Covalent drug discovery efforts are growing rapidly but have major unaddressed limitations. These include high false positive rates during hit-to-lead identification
the inherent uncoupling of covalent drug concentration and effect [i.e., uncoupling of pharmacokinetics (PK) and pharmacodynamics (PD)]
and a lack of bioanalytical and modeling methods for determining PK and PD parameters. We present a covalent drug discovery workflow that addresses these limitations. Our bioanalytical methods are based upon a mass spectrometry (MS) assay that can measure the percentage of drug-target protein conjugation (% target engagement) in biological matrices. Further we develop an intact protein PK/PD model (iPK/PD) that outputs PK parameters (absorption and distribution) as well as PD parameters (mechanism of action, protein metabolic half-lives, dose, regimen, effect) based on time-dependent target engagement data. Notably, the iPK/PD model is applicable to any measurement (e.g., bottom-up MS and other drug binding studies) that yields % of target engaged. A Decision Tree is presented to guide researchers through the covalent drug development process. Our bioanalytical methods and the Decision Tree are applied to two approved drugs (ibrutinib and sotorasib)
the most common plasma off-target, human serum albumin
three protein targets (KRAS, BTK, SOD1), and to a promising SOD1-targeting ALS drug candidates.