automation technologies).
Start Year 2011
 
Description Erasmus Medical Center 
Organisation Erasmus University Medical Center
Country Netherlands 
Sector Academic/University 
PI Contribution We have provided all of the recombinant viruses which have been used extensively in the monkey model. We host visitors from Rotterdam at QUB, where we train them in bioimaging, immunohistochemisty and pathological assessements.
Collaborator Contribution This partnership has allowed us access to a macaque model of measles virus pathogenesis which is not available in the United Kingdom. Scientists from Rotterdam participate in small animal experimentation in The Queen's University of Belfast (QUB) and have been instrumental in developing our immunological capabilities.
Impact This collaboration enabled me to make an application to the Bill and Melinda Gates Foundation Grand Challenges in Global Health Initiative/Foundation of the National Institutes for Health which has recently been funded.
Start Year 2006
 
Description Harvard Medical School, Harvard University 
Organisation Harvard University
Country United States 
Sector Academic/University 
PI Contribution We are using the systems developed to generate recombinant viruses which can be used to dissect how the virus interefers with the inate immune response on a mechanistic level.
Collaborator Contribution Our collaborator, Michaela Gack (HMS), has used the viruses we generate to dissect the role of the PPD-1 pathway in dendritic cells.
Impact Paper in preparation.
Start Year 2013
 
Description University of Würzburg 
Organisation University of Wurzburg
Department Institute of Virology and Immunobiology Würzburg
Country Germany 
Sector Academic/University 
PI Contribution We have provided many recombinant viruses which have been used extensively. We host visitors from Würzburg in The Queen's University of Belfast and train them in reverse genetics technologies, pathological assessment and bioimaging.
Collaborator Contribution We organise an annual, invitation only, European Measles Virus Workshop in Würzburg. There we critique data, share ideas, formulate strategy and distribute reagents to help facilitate the ongoing programmes of work in the partner laboratories.
Impact I have a rolling visiting professorship which is sponsored by Sonderforschungsbereich 479 (an interdisciplinary integrated twelve year programme of work funded by the Deutsche Forschungsgemeinschaft (DFG). This enables me to spend between two to four weeks working in Würzburg each year. As a result of this, alongside the establishment of the International Graduate Centre in the University of Würzburg, I now co-supervise three PhD students who work in the Institute who are supported by the DFG.
 
Description Northern Ireland Multipe Sclerosis Society 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact We hosted representitatives from the Multipe Sclerosis Society in Northern Ireland who toured the bioimaging facilities in the University and met with scientists undertaking MS related research.

This visit consolidated the University's already strong relationship with the MS Society and they appreciated how the funds they had contributed to purchase a state-of-the art confocal microscope had allowed the development of a vibrant bioimaging community within the University.
Year(s) Of Engagement Activity 2009
 
Description SCIART 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? Yes
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Under the auspices of the University's Bioimaging Core Technology Unit, of which I am co-academic lead, we host final year students from St. Mary's University College (191 Falls Road, Belfast, Northern Ireland) who take inspiration from photomicrographs produced using our confocal and electron microscopes. Thereafter the students produce a range of art works, some of which are on display in the unit.

The Head of the Art Department at St. Mary's presented a paper entitled, "An exploration of SCIART in various educational contexts", at the 32nd National Microscopical Society of Ireland conference, which was held in The Queen's University of Belfast.
Year(s) Of Engagement Activity 2008,2009
 
Description SGM Blog Interview at ESV Meeting 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact I was interviewed by the SGM for their monthy blog: Microbe Post

Twitter interest (@10queues)
Year(s) Of Engagement Activity 2013
URL http://microbepost.org/2013/11/11/microbe-talk-november-2013/
 
Description This Week in Virology #286 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact This Week in Virology (TWiV) is the most widely listened to virology podcast in the world. It has international reach and is listened to by both professional scientists and the general public.

Significant number of colleagues and press follow up. Excellent recruiting tool for PDRAs
Year(s) Of Engagement Activity 2014
URL http://www.twiv.tv/2014/05/25/twiv-286-boston-twiv-party/