Endogenous effectors of human liver glycogen phosphorylase modulate effects of indole-site inhibitors

1 Metabolic Research Laboratory, Section of Endocrinology, Metabolism, and Nutrition, Minneapolis Veterans Affairs Medical Center and 5 Departments of Medicine and Food Science and Nutrition, University of Minnesota, Minneapolis, Minnesota; Departments of 2 Cardiovascular and Metabolic Diseases, 3 Molecular Sciences, and 4 Pharmacokinetics, Dynamics, and Metabolism, Pfizer Global Research and Development, Groton Laboratories, Groton, Connecticut Submitted 22 June 2004 ; accepted in final form 28 March 2005 Phosphorylase is regulated by a number of small-molecular-weight effectors that bind to three sites on the enzyme. Recently, a fourth site referred to as the indole-inhibitor site has been identified. Synthetic compounds bind to the site and inhibit activity. However, the effects of these compounds in the presence of other endogenous effectors are unknown. We have determined the effects of four indole derivative glycogen phosphorylase inhibitors (GPI) on recombinant human liver glycogen phosphorylase a activity. The GPIs tested were all potent inhibitors. However, the endogenous inhibitors (glucose, ADP, ATP, fructose 1-phosphate, glucose 6-phosphate, UDP-glucose) and the activator (AMP) markedly reduced the inhibitory effect of GPIs. Consistent with these in vitro findings, the IC 50 for the inhibition of glycogenolysis in cells and the liver drug concentration associated with glucose-lowering activity in diabetic ob / ob mice in vivo were also significantly higher than those determined in in vitro enzyme assays. The inhibitory effect of indole-site effectors is modulated by endogenous small-molecular-weight effectors of phosphorylase a activity. However, at higher concentrations (10–30 µM), the GPI effect was dominant and resulted in inhibition of phosphorylase a activity irrespective of the presence or absence of the other modulators of the enzyme. glucose Address for reprint requests and other correspondence: N. Ercan-Fang, Minneapolis VA Medical Center, One Veterans Dr. (111G), Minneapolis, MN 55417 (e-mail: ercan001{at}umn.edu )