In vivo studies in rat and rabbit

Besides mouse and monkey, the species employed in general toxicity assessment, rat and rabbit, were also investigated as preclinical species for reprotoxicity testing. Plasma pharmacokinetics was assessed either in preparatory PK‐studies (rat) or from regulatory toxicity studies (rabbit). In Wistar rats, PK‐samples were collected after an intravenous bolus injection at 0.1, 0.25, 0.5, 1, 2, 4, and 6 h postadministration. Plasma PK in Chinchilla rabbits were recorded after intravenous infusion (into the ear vein) at various dose levels (i.e., 50, 150, or 450 mg/kg/d; 85, 255, or 764 μmol/kg/d) with the following sampling times: before, and 2, 4, 4.25, 4.5, 5, 7, and 10 h after start of infusion (4 h). The test item was administered dissolved in phosphate buffer saline (pH 7.4) and plasma concentrations were analyzed applying LC/MS/MS.

Animal studies have been approved by the appropriate animal welfare authority (Regierung von Oberbayern, Munich, Germany) and were conducted in compliance with European and federal guidelines for the use and care of laboratory animals.

Allometric scaling is a basic interspecies scaling method that employs in vivo pharmacokinetic parameters to extrapolate from animal models to human. As metabolism of cilengitide was shown to be minimal across species and the mode of administration is i.v., simple allometric scaling based on log body weight was considered sufficient and applied using the following equation:

Y = pharmacokinetic parameter (CL, Vdss); a = allometric coefficient; b = allometric exponent; W = body weight.

Half‐life values were estimated based upon the predicted primary parameters of clearance (CL) and volume of distribution at steady state (Vd_ss).