Stage 
A
Perform discovery, safety, immunogenicity and efficacy evaluation in initial animal model
Gate 
A
Progress to proof of concept (POC) studies in animals
Stage/Gate A
Project Management
Main Activities
  • Draft the Target Product Profile (TPP) with indication, target population, etc.
  • Set activities, deliverables and criteria to pass Gate A
CRITERIA REQUIRED
  • TPP with primary indication prepared
  • Activities, deliverables and criteria to pass Gate A agreed and finalised
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Guidance
Guidance

The Target Product Profile (TPP) is first drafted at stage A. The TPP lists key characteristics of the vaccine candidate and evolves as the project progresses. A standard TPP specifies the primary indication (for example, prevention of TB disease), the target population (for example, adolescents and adults), a description of the active drug substance (for example, a live attenuated recombinant BCG that expresses given, new antigens), the drug product (with additional excipients), and the general characteristics of the vaccine candidate. Additional information includes presentation (liquid, lyophilized), dose route and schedule and manufacturing aspects (for example, number of doses and affordability). Information related to clinical aspects will include: safety and reactogenicity, immunogenicity (immune response, mechanism of protection) and efficacy (prevention of disease, speed of onset of immunity), the number of administrations and schedule (multiple, time between doses), the route of administration , durability of protection (lifetime or time-limited), and whether co-administration with other vaccines (and potential interference) is anticipated. The safety, immunogenicity and efficacy of the vaccine candidate will be documented in animals first, then in humans.

In anticipation of the regulatory strategy the draft TPP should include registration (in relevant countries), as well as business (for example, analysis of competitive advantage). Where possible, the targeted countries and an estimate of the market potential are included, as these might have an influence on the design of the vaccine, and its geographic development.

At the early, draft stage of the TPP, vaccine candidates are in construction with limited characterisation and data. It is recommended to avoid ‘unknown’ or ‘to be determined’, and rather set targets and specify how information will be obtained, for example, a specific type of study for safety or efficacy. Often, the TPP contains a column of criteria that MUST be met, and another column of criteria that are ‘Nice to have’.

There are multiple sources for general guidance on preparation of a TPP which include:
Lee and Burke, Vaccine 2010, FDA Guidance and UK MRC guidance. For guidance with more specificity for TB vaccines it is suggested to review the package insert for commercial BCG (BCG SSI, BCG SII and BCG Tice) and the WHO published Preferred Product Characteristics (PPC) for TB vaccines. TBVI will shortly publish a generic TPP for a TB vaccine for Therapeutic applications.

Stage/Gate A
Product Characterization and quality
Main Activities
  • Screen and select antigen(s), adjuvants, other excipients and delivery system
  • Characterise vaccine candidate. In particular, demonstrate antigen expression and purity (e.g. proteins)
CRITERIA REQUIRED
  • Antigens, adjuvants, other excipients, delivery system selected
  • Characterisation tests defined and characteristics of the vaccine candidate documented
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Guidance
Guidance

Discovery is the stage where the technology and design of TB vaccine candidates are made, and when TB antigens, adjuvants, delivery system are screened based on expression (yield) and immune response in animals.

Key for the characterisation of the drug substance are assays to measure its identity, potency, purity, and stability. These assays require specific development and depend on the type of vaccine, for example a TB subunit vaccine will require a different assay for identity than a recombinant BCG. Identification and expression of the substance often includes gel electrophoresis, western-blot, or Enzyme Linked Immunosorbent Assay (ELISA) for a protein, and Polymerase Chain Reaction (PCR) for recombinant DNA. Potency could be bacterial viability (colony-forming unit) for a recombinant BCG, coupled to a marker assay for an immunological relevant effect representing protection. For a subunit vaccine it could be the concentration linked to a protective immunological effect, and for a viral vectored vaccine it would be titre linked to a similar output. For TB vaccines, T-cell epitopes and associated responses can be evaluated as markers for immunogenicity and potency. Examples include- antigenspecific stimulation of Interferon gamma production, measured by ELISpot, other cytokines, specific T-cell populations such as polyfunctional T-cells. Purity assays measure contaminants that originate from the expression system and culture medium used, such as host cell protein, host cell DNA, endotoxin or serum components, or from the process conditions used, e.g. detergents (Triton) or DNAse (benzonase).

