JDQ443, a Structurally Novel, Pyrazole-Based, Covalent Inhibitor of KRASG12C for the Treatment of Solid Tumors.

Rapid emergence of tumor resistance via RAS pathway reactivation has been reported from clinical studies of covalent KRASG12C inhibitors. Thus, inhibitors with broad potential for combination treatment and distinct binding modes to overcome resistance mutations may prove beneficial. JDQ443 is an investigational covalent KRASG12C inhibitor derived from structure-based drug design followed by extensive optimization of two dissimilar prototypes. JDQ443 is a stable atropisomer containing a unique 5-methylpyrazole core and a spiro-azetidine linker designed to position the electrophilic acrylamide for optimal engagement with KRASG12C C12. A substituted indazole at pyrazole position 3 results in novel interactions with the binding pocket that do not involve residue H95. JDQ443 showed PK/PD activity in vivo and dose-dependent antitumor activity in mouse xenograft models. JDQ443 is now in clinical development, with encouraging early phase data reported from an ongoing Phase Ib/II clinical trial (NCT04699188).

An Alternative Thiol-Reactive Dye to Analyze Ligand Interactions with the Chemokine Receptor CXCR2 Using a New Thermal Shift Assay Format.

Integral membrane proteins (IMPs) play an important role in many cellular events and are involved in numerous pathological processes. Therefore, understanding the structure and function of IMPs is a crucial prerequisite to enable successful targeting of these proteins with low molecular weight (LMW) ligands early on in the discovery process. To optimize IMP purification/crystallization and to identify/characterize LMW ligand-target interactions, robust, reliable, high-throughput, and sensitive biophysical methods are needed. Here, we describe a differential scanning fluorimetry (DSF) screening method using the thiol-reactive BODIPY FL-cystine dye to monitor thermal unfolding of the G-protein-coupled receptor (GPCR), CXCR2. To validate this method, the seven-transmembrane protein CXCR2 was analyzed with a set of well-characterized antagonists. This study showed that the new DSF assay assessed reliably the stability of CXCR2 in a 384-well format. The analysis of 14 ligands with a potency range over 4 log units demonstrated the detection/characterization of LMW ligands binding to the membrane protein target. Furthermore, DSF results cross-validated with the label-free differential static light scattering (DSLS) thermal denaturation method. These results underline the potential of the BODIPY assay format as a general tool to investigate membrane proteins and their interaction partners.