Skip to main content

Protein binding

Following a drug's entrance into and its appearance in the systemic circulation, only the free and unbound fraction is capable of a pharmacological (or toxicological) effect. The extent of plasma protein binding can be determined in vitro or ex vivo by equilibrium dialysis; ultrafiltration or ultracentrifugation.
  • Regulatory compliance

  • Diverse methods

  • Analysis for efficacy and safety

The partitioning of a test article into erythrocytes vs. plasma can be determined in vitro or ex vivo in animal and human blood. Microsomal protein binding can be evaluated by equilibrium dialysis. Data describing the how the drug binds to plasma proteins and how blood-to-plasma partitioning works are required to aid in the evaluation of pharmacological, pharmacokinetic and toxicological data. (Also see SEND 3.1)

Regulatory considerations for plasma protein binding, blood to plasma partitioning and microsomal protein binding studies

Investigation of the plasma or microsomal protein unbound fraction of a test article is critical for characterizing the in vivo pharmacokinetic profile of dose, exposure, efficacy and safety margins in clinical pharmacology. Cross-species assessment of plasma protein binding is an IND requirement and detailed in the ICH M3(R2) guidance.

Methods

 

Deliverables

Plasma protein binding assays will identify the extent of test article binding, providing a percent bound and unbound, the time to dialysis equilibrium, and concentration dependence profile in human and other selected preclinical species. The blood-to-plasma partitioning will provide a partitioning coefficient. These data can then be used to calculate free fraction in plasma to allow correction of in vivo exposures to profile efficacy and safety margins.  

Microsomal protein binding assays will identify the extent of the test article binding to human liver microsomal proteins for correction of IC50 or other parameters to account for free drug fraction.