The characteristics of modified TB as live attenuated TB vaccine have been published in the ‘Geneva Consensus’ publications (Geneva consensus and 2nd Geneva consensus).

The following guidelines on quality control of vaccine development provide further information: ICH Q8 Pharmaceutical Development, August 2009 (ICH Q8), ICH Q11 Development and Manufacture of Drug Substances (chemical entities and biotechnological/biological entities), May 2012  (ICH Q11) and ICH Q2, Validation of Analytical Procedures: Text and Methodology, nov 2005 (ICH Q2).

Stage/Gate A
Production process
Main Activities
  • Select expression system and lab-scale production process for generating pre-clinical material
  • Select specific strain
CRITERIA REQUIRED
  • Suitable expression system and process that can produce target product quantity and quality selected
  • Specific strain(s) selected and history recorded
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Guidance
Guidance

The criteria to select the production technology, beside in-house knowledge and experience, should be the target quality and quantity of the vaccine to be produced. It will influence the decision for the biological expression system and associated target production process that can deliver the target product criteria. Different production processes can be evaluated for suitability with a non-standardised monitoring of product quantity and quality (research quality). The most critical step is the selection of the biological expression system, as it determines the upstream part of the process and quality features of the product.

An initial step is the selection of strain(s). It is important to document the history of the strain as it will become the source of the Master Cell Bank (MCB) and Working Cell Bank (WCB), or the Master Virus Seed and Working Virus Seed Lot for viral vectors.

A target cGMP compliant process exists for BCG suspension cells and this process could be used as the platform technology for the development of a production process for modified BCG and attenuated Mycobacterium tuberculosis (Mtb) vaccines. Note that the traditional method of pellicle culture of BCG which was established in the pre-GMP era is difficult to match with cGMP guidelines.

At the end of this stage, an expression system and lab-scale process must be selected and used to produce pre-clinical R&D material. This process should, in theory, be based upon initial tests, literature and previous projects, and deliver the estimated quantity and quality of the TB vaccine candidate(s).

Stage/Gate A
Preclinical Safety
Main Activities
  • Identify in vitro and in vivo models to test for safety
  • Test preclinical safety elements relevant to candidate
CRITERIA REQUIRED
  • Safety tests identified
  • Safety evaluated; no evidence to suggest lack of safety
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Guidance
Guidance

Safety testing at this Stage should first demonstrate that the vaccine material is suitable for use in animals. For example, protein components of protein/ adjuvant vaccines should be tested for and contain minimal levels of endotoxin, ensuring both that the candidate is safe for use in animals and that interpretation of immunogenicity and/ or efficacy data is not confounded by the presence of  innate stimuli. Further evaluations of safety at this Stage are generally observational, and can be included in immunogenicity and efficacy studies. These should reveal an absence of safety signals such as excessive local inflammation at the site of injection or indications of anaphylaxis with repeated inoculations. Safety characteristics specific for live attenuated whole cell vaccines (strains of Mycobacterium tuberculosis (Mtb)or modified BCG) are described in Walker et al., 2010, and are aimed at demonstrating safety better than BCG, for example in the immunocompromised SCID mouse model.

Stage/Gate A
Preclinical Immunogenicity
Main Activities
  • Evaluate immunogenicity
  • Compare to benchmark, if applicable
CRITERIA REQUIRED
  • Evidence of relevant immunogenicity to antigens in at least one animal species
  • Above baseline and/or benchmark (if applicable) responses to antigens preferred
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Guidance
Guidance

