Applying Two Orthogonal Methods to Assess Accuracy of Plasma Protein Binding Measurements for Highly Bound Compounds

Significant advances have been made over the years to accurately measure plasma protein binding (PPB) of highly bound compounds. However, because of perceived uncertainty based on historical suboptimal methods and limitation of radiochemical purity of radiolabeled materials, current regulatory guidelines recommend using an arbitrary cutoff fraction unbound (fu) of 0.01 as the lower limit for drug-drug interaction (DDI) prediction. This can result in significant overprediction of DDI for highly bound compounds, unnecessary DDI clinical trials and more restrictive drug product labels. To build confidence in the accuracy of PPB measurement for highly bound compounds, 2 orthogonal methods, equilibrium dialysis and ultracentrifugation, are assessed in this study to measure PPB of 10 highly bound drugs (fu < 0.01). The results show that the 2 very different methods yield comparable fu values, generally within 2-fold of each other. The data suggest that PPB of highly bound compounds can be measured accurately using current state-of-art methods, and the experimental fu should be used for DDI prediction to provide a more realistic evaluation of DDI risk in the clinic.