CPSA Digest 2003

Day 3

WeOA2

Implications of Metabolites in Safety Testing (MIST) Guidelines

Issues

• Are there major human circulating metabolite(s) to which animals have not been adequately exposed?
• Is there is a unique metabolite in human plasma to which the safety species are not being exposed?
• Is any metabolite active?
• Does any metabolite possesses a structural alert or arise from a reactive intermediate?

General Practice
In industry, researchers rely on 14C-ADME clinical studies to define actual in vivo human metabolism. However, these studies are often not conducted until early phase II of a clinical program. Normally, only the parent drug is quantified to support early preclinical safety and toxicology studies. Human metabolites are not routinely measured in the early preclinical safety and toxicology program. Thus, bridging studies (ancillary toxicokinetic studies) may be necessary to assess exposure to major human metabolites and establish a safety margin from non-clinical species.

Problems with these approaches

• Qualitative and/or quantitative changes in metabolite(s) can occur following multiple doses that are not predicted by single dose 14C -ADME clinical studies.
• The identification of CYP450, UGT or other enzymes responsible for the formation of relevant in vivo human metabolites of a drug is, at best, "guess work" based solely on in vitro assessment without human data.
• The determination of CYP450, UGT or other enzymes responsible for the formation of major in vivo human metabolites is critical to designing appropriate drug-interaction studies.

Schering-Plough Approaches
Schering-Plough proactively addresses some of these issues early in clinical development, such as routinely profiling metabolites in plasma and urine collected from first-in-man (FIM) studies. Since these studies are non-radiolabeled, samples are surveyed by full scan LC-MS/MS for metabolites previously observed from in vivo animal and in vitro studies.

These initial "cold" studies do not necessarily limit the ability to discover new metabolites because of advances in the LC-MS technologies and scanning options. Also, many experimental drugs contain easily identifiable atoms such Cl or Br which have a definitive isotope pattern.

Metabolism data generated from FIM studies have limited quantitative value due to unknown intrinsic mass spectrometry response of metabolites relative to the parent drug, extraction recovery and stability of drug derived material during sample collection, handling, processing, and analysis conditions.

Nonetheless, a circulating human metabolite which achieves a significant (50%) response relative to parent drug deserves follow-up. Reference standards are synthesized as necessary for confirmation. Depending on structure and/or fragmentation pattern, an interference check is performed against the validated bioanalytical method for the parent drug. It can then be decided early in clinical development whether to validate a bioanalytical method for metabolite(s) and whether studies should be designed to assess exposure to metabolite in preclinical species.

• Detect and identify
      - Human specific metabolites
      - Important single vs. multiple dose differences in metabolism
• Provide feedback about the quantitative bioanalytical method
      - Verify selectivity (metabolite interference)
      - Determine need to validate for additional analyte(s)
• Offer guidance to Drug Safety assessment of exposure multiples to major circulating human metabolites
• Verify responsible P450, UDP-glucuronosyltransferase (UGT), etc. determined in vitro leading to the design of appropriate clinical drug-drug interaction studies

References
Thomas A. Baillie, Mitchell N. Cayen, Hassan Fouda, Ronald J. Gerson, James D. Green, Scott J. Grossman, Lewis J. Klunk, Bernard LeBlanc, Darcy G. Perkins and Lisa A. Shipley, "Drug Metabolites in Safety Testing," Toxicol. Appl. Pharmacol. 182 188-196 (2002).

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