There has been an increasing focus on large molecule therapeutics and pharmaceutical companies have increasingly aligned their development pipelines in that direction. This has resulted in more biological therapeutics coming to market. In 2020, 15 of the 53 new approved drugs were large molecules.
Historically, bioanalytical support for large molecule development has utilized ligand binding technologies. LBA has been the platform of choice for pharmacokinetic, pharmacodynamic and anti-drug antibody analyses relying on the quality of reagents to help provide sensitive and selective assays.
However, focusing only on one technology can significantly hinder the bioanalytical strategy needed for today’s biologics. The improvement in mass spectrometric technology and growing focus on quantitation of large molecules using LC-MS/MS provides an ideal complement to the more established ligand-binding approaches.
We have long exploited the ability to deliver sensitive & specific assays for small molecules by taking advantage of an analyte’s chemical properties. Optimization of extraction, chromatography and mass spectrometric parameters make it possible to determine low concentrations of analytes in the presence of other matrix components. Combining the principles of ligand binding for target analyte enrichment with LC-MS/MS provides an alternative platform for quantitation of large molecules. The addition of an enzymatic digestion to produce a range of characteristic peptides provides surrogates that are more compatible with the mass range of many quantitative MS instruments.
LC-MS/MS provides significant advantages to quantitate large molecules. Hybrid mass spectrometry only requires 1 reagent for enrichment and this reagent does NOT need to be extremely selective. The detection and selectivity is provided by the mass spectrometer on the back end. It can readily differentiate the target molecule from potential interferences and can furthermore, differentiate similar analytes with simple sequence changes. Judicial selection of the surrogate peptides gives the ability to monitor specific parts of the molecule which can be useful where there are stability concerns. The MS assay can also be more easily multiplexed. Monitoring different peptides may also result in enhanced sensitivity.
KCAS is a strong advocate of assessing the relative merits of available technologies to answer a project’s needs and using that decision to drive project support. In the past, our ligand binding team provided the majority of our large molecule bioanalysis support. However, more recently, the power and selectivity of LC-MS/MS in large molecule bioanalysis is moving this technology into the mainstream for many PK and biomarker needs. This presentation will talk about the evolving role of LC-MS/MS in the bioanalysis of large molecule therapeutics and biomarkers, specifically where we see hybrid LCMS emerging as a front runner in generic/universal assays, as well as specific assays. We will discuss how we consider/decide which technology to pursue (LBA vs LCMS). We will also present case studies highlighting both.