Evaluation of serum protein binding by using in vitro pharmacological activity for the effective pharmacokinetics profiling in drug discovery.

The establishment of a new index for the profile of serum protein binding was analyzed theoretically. The in vitro pharmacological activity ratio of the inhibition constant in the absence of serum protein to that in its presence (activity ratio), which represents the extent of specific binding to serum protein, was suggested as the new index. To clarify the usefulness of the activity ratio, theoretical analysis by the activity ratio for 3% human serum albumin was examined in comparison with conventional methods of equilibrium dialysis. In-house very late antigen-4 antagonists were used as model compounds, whose pharmacokinetics were strongly influenced by serum protein binding. Although the theoretical and actual unbound fractions were similar, the latter tended to be slightly lower than the former. This small difference was considered to correspond to nonspecific binding. These results suggested that the specific and nonspecific binding could be discriminated by comparing the activity ratio data with those of conventional methods. Moreover, the activity ratio was suggested to be useful in profiling the influence of protein binding on pharmacokinetics. In conclusion, it was considered that the activity ratio could avoid the risk of misleading interpretation by nonspecific binding in pharmacokinetics/pharmacological activity. Moreover, the activity ratio was considered to be valuable as one of the useful parameters in pharmacokinetics profiling and as a tool of rational drug design for drug discovery.