Mechanistic Insights of a p53-Targeting Small Molecule.

Area of Science:

• Oncology
• Pharmacology
• Molecular Biology

Background:

• Restoring wild-type (wt) or mutant (mut) p53 is a key strategy for cancer therapy.
• The tryptophanol-derived oxazoloisoindolinone scaffold has yielded promising p53 reactivators.
• RVJB59 is a potent (R)-tryptophanol-derived oxazoloisoindolinone with enhanced activity and selectivity over SLMP53-1.

Purpose of the Study:

• To investigate the pharmacokinetic profile and mechanism of action of RVJB59.
• To evaluate the antiproliferative activity of RVJB59 and its metabolites.
• To assess the interaction of RVJB59 with p53 and its in vivo efficacy.

Main Methods:

• In vitro metabolic degradation studies using human liver microsomes and rat liver S9 fractions (LC/HRMS/MS).
• Synthesis and antiproliferative evaluation of RVJB59 metabolites against HCT116 cells.
• Differential Scanning Calorimetry (DSF) studies on p53 thermostability.
• Covalent binding assays using glutathione and wt p53 (HRMS/MS).
• In vitro 3D spheroid and in vivo chicken embryo models.

Main Results:

• RVJB59 exhibits low in vitro clearance and its main metabolites show reduced antiproliferative activity.
• RVJB59 enhances the thermostability of wt and R273H p53 DNA-binding domains, with selective binding to Cys141 of wt p53.
• RVJB59 reduces 3D tumor spheroid growth and demonstrates antiangiogenic potential in vivo.

Conclusions:

• RVJB59 is a low-clearance compound with a favorable pharmacokinetic profile.
• RVJB59 selectively targets and stabilizes p53, leading to anti-cancer effects.
• RVJB59 shows significant potential as a p53-activating therapeutic agent for cancer treatment.