Over the past two decades considerable industry and regulatory discussion has focused around the importance of drug metabolites as potential sources of drug toxicity.
The issue of metabolite-mediated toxicity, and when it should be assessed, has become known as metabolites in safety testing (MIST). Essentially, it tackles the following question: When does a human metabolite become important? It is crucial to MIST to identify the major circulating human metabolites and ensure the species used in safety assessment are exposed to the same metabolites, at similar levels of exposure.
What do the regulators say?
A regulatory guidance framework is available for drug developers to help ensure the safety assessments (toxicological testing) conducted in animals (typically a rodent and non-rodent) are relevant to assess potential human drug toxicities.
In 2016, the FDA updated the issued guidance entitled ‘Safety testing of drug metabolites.’ Here’s a quick recap of the principal recommendations regarding the concept of MIST:
- To encourage the identification of differences in drug metabolism between animals used in nonclinical safety assessments and humans as early as possible during the drug development process
- Generally, metabolites identified only in human plasma or metabolites present at disproportionately higher levels in humans than in any of the animal test species should be considered for safety assessment
- Human metabolites that raise a safety concern are those formed at greater than 10 percent of total drug related exposure
These guidance documents lay out criteria describing when a human metabolite needs to be measured in laboratory animal species. They also define the circumstances under which direct testing of a metabolite in animal toxicology studies would be needed to provide a risk assessment.
It is vital to stress the importance of understanding drug metabolism as a key driver of pharmaceutical drug development. Ultimately, how a drug is metabolised is often what can drive pharmacokinetics, hepatotoxicity and drug-drug interactions. Undeniably, it is also a major driver of MIST for the reason that drug metabolism generates the metabolites. For any new drug in development, therefore, an early understanding of potential MIST issues (human-specific metabolism) is crucial to help ensure the metabolic relevance of the animal models used in its safety assessment (toxicity testing).
What safety assessment is required?
The definitive human metabolism data set is obtained from conducting a human radiolabelled (14C) AME study (absorption, metabolism, excretion), where a radioactive form of a drug is given to a small number of human volunteers (n = 6-8 cohorts). Typically, this vital study is often conducted during late phase development (after Phase II/proof of concept). This may seem a prudent approach given these human radiolabelled studies are not inexpensive to run. However, if the definitive human data set is obtained too late, it can result in significant delay(s) in the NDA/approval process should it reveal a disproportionate human metabolite to which animal models used in safety assessment were not adequately exposed.
Any major human-specific plasma metabolites will require additional safety assessment, which in turn requires a large scale synthesis of the metabolite - a time-consuming and expensive late-stage requirement. Ultimately, possible termination of the drug may be warranted if toxicity is observed with any of the metabolite(s).
How to assess the risk of MIST
To assess MIST risk, an integrated use of both in vitro and in vivo studies should be employed, such as given in the figure below, to inform whether human-specific metabolism could be an issue. If it is, the human 14C AME in vivo study should be prioritized and conducted with some urgency to obtain the definitive human metabolite profile.
In vitro techniques for early de-risking have the ability to generate human metabolic data (from microsomes and/or hepatocytes) early in the development process. Metabolic profiles can be quickly obtained using human hepatocytes and compared with animal species to investigate the formation of possible human-specific, or disproportionate, metabolites. These in vitro data can then be compared to in vivo metabolic data (plasma, blood, feces) to gain an understanding of the strength of the in vitro - in vivo extrapolation (IVIVE), which is pivotal to the MIST assessment.
Learn more about identifying strategies to detect early MIST issues in our whitepaper ‘Pharma de-risking: Approaches to reduce liability to major metabolically-driven causes of drug attrition’
What do the safety assessment results mean?
If after conducting a MIST strategy, there are no major differences between the metabolites formed in the relevant animal models in vivo and the human in vitro (hepatocyte) metabolism, then there is a scientific, rational basis for not needing to prioritize a human 14C AME study early, as the animal in vivo metabolites and exposures are likely to be reflective of human metabolism.
However, if there are differences between animal in vivo and human in vitro (hepatocyte) metabolite profiles, there is risk of a MIST issue (inappropriate animal species used for safety assessment and/or human-specific metabolism) and there is therefore a need to prioritize the human 14C AME study early to obtain the definitive human metabolite data.
Explore MIST in more detail by streaming Guy Webber’s on-demand webinar ‘Metabolites in safety testing.’
About the author
Guy Webber, Scientific Manager, In vitro and drug-drug interaction sciences at Envigo, has 25 years’ experience in Pharma and Contract Research, working on hundreds of drug molecules for many different clients.