P.Burtin, J. vanBree, F. Mentre, S. Hensel, J.L. Steimer
SANDOZ PHARMA LTD. Drug safety Dept, Basle, Switzerland and INSERM U436, Paris, France.
Rationale.
The main objectives of Toxicokinetics are to assess the animal exposure after drug administration across dose levels and genders, and for repeated vs single administration. In rodents, Toxicokinetics is commonly assessed in a so-called satellite group of animals which is run in parallel to the main group devoted to toxicologic evaluation. This strategy is justified by the risk of altering the health of the main animals due to repeated sampling, and thereby jeopardizing the toxicologic evaluation. But new regulations in several countries tend to restrict blood withdrawal in animals; in addition, the relatively extensive blood withdrawal in satellite rodents may bias the kinetic results.
Objective.
Our objective was to evaluate wether the exposure could be assessed directly in the main group of animals from sparse samples. The only acceptable designs in the main group consisted of one single sample per animal repeated on 2 or 3 different study days. Our hypothesis was that the larger number of animals in the main group would overall compensate for the small number of samples per animal, and that the animals were sufficiently similar to support extremely sparse designs.
Methods.
The 13-week oral study of Drug D in rats with 3 dose levels involved both classical sampling in the 24 satellite animals and sparse sampling in the 60 main animals. In the satellite group, each animal was sampled at 0, 0.5, 1, 2, 4, 7, and 24 H post-dose on each of the first and last study days. In the main group, each animal was sampled at one out of five possible sampling times (0.5, 1, 2, 7, and 24 H post-dose) on the first and last study days. The sampling time was identical for each animal on the two occasions, and there were two animals per sampling time for each dose*gender combination (10 animals per dose*gender). The analysis of the satellite and main group kinetic data was performed by two independent investigators. In the satellite group, the individual exposure was estimated from the trapezoidal area under the curve (AUC) and the observed maximal concentration (Cmax). In the main group, the sparse data were analysed with two methods. On one hand, model-free mean estimates of exposure were computed, i.e. the trapezoidal AUC(0-24) under the averaged naive pooled profile and the mean observed Cmax for each dose*gender*study day combination. On the other hand, the kinetics were analysed with NONMEM and a one- compartment model found to be appropriate in earlier studies with compound D. Comparisons of results were based i) on the direct inspection of blood levels from the satellite and main groups, ii) on the qualitative conclusions drawn by the independent investigators regarding dose, gender, and study day effect, iii) on the mean AUC and Cmax estimates.
Results.
i) Concentration-time values of the main animals tended (not significantly) to be higher than those of the satellite animals.
ii) All techniques led to the same major qualitative conclusions: the exposure pattern was dependent on dose, and females were more exposed than males. In the main group, the two methods found a trend for accumulation between the first and the last study day, which was not detected in the satellite group. The model-based analysis of the main group data with NONMEM indicated that dose influenced the distribution but not clearance, that clearance was 20% lower in females than males, and 25% lower on the last than on the first study day.
iii) The AUC and Cmax estimates were similar between the three evaluations. There was a general trend for model-free exposure estimates to be higher than the model-based estimates.
Conclusions.
The very sparse sampling design applied to the main group yielded adequate toxicokinetic information about drug D, even with a very simple model-free analysis. The population analysis with NONMEM provided additional insight about drug disposition and the influence of the covariates. In addition, it provides Bayesian estimates of the individual animals' exposure for further exploration of the concentration-toxicity relationship.