Our goal is to find novel non-competitive inhibitors of phosphomannose isomerase (PMI) that can be used as a therapeutic for treating patients with Congenital Disorder of Glycosylation Type Ia (CDG-Ia). This is a rare autosomal recessive metabolic disorder with multisystemic symptoms. Patients have decreased activity of phosphomannomutase 2 (PMM2) that impair the conversion of Mannose-6-P (Man-6-P) to Mannose-1-P leading to defective N-glycosylation. There is no therapy for CDG-Ia patients and many die. CDG-Ib patients have PMI, and this enzyme mutated interconverts Man-6-P and fructose-6-P (F-6-P). However, dietary supplements of mannose completely reverse the symptoms of CDG-Ib patients, some of which overlap with the symptoms of CDG-Ia patients. Unfortunately, mannose does not help CDG-Ia patients because their normal PMI activity catabolizes mannose-derived Man-6-P; the ratio of PMI: PMM activity is simply too high to increase intracellular Man-6-P. We propose that CDG-Ia patients will benefit from dietary mannose if we simultaneously reduce PMI activity with a non-competitive inhibitor. This would allow a modest intracellular accumulation of Man-6-P and drive metabolic flux into the glycosylation pathway using the residual PMM2 activity. The purpose of this application is to identify PMI-specific, cell-permeable, non-competitive inhibitors of human PMI using the MLSCN facilities and resources, including the MLSMR compound collection. PMI assays are well-established, and will be adapted to fluorescent readout in 384-well plate format. Confirmed hits will be validated using live cell-based PMI assays that measure metabolic flux of mannose into glycosylation vs. glycolysis. A successful outcome would initially be the identification of a novel chemical probe, and ultimately to the development of potential therapeutics to be used in conjunction with mannose therapy for CDG-Ia patients, who have no other therapeutic options. ? ? ?
