Preclinical Development of GT-14, a Novel Inhibitor of Gαi2 Protein: Comprehensive Evaluation of Physicochemical, Metabolic Characteristics and Tissue Distribution

GT-14, identified as [(E)-4-((1-(1-methyl-1H-indol-2-yl) ethylidene)amino)phenol], is a novel inhibitor targeting the Gα_i2 protein, which is crucial in facilitating cell migration and invasion in prostate, ovarian, and breast cancer cells. therefore a valuable target for treating metastatic castration-resistant prostate cancer (mCRPC). In this study, GT-14's physicochemical properties, permeability, metabolic behavior, and tissue distribution were assessed. The results showed that GT-14 exhibited very slight aqueous solubility at room temperature (0.11 mg/mL) but was soluble in solvents including dimethyl sulfoxide and dimethyl acetamide, and sparingly or slightly soluble in several cosolvents. GT-14 exhibited a distinct pH-dependent solubility profile, being stable across a broad pH range (1.2-7.4) but degrading in strongly basic conditions. It exhibited high permeability (1.3 x 10^-5 cm/s) in Caco-2 cell culture models and therefore identified as a BCS II compound. Hepatic microsomal studies revealed that GT-14 underwent Phase I metabolism, with more than 90% remaining in 60 min incubation in rat liver microsomes. A stable co-solvent formulation was developed to enable intravenous administration for pharmacokinetic studies. Previous pharmacokinetic studies showed that GT-14 exhibited biphasic disposition with a terminal plasma elimination half-life of 268.07 minutes (> 4 hours). Tissue distribution analysis indicated the highest concentration of GT-14 in the prostate, followed by the kidneys, lungs, heart, and liver. Our study presents an early-stage preclinical drug development roadmap that integrates modern technologies for efficiency and success, using GT-14 as a model compound. It showed promising characteristics, reinforcing its potential as a new therapeutic agent for mCRPC.