Pathfinder:Experimental Human Challenge with Genetically Modified Commensals to Investigate Respiratory Tract Mucosal Immunity and Colonisation

Lead Research Organisation: University of Southampton
Department Name: Clinical and Experimental Sciences

Abstract

We will genetically modify a bacterium and then perform two pilot controlled human infections to road test the technique for investigating human immunity and discovering and testing new vaccines. The proposal is complex - we have to gain regulatory approval both for deliberate release and human work. In the future, vaccines for such diseases as pneumonia, meningitis and whooping cough will be more sophisticated than those in the current schedule, and will likely take the form of nose drops that contain molecules providing nuanced immunity in the nose and throat. This is the critical first entry point of bacteria that cause these diseases, and the key to new generations of vaccines is to understand how natural immunity blocks successful entry. Stopping colonisation by pathogens stops transmission to other people (herd protection). We know that proteins on the surface of bacteria help them gain a foothold. These proteins may also elicit immunity that could be harnessed as vaccines. The `signature` of the bacteria - the proteins on or within the bacterium - vary widely between bacterial families. This variation renders current protein vaccines effective against some, but not all, the pathogens to which we are exposed.
To make the future vaccines we need to understand more. Most of our knowledge is from laboratory studies with cells. There are some animal models but they are imperfect. Experimental human challenge is a powerful technique in which people volunteer to be infected with microorganisms in order to study the immune response to the whole organism, and the efficacy of vaccines or treatments. Our group was one of the first to infect the nose and throat with bacteria; we used a commensal Neisseria lactamica (`friendly bacteria`) and have infected over 350 human volunteers and shown it is safe and informative. However, the precision required for experimental medicine will follow if we can control and compare the signature of bacteria that we inoculate. This requires genetic modification.
In the first part of the project we will genetically modify N.lactamica. This should be straightforward because we have made a prototype. However we will then need extensive laboratory testing to show that the organism is not more hazardous than the wild type. This will involve seeing how easily it can be killed by antibiotics and human blood, and also how `stable` its genome is (ie is it more likely to be changed genetically into something more dangerous when it is inside the nose). We will need to do this before we approach authorities for permission to allow those people who are challenged to walk out into the community whilst they are still carrying the bacteria in their noses. We contacted DEFRA (the government body concerned with deliberate release) and they have told us what information we will need to provide.
On approval, we will enrol participants who will be admitted to our hospital research facility for 48 hours and undergo controlled infections. They will then be discharged with clear instructions how to prevent transmission to others. From the volunteers, samples will be taken of throat and nose fluid, and blood, to isolate carried bacteria (in the throat) and detect the cells and soluble products of the immune response, which will be characterised. Bacteria harvested from the throats will be genome sequenced to check carefully that the bacteria remain stable genetically.
Once this is done we will repeat the study - this time to discover whether the natural mechanisms bacteria employ to conceal their signatures (phase variation) cause a different immune response. This is important to know because it informs how to make sure future challenges of this type use the right engineering to address research questions.
This study will be a true pathfinder - this technique will fast-track discovery of the bacterial molecules - and the host response to them - that are critical for colonisation of humans by pathogens.

Technical Summary

This is a pathfinder study to establish the utility of experimental human challenge with a genetically modified commensal bacterium as a future method to investigate pathogenesis, immunity and to discover and test new vaccines. The commensal bacterium Neisseria lactamica (Nlac) does not have a polysaccharide capsule , so any adaptive immune response to this bacterium after colonisation of the nasopharynx must be directed at non-capsular antigens, providing a good platform for assessing non-anti-polysaccharide immunity against colonising bacteria in the upper respiratory tract. A wild-type strain of Nlac Y92-1009 (Nlac), will be chromosomally modified to express the strongly immunogenic meningococcal antigen PorA (GM-Nlac). GM-Nlac will be fully characterised, in particular with respect to enhanced antimicrobial or serum resistance, and altered genome stability cf Nlac. An application will then be made to DEFRA and the National Research Ethics Service to perform a controlled infection study in which non-smoking male participants aged 18-45 will be infected intranasally at a dose of 10,000 colony forming units (selected because we previously have infected >350 volunteers with Nlac at this dose) and then permitted to re-enter the community after challenge and followed up for 4 weeks. In a comprehensive serological and cellular immunological study we will define the nature and kinetics of the PorA-specific immune response to colonising GM-Nlac and measure recombination in carried isolates over the ensuing period of carriage. PorA is phase variable so we will determine the effect of phase variation in a second challenge experiment by comparing the serological and cellular responses to GM-Nlac expressing PorA under a phase variable promoter with one that is constitutively highly expressed. We will also compare the immune response to Nlac antigens between GM-Nlac and Nlac infected participants, to understand the effect of immunodominant antigens on immunity.

