OR05-2 Targeted Inhibition of Glutamate Dehydrogenase by Alpha-Tocopherol: A Potential Novel Treatment for Hyperinsulinism Hyperammonemia Syndrome

Congenital hyperinsulinism is a rare genetic disorder that causes severe hypoglycemia and can lead to permanent brain damage if inadequately controlled. Gain of function mutations in glutamate dehydrogenase (GDH) result in hyperinsulinism hyperammonemia (HI/HA) syndrome, the second most common cause of congenital hyperinsulinism. Current therapies inhibit insulin secretion but do not target GDH overactivity directly and thus fail to treat the spectrum of clinical manifestations observed, which in addition to hyperinsulinism and hyperammonemia, include seizures and developmental delays. The neurological symptoms are profound and occur independent of the hypoglycemia. We investigated the efficacy of alpha-tocopherol as a targeted GDH inhibitor in vitro and in vivo models of HI/HA syndrome. Human embryonic kidney (HEK293T) cells were transduced with lentivirus to overexpress either wild-type GDH or H454Y GDH, a well-characterized disease-causing mutant. GDH enzyme kinetics were determined spectrophotometrically in cell homogenates perfused with escalating concentrations of alpha-tocopherol on a background of 10mM glutamine. To evaluate the inhibitory effect of alpha-tocopherol on GDH in vivo, wild-type (WT, n=10) and H454Y GDH transgenic (Tg, n=13) adult mice were treated with oral alpha-tocopherol (75 μg/g body weight/dose) or vehicle twice daily for 3 doses and then fasted for 6 hours. Nadir fasting plasma glucoses were measured as a clinical correlate of GDH activity and compared between alpha-tocopherol and vehicle-treated wild-type and transgenic mice, respectively. Alpha-tocopherol effectively inhibited activity of wild-type and H454Y mutant GDH in HEK293T cells with 50% inhibitory concentration (IC50) of 4.1 and 3.1 μM, respectively. In vivo, nadir fasting plasma glucose was significantly higher in alpha-tocopherol-treated versus vehicle-treated WT and Tg mice (WT: mean nadir plasma glucose 94.3 ± 9.6 mg/dl v. 80.1 ± 9.6 mg/dl, p=0.003; Tg: mean nadir plasma glucose 71.9± 11.3 mg/dl v. 57.2 ± 12.6 mg/dl, p=0.002). Alpha-tocopherol effectively inhibits GDH in vitro and ameliorates fasting hypoglycemia in vivo in a mouse model of HI/HA syndrome. Based upon these findings, alpha-tocopherol is a promising potential treatment for HI/HA syndrome. Our next step will be to study the safety and efficacy of oral alpha-tocopherol supplementation in human subjects with HI/HA syndrome.