DRUG DISCOVERY / DEVELOPMENT & DELIVERY a generic approach to the Validation of Small Molecule Lc-MS/MS Biomarker assays

In recent years, there has been a large increase in the use of both exploratory and validated biomarkers1, as they have been formally incorporated into the drug development process as indicators of the pharmacodynamic effect of drugs. a “biomarker” is a substance that is measured as an indicator of the normal biologic process, pathogenic process or pharmacologic response to a therapeutic intervention. However, while there is clear and comprehensive guidance on expectation of the approach and quality requirements for the generation of regulated pharmacokinetic or bioequivalence data2,3 this is not the case for biomarker data. Recent studies have seen the development of an approach to the validation of laboratory biomarker assays in support of drug development (Lee et al.). However, the focus has been on the use of ligand binding assays for the measurement of biomarker concentrations in ex vivo body fluids and tissues. The fact that biomarkers come in all shapes and sizes, and many are small molecules that lend themselves to the use of liquid chromatography – mass spectrometry (LC-MS), has been neglected. The traditional expectations for small molecule assays, in terms of performance, are significantly different from ligand binding assays for the generation of pharmacokinetic data5. Despite this, there is currently no published approach to validation of LC-MS/MS methods for endogenous small molecule biomarkers. Members of the pharmaceutical industry have struggled with endogenous assay validation for many years, and there is often confusion over which approach to take. The presence of endogenous analytes in a control matrix presents an added analytical challenge that must be overcome if small molecule biomarker assays are to be developed and characterised. There is therefore a need for a generic approach to endogenous assay validation that can be successfully applied in most cases of small molecule biomarker analysis. Some contract research organisations (CROs) are skilled in conducting this generic approach to small molecule biomarker validation of endogenous assays. As these CROs are extremely familiar with the method, they have the expertise to perform endogenous assay validation in the most effective way. Although there are no specific Food and Drug Administration (FDA) regulations for the validation of endogenous compounds, CROs performing endogenous assay validation are equipped to meet regulatory expectations and work with the scientific rigour required by such authorities. In this article, a generic approach to the validation of LC-MS/MS endogenous small molecule biomarkers methods is presented for consideration as the optimal bioanalytical method for this type of assay. The recommendation is to use a surrogate matrix over traditionally used controlled matrices. This generic approach is one that is likely to be successful in most cases, particularly when conducted by CROs who have the confidence and familiarity needed to perform it efficiently and correctly.