The Plasma Inhibitory Assay is a tool for measuring the efficacy of inhibitors in the presence of plasma protein
One factor determining the bioavailability is the impact of plasma proteins binding to a drug, potentially rendering it ineffective. We needed to measure the plasma inhibitory activity (PIA) on FLT3 inhibitors for a drug discovery study.
We have employed our FLT3 Cellular Phosphorylation Assays to determine the efficacy of two FLT3 inhibitors, Sorafenib and Sunitinib, in the presence and absence of plasma proteins. In an in vitro study, we incubated the inhibitors with either a plasma-free medium or a plasma-containing medium before subjecting them to the cellular phosphorylation assay with the readout of FLT3 autophosphorylation via ELISA. The reference inhibitors are examples of inhibitors being susceptible (Sorafenib) and unsusceptible (Sunitinib) to plasma binding.
The determination of bioavailability in mice is performed similarly: At different time points after drug administration blood samples are taken and the effective compound concentration will be measured via the Cellular Phosphorylation Assay. The PIA assay is used in clinical trials as a surrogate for pharmacodynamic analysis of FLT3 inhibitors in patients.
Determination of the Plasma Inhibitory Activity on Drugs based on the Cellular Phosphorylation Assay.