System-wide profiling by proteome integral solubility alteration assay of drug residence times for target characterization

Abstract

Most drugs used in the clinic and drug candidates target multiple proteins, and thus detailed characterization of their efficacy targets is required. While current methods rely on quantitative measurements at thermodynamic equilibrium, kinetic parameters such as the residence time of a drug on its target provide a better proxy for efficacy in vivo. Here, we present Residence Time Proteome Integral Solubility Alteration (ResT-PISA) assay which provides monitoring temporal protein solubility profiles after drug removal (off-curve) in cell lysate or intact cells, quantifying the lifetime of drug-target interaction. A compressed version of the assay measures the integral under the off-curve enabling the multiplexing of binding affinity and residence time assessments into a single proteomic analysis. We introduce a combined scoring system for three parametric dimensions to improve prioritization of targets. By providing complementary information to other characteristics of drug-target interaction, ResT-PISA approach will be useful in drug development and precision medicine.

Merging Multi-OMICs with Proteome Integral Solubility Alteration Unveils Antibiotic Mode of Action

“…6(D)). Regarding the validation of the pre-defined target like flavodoxin, it is paramount to mention that some targets exhibit very limited shifts in solubility changes upon drug binding, 12,30,31 making the detection of the target difficult at a statistically significant level. In this regard, we have observed shifts in the solubility of flavodoxin for all the compounds, however, only in the case of compound IVj the change was statistically significant (Fig.…”