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The development of phage-based vaccines for animal health.

Primary Supervisor: Dr Andrew Millard, Department of Genetics and Genome Biology

Secondary supervisor:Dr Ed Galyov , Prof Martha Clokie and Prof Venugopal Nair, Pirbright Institute

PhD project title: The development of phage-based vaccines for animal health

University of Registration: University of Leicester

Project outline:

The challenge: to develop good and cheap phage-based vaccines against animal diseases at scale required for mass vaccination of agricultural animals.

BACKGROUND: Bacteriophages are the most abundant biological entity on the planet, with 1031 in the biosphere. Despite being a natural part of the human microbiome, naturally occurring bacteriophages are known to induce adaptive immune responses. It is this natural response that we plan to exploit to use phages as a vaccine. Furthermore, phages are very cheap to produce in large quantities. There is a large diversity of phage types, with several having been previously used for vaccine applications, including phages T4, M13, MS2, and T7[1]. We will utilise phage T7, which has previously been used to develop vaccines against viruses causing influenza, Foot & mouth disease and mononucleosis. In all cases vaccination with recombinant T7 displaying antigens from the pathogenic viruses has resulted in the production of neutralising antibodies [1].

It is well established that proteins can be displayed on the capsid of T7 by fusing the antigen to the minor capsid protein of T7 (10B). By displaying on the capsid head in a repetitive and organised manor (as a result of the structure of the capsid) a higher immunogenicity of foreign antigens to B cells is thought to occur. A limiting factor in the modification of the capsid head is the ability to rapidly alter and engineer the genome of T7 to display these antigens. We are uniquely positioned to do this after having now optimised the ability to rapidly engineer the genome of phage T7 [2]. Possible disadvantages of using phages as vaccines are the introduction of a biological entity that can potentially replicate uncontrollably, if it finds a suitable E. coli host and phages do not always have strong adjuvant characteristics.

PROPOSED WORK:

Objective 1: Produce a non-replicate version of phage T7 with the DNA polymerase replaced by a large CpG island

Objective 2: Engineer the non-replicative, strong adjuvant form of T7 with (in first instance) fragments of structural antigens from the MDV virus, a causative agent of Marek’s disease, expressed on the capsid and perform preliminary characterisation of the phage and levels of MDV’s peptides exposed on its surface. Normally ~45 10B subunits are incorporated on the capsid, so similar number of MDV’s peptides would be expected to be displayed.

Objective 3. To assess safety, immunogenicity and protectivity of the T7 MDV vaccine prototype in vitro and relevant animal systems.

We have already produced non-replicative forms of T7 by alteration of the tail fibres and can easily adjust this to remove the T7 DNA-polymerase. We will introduce a large CpG island into the carrier phage genome as it is known CpG islands can act as adjuvants that are known to boost the immune response [3].We will then construct a non-replicative adjuvant version T7 phage with multiple copies of peptides from the MDV displayed on its capsid.

Objective 4. Develop a suite of plasmids that rapidly allows different antigens to be displayed in multiple combinations on phage T7.

POTENTIAL IMPACT: Unlike most of subunit vaccines, a phage-based vaccine will be stable and very cheap to manufacture, and the technology easily transferable to multiple users. It will use a highly stable carrier base and will not require adjuvant. It will be likely recognised by the immune system as a “virus” thus eliciting broad humoral and cellular immune responses that are essential for protectivity.

References:

  1. Hess, K. L. et al. Bioeng. Transl. Med. (2020) 2 Grigonyte, A. M. et al. Viruses (2020) 3 Krieg, A. M. Annu. Rev. Immunol. (2002)

BBSRC Strategic Research Priority: Sustainable Agriculture and Food: Animal Health and Welfare & Microbial Food Safety. Understanding the Rules of Life: Immunology & Microbiology

    Techniques that will be undertaken during the project:

    • Molecular biology techniques
    • Phage engineering
    • Protein expression and analysis
    • Vaccine testing in vitro and in vivo

    Contact: Dr Andrew Millard, University of Leicester