Microdose trials test compounds in doses of 100 µg or less. Using such low doses means that LC/MS/MS assay sometimes isn’t sensitive enough, in which case radiolabelled compound is needed to enable an acceptably sensitive and precise assay. Carbon-14 is the radionuclide of choice, because tiny, harmless amounts can be quantified by Accelerator Mass Spectrometry (AMS), which offers sensitivity in the picogram to attogram range. We’ve successfully collaborated with AMS laboratories such as Xceleron in the early identification of viable drug candidates, and the elucidation of metabolic pathways.
Absolute bioavailability – a valuable application of microdosing
Most medicines are given by mouth, and it’s really helpful (and potentially important) to know what fraction of an oral dose reaches the systemic circulation. That’s the absolute bioavailability of the drug, which can be determined by measuring plasma concentrations after an oral and an IV dose. Formerly that required the development of an intravenous formulation, plus a programme of IV toxicology studies. Now, however, it’s possible to give the IV dose as a microdose, after just one 2-week tox study in rats. Given that the dose in humans is 100 µg or less, the IV formulation is very simple.
A typical trial to measure absolute bioavailability may be done in as few as 6 healthy volunteers, who each receive a single, therapeutic, oral dose of IMP containing no radioactivity. At about the time of peak plasma concentrations of oral drug – typically 1.5 – 2 h after oral dosing – the same subjects all receive an IV injection of a microdose of the IMP, which contains a ‘light label’ of carbon-14. Serial blood samples are then analysed for concentrations of unlabelled IMP, usually by LC/MS/MS, and carbon-14-labelled IMP by the AMS method. The results allow separate calculation of the plasma AUC attributable to the oral dose and to the IV dose. The ratio of AUC oral:IV is the absolute oral bioavailability of the IMP. It’s a simple and reliable method, and it’s now included in many drug development programmes.
Absolute bioavailability and ADME – 2 trials for the price of 1?
It makes perfect sense to add an assessment of Absorption, Distribution, Metabolism and Elimination (ADME) to your absolute bioavailability trial. You can use the same subjects, and the same source of carbon-14-labelled IMP. About 1 week after the completion of blood sampling in the oral dose and IV microdose phase, the subjects receive a therapeutic dose of IMP labelled with carbon-14, followed by serial blood sampling, and total collections of urine and faeces – typically for 7 days after dosing. Analysis by liquid scintillation counting and/or AMS yields ADME data, which can extend to characterisation and quantification of the IMP’s metabolites.
Microdosing in Phase 0 trials
Microdose trials, or Phase 0, are sometimes used to gain preliminary information about the pharmacokinetic properties either of a single compound, or of several compounds that are competing for development resources, before committing time and resources to Phase I trials. Human microdosing has now been used by at least 15 of the 20 largest pharmaceutical companies in drug development, and the technique has been provisionally endorsed by the European Medicines Agency (EMA) and the USA Food and Drug Administration (FDA).
At HMR we’ve successfully completed 8 clinical trials using microdoses of carbon-14-labelled IMP. We now do 1–2 such trials a year.
We have a dedicated radiopharmacy, including an aseptic suite, for preparing radiopharmaceuticals. We prepare injections and intravenous infusions, and do terminal sterilisation using a validated filtration procedure. Our radiopharmacy is experienced in preparing microdoses as well as conventional doses of radiolabelled labelled IMP and non-IMP.
We use on-site liquid scintillation counting (LSC) to QC our radiolabelled formulations of IMP. We also use LSC to estimate the extent of adsorption of IMP to the materials used in its preparation, such as plastic tubing and filters.
On our wards, we strictly segregate areas used for trials of ‘light label’ radioactivity and conventional doses of radionuclides. The same strict segregation is applied in the HMR laboratory, where the samples are processed.
We hold an Environment Agency licence to store and dispose of radioactive materials. Our ARSAC licence holder is on-site. Our permit from the Health and Safety Executive (HSE) allows us to use carbon-14, technetium-99m, chromium-51, iodine-125, caesium-137+ and molybdenum-99.
Our work with radioactive materials complies with all relevant acts and regulations, including: the Health and Safety at Work Act 1974, the Environmental Permitting Regulations 2016 (EPR16); the Ionising Radiations Regulations 2017 (IRR17); the Ionising Radiation (Medical Exposures) Regulations 2017 (IR[ME]R17); and the Carriage of Dangerous Goods and Transportable Pressure Equipment Regulations 2009, with amendments (2011/2013).
Lappin G, Boyce MJ, Matzow T, Lociuro S, Seymour M, Warrington SJ. A microdose study of 14C-AR-709 in healthy men: pharmacokinetics, absolute bioavailability and concentrations in key compartments of the lung. Eur J Clin Pharmacol 2013; 69: 1673–1682
Ambery C, Young G, Fuller T, Lazaar AL, Pereira A, Hughes A, Ramsay D, van den Berg F, Daley-Yates P. Pharmacokinetics, Excretion, and Mass Balance of [14C]-Batefenterol Following a Single Microtracer Intravenous Dose (Concomitant to an Inhaled Dose) or Oral Dose of Batefenterol in Healthy Men. Clin Pharmacol Drug Dev 2018; 7: 901–910
Okour M, Derimanov G, Barnett R, Fernandez E, Ferrer S, Gresham S, Hossain M, Gamo FJ, Koh G, Pereira A, Rolfe K, Wong D, Young G, Rami H, Haselden J. A human microdose study of the antimalarial drug GSK3191607 in healthy volunteers. Br J Clin Pharmacol 2018; 84: 482–489