Cross-industry demonstration of the validity of the mixed matrix method for the assessment of cross-species exposure coverage of human circulating drug metabolites.

The mixed matrix method (MmM) is a widely used approach by the pharmaceutical industry for early assessment of whether exposures to major human circulating metabolites, of traditional small-molecule drugs, are adequately covered by the species used for toxicology assessment, which is a key requirement of the safety testing of drug metabolites (metabolites in safety testing guidelines). However, questions remain regarding its accuracy and utility in replacing conventional bioanalytical approaches. Furthermore, the available literature on the topic is not fully consistent in terms of how the assay should be conducted. As a result, encouraged by health authority advice on this topic, a cross-industry group under the European Federation of Pharmaceutical Industries and Associations was formed to: (1) further investigate the MmM accuracy, including a robust statistical analysis covering a diverse chemical space of commercially available drugs and drug candidates as well as their metabolites; (2) propose recommendations for best practice including a decision tree that the industry should consider when using the MmM; and (3) discuss whether the MmM could be used to support metabolites in safety testing assessment and could potentially be included into new drug application submissions without the need for additional measurements using the conventional bioanalytical approach. The outcome of this European Federation of Pharmaceutical Industries and Associations assessment shows that MmM measured exposure ratios of 1.9 and 1.4 are statistically sufficient to demonstrate adequate exposure coverage of human metabolites above 50% or between 10% and 50% of drug-related exposure, respectively, by toxicology species. The aim is to encourage both industry and regulatory agencies to consider MmM as an acceptable approach to compare major human circulating metabolite exposures across species. SIGNIFICANCE STATEMENT: The outcome of our mixed matrix method assessment showed that measured exposure ratios of 1.9 and 1.4 are adequate to demonstrate coverage of human metabolites above or below 50% drug-related exposure by toxicology species. Recommendations for best practice and a decision tree for conducting metabolites in safety testing evaluations are proposed. Our investigations show that mixed matrix method data are sufficiently robust for the intended purpose and that the assay provides an opportunity to streamline drug development and reduce the need for resource-intensive bioanalysis and certain animal studies.