Small Molecule Inhibitors of Constitutively Active Gsa

Abstract

Constitutive activation of Gsα is seen in the genetic disorder McCune‐Albright Syndrome, as well as some endocrine adenomas and in tissues affected by cholera toxin. Random mutagenesis of constitutively active Gα subunits in yeast identified three sites that, when mutated, suppressed the constitutive activity caused by R201H: F142, R231, and L266. Because R231 is on the surface of the protein, near the GTP‐binding site it was selected as a candidate site for drug targeting, with the goal of developing inhibitors of the constitutively active Gs alleles. Docking molecules in silico to the R231 binding pocket on the surface of Gs identified commercially available molecules with high potential to bind to the suppressor site. Eighteen molecules with high predicted binding affinity were tested for the ability to decrease the elevation in basal cAMP observed in HEK cells overexpressing the constitutively active Gs‐R201H allele. Two molecules, 6‐oxo‐N‐{[2‐(pyrrolidin‐1‐yl)pyridin‐4‐yl]methyl}‐1,6‐dihydropyridazine‐3‐carboxamide and N‐{[3‐(3‐methylphenyl)phenyl]methyl}‐6‐oxo‐1,6‐dihydropyridazine‐3‐carboxamide were able to significantly reduce cAMP levels by at least 10% from baseline. Neither of these molecules significantly reduced basal cAMP levels in cells transfected with wild‐type Gs constructs. These molecules will form the basis for further studies to develop antagonists of mutant Gs proteins associated with human disease.This work was Supported by NIH grant 1R15ED020190‐01