Extremes of Clinical and Enzymatic Phenotypes in Children With Hyperinsulinism Caused by Glucokinase Activating Mutations

OBJECTIVE

Heterozygous activating mutations of glucokinase have been reported to cause hypoglycemia attributable to hyperinsulinism in a limited number of families. We report three children with de novo glucokinase hyperinsulinism mutations who displayed a spectrum of clinical phenotypes corresponding to marked differences in enzyme kinetics.

RESEARCH DESIGN AND METHODS

Mutations were directly sequenced, and mutants were expressed as glutathionyl S-transferase–glucokinase fusion proteins. Kinetic analysis of the enzymes included determinations of stability, activity index, the response to glucokinase activator drug, and the effect of glucokinase regulatory protein.

RESULTS

Child 1 had an ins454A mutation, child 2 a W99L mutation, and child 3 an M197I mutation. Diazoxide treatment was effective in child 3 but ineffective in child 1 and only partially effective in child 2. Expression of the mutant glucokinase ins454A, W99L, and M197I enzymes revealed a continuum of high relative activity indexes in the three children (26, 8.9, and 3.1, respectively; wild type = 1.0). Allosteric responses to inhibition by glucokinase regulatory protein and activation by the drug RO0281675 were impaired by the ins454A but unaffected by the M197I mutation. Estimated thresholds for glucose-stimulated insulin release were more severely reduced by the ins454A than the M197I mutation and intermediate in the W99L mutation (1.1, 3.5, and 2.2 mmol/l, respectively; wild type = 5.0 mmol/l).

CONCLUSIONS

These results confirm the potency of glucokinase as the pancreatic β-cell glucose sensor, and they demonstrate that responsiveness to diazoxide varies with genotype in glucokinase hyperinsulinism resulting in hypoglycemia, which can be more difficult to control than previously believed.Hypoglycemia in infants with congenital hyperinsulinism has been associated with mutations that affect the regulation of insulin secretion by all three major classes of metabolic fuels: glucose, amino acids, and fatty acids (1–6). The most common of these disorders is caused by recessive mutations of the β-cell ATP-sensitive K⁺(K_ATP) channel; these mutations cause severe neonatal hypoglycemia that does not respond to medical therapy with diazoxide, a K_ATP channel agonist, and often requires near-total pancreatectomy (7,8). Other genetic forms of congenital hyperinsulinism, such as dominant mutations of glutamate dehydrogenase, cause less severe disease, with hypoglycemia that may not be recognized until childhood or even adult life and that responds well to diazoxide therapy (4,9–11). In 1998, the first case of hyperinsulinism caused by a dominant gain-of-function mutation of glucokinase was reported (12). This remains one of the rarest forms of hyperinsulinism, and information on its clinical and biochemical manifestations is limited because only a few cases have been reported subsequently (13–19). Most of these cases have been identified because of family histories of hypoglycemia with dominant patterns of transmission, and most affected individuals were reported to have relatively mild disease that could be managed medically with diazoxide.Glucokinase catalyzes the first step in glucose metabolism in pancreatic β-cells and liver (20). It exists as a monomer in three conformations that control catalytic function: a closed form, an open form, and a super open form (21). Transitions between these conformations are controlled by glucose concentration, giving a sigmoidal enzyme activity curve, as well as by allosteric modulators. Binding of novel glucokinase activator molecules, such as RO0281675, to the allosteric site increases glucokinase activity, resulting in both augmented hepatic glucose uptake and lowering of the β-cell threshold for glucose-stimulated insulin release (22). In the liver, glucokinase enzyme activity is inhibited by binding of glucokinase regulatory protein, which also leads to nuclear sequestration of the enzyme (23).Glucokinase serves a critical physiological function as the β-cell glucose sensor. It determines the glucose threshold for insulin release because of the low affinity of the enzyme for its substrate, glucose (half-maximal activity, S_0.5, occurs at 7.5 mmol/l glucose). Heterozygous mutations that reduce enzyme activity cause a subtype of maturity-onset diabetes of the young 2 (MODY2), whereas, as noted above, heterozygous activating mutations cause hypoglycemia. Expression of these activating mutations shows increased affinity for glucose with elevations of calculated enzyme activity indexes and lower calculated glucose thresholds for insulin release (24).Based on the initial cases reported, glucokinase hyperinsulinism has been assumed to be a mild form of hypoglycemia that can easily be managed medically. However, one reported case with a more severe clinical phenotype of uncontrollable hypoglycemia suggests that the range of manifestations of glucokinase hyperinsulinism may be greater than has been appreciated (14). The purpose of this report is to describe three children with hyperinsulinism caused by de novo glucokinase mutations who exhibit marked differences in responsiveness to medical therapy that correlate with differences in enzyme activity indexes.