ADME Studies in Early Clinical Development: Addressing MIST
Mallé Jurima-Romet, Ph.D.
Senior Director, Drug Development
As discussed in the introductory section of this newsletter, “Understanding Human ADME”, the timing of human ADME studies is shifting earlier in the clinical development of a new drug. Part of the impetus for this shift can be attributed to the 2002 landmark publication of PhRMA/FDA recommendations on the safety assessment of circulating drug metabolites1. Subsequently, the FDA introduced a Guidance for Industry – Safety Testing of Drug Metabolites (“MIST”), published in final form in 20082, and the ICH M2(R2) guidance, published in 2009.3 These regulatory guidances have focused industry attention on the need to determine if human metabolites are adequately evaluated during non-clinical safety studies. The rationale for MIST is that drug metabolites potentially have a role in causing or contributing to the toxicity of a drug. If a metabolite is a unique human metabolite, or more commonly, if a circulating metabolite is present at disproportionately higher levels in humans than in the animal species used in toxicology studies, additional non-clinical safety assessment studies may be required. Per ICH guidance, a metabolite present in human plasma at 10% or more of total drug-related material should be present at equal or greater abundance in the circulation of at least one toxicology species. Exceptions are possible for lower risk metabolites such as most Phase II conjugated metabolites. In practice these guidances require metabolite profiling in humans during early clinical development. The challenge to the drug developer is deciding how to invest resources appropriately to address the MIST requirements at a stage of development when the risk-benefit ratio of a drug is still largely unknown.
Even before initiating clinical studies, it is feasible and advisable to learn about the human metabolism of a drug and compare metabolite profiles in humans and animal species. This can be done using in vitro systems such as hepatocytes, liver microsomes and recombinant enzymes, coupled with LC/MS-MS analytical technology. However, the data obtained from such investigations generally are not quantitative. Moreover, in vitro systems are limited in their ability to provide a complete metabolizing system, when compared to the in vivo situation. To supplement the animal in vitro metabolite data with more physiologically relevant in vivo data, plasma and excreta samples collected from animals in pharmacological and toxicological studies can be qualitatively profiled by LC-MS/MS. Quantitative metabolism studies in animals have been traditionally conducted using a single dose of 14C-labelled drug. More recently, the need for radiolabeled animal metabolism studies has been questioned4.
The first opportunities to collect in vivo human metabolite data are in the First-in-Human Phase I single ascending dose (SAD) and multiple ascending dose (MAD) studies. However, metabolite profiles from in vivo plasma and urine samples analyzed by LC-MS/MS are qualitative or semi-quantitative at best. This may be due to the unavailability of reference standards for metabolites and so the analytical assays would not have been validated for all potential metabolites. Therefore, it is challenging for a drug developer to make a decision on whether to further investigate the activity and safety of drug metabolites based on the qualitative or semi-quantitative LC-MS/MS data from Phase I studies.
Nonetheless, it is prudent to look for signals for potential active metabolite(s) in early Phase I studies. For example, if parent drug exposures are very low but dose dependent adverse effects associated with target pharmacology are observed, this may be an indication that one or more active metabolites are produced. In such a situation, metabolite structural determination of plasma samples by LC-M/-MS or Q-TOF MS should be attempted and metabolites isolated from HPLC fractions tested for activity. Active metabolites will need to undergo non-clinical safety assessment in toxicology and safety pharmacology studies analogous to the testing required for the parent drug. If none of the animal species commonly used in toxicology studies form the human active metabolite, or if they demonstrate lower exposures compared to human exposures, it may be necessary to synthesize the metabolite in larger quantities for direct administration to animals in toxicology and other safety studies.
A radiolabel ADME study in humans, using a single dose of radioactive parent drug, has been the mainstay study enabling comprehensive identification and quantification of drug metabolites. Before the human ADME study can be conducted, a whole body autoradiography study in rats is required to obtain dosimetry data. Since ideally the dose selected for the human study should be clinically relevant, it makes sense to conduct the radiolabel ADME study after enough efficacy data is available to estimate a clinically relevant dose. On the other hand, delaying the human ADME study to later in Phase II is inherently risky. Failing to identify unique or disproportionate human metabolites early on can lead to significantly longer development timelines if it becomes necessary to add a separate set of toxicology studies late in the development program.
There are some innovative strategies that drug developers can apply to address MIST early in clinical development. One approach is to administer an estimated “therapeutic” dose containing a microdose of a radiolabelled drug in the Phase I SAD study. As explained in the introductory section of this newsletter, an animal tissue distribution study is not required to support dosimetry calculations for a microdose study since the amount of radioactivity administered is so minute. From the SAD/14C-labelled microdose, 14C is measured by AMS (accelerator mass spectrometry) for metabolite quantification, while mass spectrometry and possibly NMR are used for structural elucidation5. To obtain sufficient levels of metabolites for analysis, an “AUC pool” can be created by pooling plasma samples across the collection period, with volume adjustment proportional to the collection period. Any metabolites that are found to be >10% of total drug-related material should then be monitored in repeat dose animal toxicology studies to determine if they are present at higher levels. Toxicokinetic samples from previously conducted or ongoing toxicity studies can be used for this purpose, provided that the stability of metabolites is not an issue. If metabolite exposure in at least one species is greater than in the human, then no additional safety studies of the metabolite are needed.
In summary, the MIST guidances have defined the knowledge needed for the metabolism of a new drug in humans and animals. The ultimate goal is to minimize the potential for clinical risk. Analytical technologies have advanced to enable quantitative and structural metabolite data from human ADME studies conducted during early clinical research. This approach therefore addresses MIST at a strategically advantageous stage of development.
- Baillie T.A., et al. Drug metabolites in safety testing. Toxicol. Appl. Pharmacol. 182: 188-196 (2002).
- FDA: Guidance for Industry: Safety testing of drug metabolites (February 2008).
- ICH M3(R2): Guidance on nonclinical safety studies for the conduct of human clinical trials and marketing authorization for pharmaceuticals (June 2009).
- Obach R.S., et al. Radiolabelled mass-balance excretion and metabolism studies in laboratory animals: are they still necessary? Xenobiotica 42: 46-56 (2012).
- Lappin G. and Seymour M. Addressing metabolite safety during first-in-man studies using 14C- labeled drug and accelerator mass spectrometry. Bioanalysis 2: 1315-1324 (2010).