Determine the toxicokinetics of compounds.

Toxicokinetics is the study of the relationship between drug pharmacokinetic parameters and chemical toxicity. The lack of liver drug metabolism in the HRN™ mouse enables studies to distinguish the maximum plasma concentration from total exposure. This is crucial in the development of anti-cancer agents where the overall efficacy is a delicate balance between the therapeutic effect and toxic side effects. The pharmacological efficacy and/or toxic side effects may be more dependent on either the maximum circulating drug concentration (Cmax) or total overall exposure to the drug (AUC). Toxicokinetic studies in normal animals are limited as the Cmax and AUC are often very closely linked with each other, and it has not previously been possible to distinguish between these two parameters. However the lack of hepatic metabolism in the HRN™ mouse allows the relative contribution of each parameter to toxicity and efficacy to be determined. The results from this type of experiment can have profound implications for optimising the clinical dosing regime, as the severity of adverse side effects can be minimised by changing the way in which compounds are administered.

An example to demonstrate this is the clinically used anti-cancer agent cyclophosphamide. Cyclophosphamide (CPA) is an anti-cancer pro-drug that is dependent on CYP metabolism to 4-hydroxy-cyclophosphamide (4-OH-CPA) for its therapeutic effectiveness. However its clinical use is accompanied by a number of side effects such as neurotoxicity, urotoxicity and myelosuppression. We have employed the HRN™ mouse to investigate the toxicokinetics of this drug to optimise the dosage regime.

Pharmacokinetic parameters of the active metabolite 4-OH-cyclophosphamide and the associated myelosuppression in normal and HRN™ mice following administration of 35-100mg/kg cyclophosphamide. The total exposure (AUC) and acute exposure (Cmax) to 4-OH-cyclophosphamide in HRN™ and wild type mice are shown compared to the degree of induced myelosuppression. Results demonstrate that there is extrahepatic metabolism of CPA as the overall exposure is the same (AUC) [Top panel], but the peak blood concentration (Cmax) [bottom panel] is lower in the HRN™as the rate of metabolism is different between the two mouse lines. This correlates with the associated myelotoxicity which is considerably reduced in the HRN™, thus demonstrating that the peak concentration is more important in determining toxicity. The area under the PK curve (AUC) and peak blood concentration (Cmax) are plotted relative to internal standards, whereas the myelotoxicity is plotted as the reduction in the number of bone marrow granulocytes relative to untreated controls.

The results of administering 50mg/Kg CPA to both HRN™and normal mice shown in the figures, demonstrate that the conversion of CPA to the active metabolite 4-OH-CPA is due to extrahepatic metabolism, and that the associated myelotoxicity is more closely related to the acute exposure (C_max) rather than the total exposure (AUC) to 4-OH-CPA. As the overall exposure to 4-OH-CPA has been demonstrated to be the major parameter that defines the therapeutic efficacy of CPA, these results suggest that it would be better to administer CPA as a continual infusion rather than acute administration which is the current clinical practice. This would not only improve the clinical effectiveness but also reduce the toxic side effects. Detailed kinetic information like this is of crucial importance when designing clinical trials to maximise the chance of successful outcome.