(1025-B) Covalent inhibitors: how we can find them in an early stage of a drug discovery program

Abstract: Over the last ten years, advances in covalent drug discovery have led to the successful development and approval of drugs, including covalent of inhibitors of BTK, Mutant EGFR, KRAS(G12C) and SARS- CoV-2 Mpro. Nowadays, covalent inhibitors are no longer identified, as in the past, by serendipity: on the contrary, well-defined multidisciplinary approaches have been established that can pave the way to the successful identification of the compounds.
Multiple drug discovery programs focused on the development of first-in-class covalent inhibitors are currently ongoing at IRBM. In this poster, we describe the assay cascade that we established to identify novel covalent pharmacophores against an undisclosed target. The starting point was a homogenous assay used to monitor the enzymatic activity of the target protein and suitable to screen covalent fragment libraries. A mutant variant of the protein with the target cysteine residue converted into alanine was prepared: as the enzymatic activity of the mutant protein was only < 50% lower than the wt, an appropriate and target-relevant counter screen assay was developed. Confirmation of the binding specificity of the hit compounds was conducted by LC/MS analysis and peptide mapping.
Biochemical characterization of the more interesting molecules (i.e. time-dependency assay, progression curve to measure the Kobs) followed along with a cell-based target engagement assay are included in the screening cascade.
This approach led to the identification and characterization of hits and the transition to the hit expansion phase with an ongoing SAR.