Affinity and Kinetic Measurements Provide Insights of Biomolecular Complex Formation and Stability, Key Components of Drug-Target Interaction Strength
Detecting and measuring biomolecular interactions are critical in the discovery and selection of biotherapeutics leads.
Affinity (KD) and kinetics measurements (Kon, Koff) can benefit every stage of biopharmaceutical discovery and development processes. From target integrity validation, hit/lead selection, drug format diversification (e.g germlining, humanization, bispecific conversion, drug conjugation, effector function modulation …), binding kinetic data are required to define strength of the drug-target complex and to predict pharmacological drug response.
Antibody Drug Format Diversification
Overview of Affinity and Kinetic Measurements Applications
Drugs that exhibit similar affinities could exhibit diverse binding mechanisms and kinetics. Hence, the evaluation of affinity constants (KD) in conjunction with association (kon) and dissociation (koff) rates provides crucial information for molecule characterization. The affinity of an interaction directly affects the dose at which a biopharmaceutical is effective, and understanding mechanism of binding has implications in efficacy and desirability of a therapeutic candidate. Dissociation rate determines the duration of target occupancy. Long residence time is a good indicator of durable pharmacological response, allowing less frequent administration of lower doses and minimization of unwanted side effects.
Octet® BLI assays have become increasingly valuable for accurately determining affinity and kinetic constants. It highlights difference in on and off constant for each interaction even though equilibrium affinities are similar.
By bringing the detection surface directly to the sample and therefore eliminating the need for microfluidics, the approach using label-free and real-time analysis enables to streamline laboratory workflows and expedite assay development. It is extremely versatile in large molecules kinetics characterization and enables rapid assay optimization to allow the analysis of drug candidates ranging from antibody, antibody fragment, aptamer lead characterization to bispefic, multispefic and conjugated proteins reformating.
Therapeutic and Target Diversity for BLI Kinetic measurements
The concentration of sample in solution (analyte) has to be known and a dilution series of at least four concentrations is required to provide accurate KD values. Furthermore, Octet® BLI platform allows also direct measurement between crude samples pre-captured on biosensor and purified target in solution.
End point kinetic measurement (steady state analysis) is another approach that can be used for very fast binding interaction. It determines the concentration of analyte that gives 50% maximum signal when at equilibrium. This analysis does provide KD value but does not give any information on the binding rate constants of these interactions.
Single dose KD determination assay enables to provide approximate KD values that can be useful during screening campaigns of targets or drug variants (e.g epitope/paratope mapping, alanine variant scanning, affinity maturation, PTM removal…).
Besides the binding parameters measurements performed on therapeutics, characterization and quality control assays can also be conducted on targets. After purification or formulations steps, the functionality of targets, whether soluble (e.g ECD, enzyme, protein fragment…) or embedded in particles (e.g liposome, VLP, viral vector, nanodisc, nanoparticle…) can be estimated through functional characterization and qualitative analysis studies. These data are crucial for validating new constructs, purity, conformational or oligomeric state and active concentration of target in the sample.
BindingSolution offers versatile BLI binding measurement service platform to provide detailed insights into your product’s biological interactions.
BindingSolution’s expertise can help you to streamline and empower your drug discovery and development processes.
Our Analytical Binding Service Platform at a Glance