In Stage A, immunogenicity studies should demonstrate that a clear immune response to the target antigens can be induced in the same animal model used to demonstrate protection. Mice are often used in early studies, but may not be appropriate for all candidates (for example, it would not be appropriate to evaluate vaccines based on CD1-associated lipids, as mice lack most CD1 molecules). Balb/C or C57Bl/6 strains of mice are frequently used but, as there may be differences in the immune response caused by MHC-restriction, an alternative is to use CB6F1 mice (see Stylianou et al 2018 and Aagaard et al., 2011. In the absence of a correlate of protection, the immune parameters measured should be relevant to immunity to Mtb and to the proposed mechanism of action of the vaccine. Due to the costs of Mtb protection studies, early-stage decisions e.g. on doses, formulations etc, would be most likely based upon immunogenicity, such as the magnitude of antigen-specific T-cells. A response above baseline is considered a minimum requirement but, if similar to others in development, the candidate should show an obvious differentiating characteristic – qualitative or quantitative. In addition to optimisation of doses and routes of administration of individual candidates, immunogenicity studies in stage A may play an important role in prioritisation or selection between different candidates. For example to select between different antigens or vectors, or even in a portfolio management context to discriminate between different vaccine types, although such decisions are most likely to be multifactorial, with immunogenicity being only one factor.

Stage/Gate A
Preclinical Protection/efficacy
Main Activities
  • Demonstrate protection in a small animal Mycobacterium tuberculosis (Mtb) infection
  • Compare to benchmark, if applicable
CRITERIA REQUIRED
  • Protection in a small animal Mtb infection model demonstrated
  • Protection statistically better than a relevant benchmark (e.g. BCG or no vaccine, or ‘best-in-class’ comparator)
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Guidance
Guidance

The ability of a candidate to reduce or limit the progression of experimental infection with Mtb must be demonstrated in an appropriate animal model. Different animal species, modes of infection and read-outs of disease are used. These are described in reviews such as Cardona and Williams 2017, Singh and Gupta 2018, Williams and Orme 2016. Typically, efficacy is first demonstrated in mice or guinea pigs. There are many examples of these species being used for testing the efficacy of TB vaccines which are illustrated in the following references; Stylianou et al 2018, Doherty et al., 2004, Reed and Lobet 2005, Williams et al., 2005, Brandt et al., 2004, Clark et al., 2016

There are two, commonly used laboratory-adapted strains of Mtb used for challenge, preferably via the respiratory route, these are the H37Rv and Erdman strains. There is no specific requirement to use these strains and it may be considered an advantage to use clinical strains. The challenge strain should be well-characterised and from a source that allows reproducibility between experiments. The design of the vaccination schedule should be informed by previously generated immunogenicity data. In the case of protection studies in mice, the strain of mice used should also be consistent with the strain used to demonstrate immunogenicity. In the absence of a correlate of protection, evaluating multiple vaccine dose levels in early-stage challenge studies should be considered. The outcome measures used to demonstrate protection typically include a reduction in bacterial burden in lungs and other relevant organs, survival, or quantifiable changes in pathological features of TB disease. A statistically significant change compared to a relevant control group must be demonstrated. Statistical power calculations based upon a pre-defined target level of protection should be used to design experiments with appropriate group sizes. The control group against which efficacy is compared must be justifiable and appropriate to the nature and intended TPP of the candidate. For example, a novel live-attenuated whole cell vaccine should show an improvement over BCG unless other significant benefits would justify an equivalent level of protection to BCG. If a candidate is a part of a heterologous prime-boost regimen, the combined regimen must be significantly more protective than the individual components. If the prime-boost regimen has BCG as a critical component e.g. boosting BCG in infants, there is an expectation that protection greater than BCG must be demonstrated. At stage A, in order to show an efficacy signal, it is advisable to use a model e.g. mice where there is greater statistical power to show the BCG-boost effect, with tractable group sizes. Appropriate comparators to judge the relative effect of the vaccine might include empty vectors or, if available, best-in-class vaccines that are already in development.  

There is no published guidance about the most suitable model to use for specific vaccine candidates but organisations such as TBVI (TBVI services) or the Collaboration for Tuberculosis Vaccine Discovery (CTVD) will provide advice via, for example, product development teams (PDT – TBVI) or the research communities established by CTVD, in particular the NHP community.