A method to quantitate the neutralizing capacity of anti-therapeutic protein antibodies in serum and their correlation to clinical impact

Abstract A robust, quantitative method for assessing the neutralizing capacity of anti-therapeutic protein antibodies was developed and tested using 4 analytical assay platforms typically used for detection of anti-drug neutralizing antibodies. The method described here utilized titration of increasing concentrations of therapeutic protein into serum containing anti-therapeutic protein antibodies, either positive control antibodies or clinical samples. Neutralizing capacities were calculated by determining the EC 50 from the titration curves. The neutralizing capacity of purified anti therapeutic protein antibodies was expressed in terms of “μg of drug neutralized per μg of anti-TP antibody” present. In the case of serum originating from clinical study subjects, the neutralizing capacity of the samples was expressed as “μg of drug neutralized per mL of serum”. A relative shift in EC 50 values was observed as the amount of serum or antibody was changed resulting in a proportional shift of the calculated neutralizing capacity. Application of this approach using different assay platforms was consistent providing evidence for its potential to be a useful approach to characterize, qualify and compare neutralizing positive control antibody preparations or clinical samples. Using this methodology, we were able to draw a clear correlation between the neutralizing capacity and the effect of these antibodies on a clinical pharmacodynamic (PD) marker. Determination of the neutralizing capacity of antibody positive samples from subjects in a clinical study indicated direct correlation of pharmacodynamic results with non-response, partial response and response to high, mid and low neutralizing capacities respectively.