Bis-Indole Derivatives as Dual Nuclear Receptor 4A1 (NR4A1) and NR4A2 Ligands
Srijana Upadhyay,
Amanuel Esayas Hailemariam,
Fuada Mariyam,
Zahin Hafiz,
Gregory Martin,
Jainish Kothari,
Evan Farkas,
Gargi Sivaram,
Logan Bell,
Ronald Tjalkens,
Stephen Safe
Affiliations
Srijana Upadhyay
Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA
Amanuel Esayas Hailemariam
Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA
Fuada Mariyam
Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA
Zahin Hafiz
Department of Chemical Engineering, Texas A&M University, College Station, TX 77843, USA
Gregory Martin
Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA
Jainish Kothari
Department of Chemical Engineering, Texas A&M University, College Station, TX 77843, USA
Evan Farkas
Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA
Gargi Sivaram
Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA
Logan Bell
Department of Chemistry, University of La Verne, La Verne, CA 91750, USA
Ronald Tjalkens
Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80526, USA
Stephen Safe
Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA
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 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.
