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CASE STUDY 2: BETTER TOXICOKINETIC DATA WHILE ADDRESSING THE 3RS
Alexis Caron and his colleagues at the department for Disposition, Safety, and Animal Research, Sanofi-Aventis, needed to maximize the production of high quality data from mice and nonhuman primates in the toxicological evaluation of an Antibody-Drug Conjugate (ADC) developed for cancer. At the same time, they needed to minimize the impact of sampling and the number of animals used. They therefore evaluated the technical aspects, applicability and relevance of miniaturized blood-sampling procedures matched with nanoliter-scale immunoassays run on Gyrolab xP workstation (Ref 7).
MORE DATA POINTS FROM FEWER MICE
The research group’s initial studies on mice showed that repeated sampling of 25 µL at the saphenous vein using a glass capillary could be done with minimal restraint and without anesthesia, which limited stress and improved recovery from the procedure. They could draw the maximum allowable volume of 150 µL blood over 24 h or 168 h without any notable effect on animal welfare. This encouraged them to determine if the microsampling technique (25 µL) could be used in real toxicokinetic studies on the ADC, by comparing with standard sampling methods (120 µL). A total of 122 mice were exposed to different doses of ADC or vehicle (buffered solution) and then sampled using standard sampling or serially sampled using microsampling techniques.
ADC concentrations in all plasma samples were analyzed using a stepwise sandwich immunoassay run on Gyrolab xP workstation, chosen for its low sample consumption. With the exception of one outlier, the results from the standard and microsampling methods generally correlated well (see Figure). Results were consistent between sexes and among dose levels and time points.
MICROSAMPLING IN NONHUMAN PRIMATES
Microsamples of 50 µL blood taken from the heel of cynomolgus macaques were compared with standard samples of 1.2 mL taken from the saphenous or femoral vein. Microsampling could be performed with minimal restraint and in the animal’s home cage. Again, the test animals had been exposed to the ADC at different doses before samples were taken at different times and analyzed for ADC using an immunoassay run on Gyrolab xP workstation. The results showed that microsampling gave a consistently lower ADC concentration compared with standard sampling, which may have been due to the difference in sampling site, or other differences in sample handling. This finding highlights the need to carefully validate microsampling methods before they are used routinely.
AUTHOR’S CONCLUSIONS
• In mice, samples could be taken with only light restraint and without anesthesia, reducing animal stress and simplifying handling
• In monkeys, microsampling with minimal constraint in home cages reduced stress and increased safety for personnel
• Microsampling enabled more data to be generated with less animal stress
• Observed toxicological effects can be correlated with drug exposure at the individual level
Bland-Altman plot of plasma concentrations of ADC in samples taken using standard sampling and microsampling methods. Adapted from Figure 4 in Ref 7.
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