Immunoassays for Measuring Serum Concentrations of Monoclonal Antibodies and Anti-biopharmaceutical Antibodies in Patients

Abstract

Monoclonal antibodies (mAbs) may be used as biopharmaceuticals to treat various diseases, ranging from oncology to inflammatory and cardiovascular affections. Trustworthy analytical methods are necessary to study their pharmacokinetics, both during their development and in post-marketing studies. Because biopharmaceuticals are macromolecules, ligand-binding assays (both immunoassays and bioassays) are methods of choice to measure their concentrations. Immunoassays are based on the capture of biopharmaceuticals by their target, which may be a circulating or membrane antigen or by an antibody recognizing their structure. Bioassays measure the activity of the biopharmaceutical in a specific in vitro test. A number of techniques have been reported, but their limits of detection and quantification vary widely. Anti-drug antibodies (ADA) against biopharmaceuticals are often formed and sometimes interfere with clinical efficacy. Accurate and reliable detection of ADA is therefore necessary. Binding of ADA is dependent on affinity and avidity, which makes quantification challenging. In this review, we discuss the benefits and limitations of each method to determine mAb levels and carefully compare ADA assays.

Overlooked Benefits of using Polyclonal Antibodies

“…Since pAbs are inherently more prone to changes over the duration of the immunization cycle, variations in performance of concern to regulatory agencies can be minimized or eliminated by choosing a mAb or rAb. Although pAbs are well suited for certain therapeutic applications as stated above, mAbs or rAbs would be a more desirable alternative when considering the potential for patients to develop anti-drug antibodies, as a single antibody, in contrast to a mixture of pAbs, would be easier to analyze for interference with clinical efficacy [41]. Similarly, mAbs have a long history of regulatory approval for therapeutics including for use as antibodydrug conjugates [42], as vaccines to rapidly emerging infectious diseases [43], to target multifactorial diseases [44], for antibody-dependent cell-mediated cytotoxicity [45], and many other clinical applications previously reviewed [4649].…”

Practical recommendations for the use of therapeutic drug monitoring of biopharmaceuticals in inflammatory diseases

“…Different methods have been developed: fluid-phase radioimmunoassay, solid-phase enzyme-linked immunosorbent assay (ELISA), reporter gene assay, enzyme immunoassay, homogenous mobility shift assay and liquid chromatography–tandem mass spectrometry 53,54,59,60. The most common format for quantifying biopharmaceuticals is ELISA, in which the biopharmaceutical is captured on a plate and detected using a secondary antibody 59. For measuring ADA, different assay formats are commercially available, in which the ADA are detected using the labeled biopharmaceutical itself 60…”

“…Only results obtained with the same assay can be compared. The implementation of a universal calibrator for quantifying ADA will eventually facilitate inter-laboratory harmonization of ADA measurements 59,64,65…”

Influence of Antigen Mass on the Pharmacokinetics of Therapeutic Antibodies in Humans

“…This description is based on: -Antibody concentration measurements which should reflect the concentration of antibody unbound to target. This is usually done by ELISA and/or HPLC-MS/MS techniques [172]; -Measurements of antigenic target unbound to mAb and, if possible, measurement of complexes or total target amounts. -Dense sampling strategies for both antibody and target amounts to ensure accurate estimates of TMDD model parameters, especially during the early phase, which is necessary to identify association (kon) and dissociation (koff) rates of mAb and targets, in addition to parameters of exchanges between central and peripheral compartiments.…”