Planned Impact

The major expected impacts of this project will be dramatic effect on the ability to study bacterial infection of the respiratory tract, and in particular to understand the mechanisms of development of mucosal and systemic immunity to colonising pathogens. One impact on society will be a shortening of the research and development time for new vaccines by Academia and Pharma, and a new way for regulators to measure the potential cost-effectiveness of new vaccines.

Pharmaceutical Industry
This initiative will provide Pharma with a very valuable tool to identify candidate antigens, but it could also be used to test new vaccines for their carriage-reduction efficacy against defined bacterial surface molecules, refining vaccines until they exert the maximal effect. This element will add greatly to their regulatory submissions, because carriage-reduction adds enhances the cost effectiveness profile of new vaccines. Our proposed model would reduce greatly the cost and duration of such assessments and is therefore likely to form part of Pharma`s submission portfolios. It will also provide a system that innovative small companies with new vaccine ideas might be able to accelerate the recognition of their new product. It is likely that we will be able to generate this new potential within 4 years of award of this grant.

3Rs
Animal models are commonly used to study the immune response to bacterial infections, but there is a growing appreciation of the importance of asymptomatic colonisation in transmission and pathogenesis of disease. In many models, bacteria are genetically modified and the animal host is used to study the immune response so our proposal will circumvent this by expanding the opportunity to use human systems in experimental medicine.

Patients and Wider Public
The population will benefit from our research because it will provide an experimental medicine technique that to fast-track the development of new vaccines which, in the future, will target colonization of the oropharynx and the herd protection that results from this.

Charities
The major charities who will most immediately engage with us will be the Meningitis Research Foundation and Meningitis Now (formerly Meningitis UK). These are patient-originated charities who span research, advocacy and support of people with meningitis. They take a very active interest in prevention of the disease and lobby the government on issues to accelerate vaccine deployment. Our model will be entirely consistent with their programmes of rapid development and deployment of new vaccines.

Government/Policy Makers
This development will increase the effectiveness of public services and policy. If this model is developed successfully, it will provide the Joint Committee for Vaccines and Immunisation (of which applicant RCR is a member) and other advisory/ regulatory bodies (eg EMA/CHMP/MHRA) with an alternative system for measuring comparative ccarriage reduction efficacies of novel vaccines. Perhaps of even greater interest is that our proposed model would be a way to develop alternative surrogate markers of efficacy alongside the use of serum bactericidal antibody concentrations in vaccine recipients The reason this is significant is because many new vaccines fall by the wayside because they do not induce bactericidal antibodies sufficiently, and yet they might be able to prevent meningococcal disease via an effect on carriage.

Training/Development
This project will be a training vehicle for young clinicians and will generate two PhD projects for graduates contributing to the two experimental work packages.

Publications

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Collini PJ (2018) HIV gp120 in the Lungs of Antiretroviral Therapy-treated Individuals Impairs Alveolar Macrophage Responses to Pneumococci. in American journal of respiratory and critical care medicine

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Dale AP (2022) Neisseria lactamica Controlled Human Infection Model. in Methods in molecular biology (Clifton, N.J.)

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Dale AP (2022) Neisseria lactamica Controlled Human Infection Model. in Methods Mol. Biol

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De Graaf H (2020) Controlled Human Infection With Bordetella pertussis Induces Asymptomatic, Immunizing Colonization. in Clinical infectious diseases : an official publication of the Infectious Diseases Society of America

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Dorey RB (2019) The nonpathogenic commensal Neisseria: friends and foes in infectious disease. in Current opinion in infectious diseases

 
Description Collaboration with Professor Andrew Gorringe, expert on Neisseria lactamica, at Public Health England 
Organisation Public Health England
Department National End of Life Intelligence Network
Country United Kingdom 
Sector Public 
PI Contribution We have invented a method to chromosomally transform Neisseria lactamica so that it could be used as a bacterial medicine
Collaborator Contribution They have provided the wild type strain Y92 for us to perform the transformation and have performed the detailed safety evaluations in cellular and animal models to enabkle an application to DEFRA for a deliberate release experiment in humans
Impact multidisciplinary
Start Year 2016
 
Title Meningococcal Infection and Modified Neisseria lactamica 
Description A method to genetically transform Neisseria lactamica to express antigens in order to prevent infection in the upper respiratory tract 
IP Reference US-2019-0307874-A1. 
Protection Patent application published
Year Protection Granted 2019
Licensed No
Impact None yet