Bis-Indole Derivatives as Dual Nuclear Receptor 4A1 (NR4A1) and NR4A2 Ligands.

Bis-indole derived compounds such as 1,1-bis(3'-indolyl)-1-(3,5-disubstitutedphenyl) methane (DIM-3,5) and the corresponding 4-hydroxyl analogs (DIM8-3,5) are NR4A1 ligands that act as inverse NR4A1 agonists and are potent inhibitors of tumor growth. The high potency of several DIM-3,5 analogs (IC50 < 1 mg/kg/day), coupled with the >60% similarity of the ligand-binding domains (LBDs) of NR4A1 and NR4A2 and the pro-oncogenic activities of both receptors lead us to hypothesize that these compounds may act as dual NR4A1 and NR4A2 ligands. Using a fluorescence binding assay, it was shown that 22 synthetic DIM8-3,5 and DIM-3,5 analogs bound the LBD of NR4A1 and NR4A2 with most KD values in the low µM range. Moreover, the DIM-3,5 and DIM8-3,5 analogs also decreased NR4A1- and NR4A2-dependent transactivation in U87G glioblastoma cells transfected with GAL4-NR4A1 or GAL4-NR4A2 chimeras and a UAS-luciferase reporter gene construct. The DIM-3,5 and DIM8-3,5 analogs were cytotoxic to U87 glioblastoma and RKO colon cancer cells and the DIM-3,5 compounds were more cytotoxic than the DIM8-3,5 compounds. These studies show that both DIM-3,5 and DIM8-3,5 compounds previously identified as NR4A1 ligands bind both NR4A1 and NR4A2 and are dual NR4A1/2 ligands.

Amino acid primed mTOR activity is essential for heart regeneration.

Heart disease is the leading cause of death with no method to repair damaged myocardium due to the limited proliferative capacity of adult cardiomyocytes. Curiously, mouse neonates and zebrafish can regenerate their hearts via cardiomyocyte de-differentiation and proliferation. However, a molecular mechanism of why these cardiomyocytes can re-enter cell cycle is poorly understood. Here, we identify a unique metabolic state that primes adult zebrafish and neonatal mouse ventricular cardiomyocytes to proliferate. Zebrafish and neonatal mouse hearts display elevated glutamine levels, predisposing them to amino-acid-driven activation of TOR, and that TOR activation is required for zebrafish cardiomyocyte regeneration in vivo. Through a multi-omics approach with cellular validation we identify metabolic and mitochondrial changes during the first week of regeneration. These data suggest that regeneration of zebrafish myocardium is driven by metabolic remodeling and reveals a unique metabolic regulator, TOR-primed state, in which zebrafish and mammalian cardiomyocytes are regeneration competent.