Example 9
To evaluate in vivo drug exposure and bioavailability, a single dose pharmacokinetic study in mice is performed. Romosozumab PARG (SEQ ID NO: 8) C-terminal variant is injected either intravenously (via tail vein) or subcutaneously at a dose of 1 mg/kg. Using nine animals per group, staggered sampling permits collection of data at a large number of time points without exceeding the maximum volume of blood that can be drawn from an individual animal. At each time point, 0.05 ml of blood is drawn. Animals 1 to 3 are sampled at 0.083, 24, 96 and 192 hours post-dose. Animals 4-6 are sampled at 1, 48, 168 and 240 hours. Animals 7-9 are sampled at 6, 72 and 192 hours. Serum is collected from the whole blood sample and test article concentration is determined by a binding immunoassay such as an ELISA (Enzyme-Linked ImmunoSorbant Assay). Changes in test article concentration over time can be used to calculate pharmacokinetic parameters via two compartment analysis. Parameters of interest include, but not limited to, area under the plasma concentration-time curve (AUC), half-life (t1/2) and clearance (CL) for each dose group. Bioavailability can be determined as the ratio of AUC for the subcutaneous dose to the AUC for the intravenous dose.
