KATP channel mutations in congenital hyperinsulinism: Progress and challenges towards mechanism-based therapies.

Congenital hyperinsulinism (CHI) is the most common cause of persistent hypoglycemia in infancy/childhood and is a serious condition associated with severe recurrent attacks of hypoglycemia due to dysregulated insulin secretion. Timely diagnosis and effective treatment are crucial to prevent severe hypoglycemia that may lead to life-long neurological complications. In pancreatic ß-cells, adenosine triphosphate (ATP)-sensitive K+ (KATP) channels are a central regulator of insulin secretion vital for glucose homeostasis. Genetic defects that lead to loss of expression or function of KATP channels are the most common cause of HI (KATP-HI). Much progress has been made in our understanding of the molecular genetics and pathophysiology of KATP-HI in the past decades; however, treatment remains challenging, in particular for patients with diffuse disease who do not respond to the KATP channel activator diazoxide. In this review, we discuss current approaches and limitations on the diagnosis and treatment of KATP-HI, and offer perspectives on alternative therapeutic strategies.

Non-coding variants disrupting a tissue-specific regulatory element in HK1 cause congenital hyperinsulinism.

Gene expression is tightly regulated, with many genes exhibiting cell-specific silencing when their protein product would disrupt normal cellular function1. This silencing is largely controlled by non-coding elements, and their disruption might cause human disease2. We performed gene-agnostic screening of the non-coding regions to discover new molecular causes of congenital hyperinsulinism. This identified 14 non-coding de novo variants affecting a 42-bp conserved region encompassed by a regulatory element in intron 2 of the hexokinase 1 gene (HK1). HK1 is widely expressed across all tissues except in the liver and pancreatic beta cells and is thus termed a 'disallowed gene' in these specific tissues. We demonstrated that the variants result in a loss of repression of HK1 in pancreatic beta cells, thereby causing insulin secretion and congenital hyperinsulinism. Using epigenomic data accessed from public repositories, we demonstrated that these variants reside within a regulatory region that we determine to be critical for cell-specific silencing. Importantly, this has revealed a disease mechanism for non-coding variants that cause inappropriate expression of a disallowed gene.

CRISPR/Cas9 ADCY7 Knockout Stimulates the Insulin Secretion Pathway Leading to Excessive Insulin Secretion.

Aim: Despite the enormous efforts to understand Congenital hyperinsulinism (CHI), up to 50% of the patients are genetically unexplained. We aimed to functionally characterize a novel candidate gene in CHI. Patient: A 4-month-old boy presented severe hyperinsulinemic hypoglycemia. A routine CHI genetic panel was negative. Methods: A trio-based whole-exome sequencing (WES) was performed. Gene knockout in the RIN-m cell line was established by CRISPR/Cas9. Gene expression was performed using real-time PCR. Results: Hyperinsulinemic hypoglycemia with diffuse beta-cell involvement was demonstrated in the patient, who was diazoxide-responsive. By WES, compound heterozygous variants were identified in the adenylyl cyclase 7, ADCY7 gene p.(Asp439Glu) and p.(Gly1045Arg). ADCY7 is calcium-sensitive, expressed in beta-cells and converts ATP to cAMP. The variants located in the cytoplasmic domains C1 and C2 in a highly conserved and functional amino acid region. RIN-m(-/-Adcy7) cells showed a significant increase in insulin secretion reaching 54% at low, and 49% at high glucose concentrations, compared to wild-type. In genetic expression analysis Adcy7 loss of function led to a 34.1-fold to 362.8-fold increase in mRNA levels of the insulin regulator genes Ins1 and Ins2 (p ≤ 0.0002), as well as increased glucose uptake and sensing indicated by higher mRNA levels of Scl2a2 and Gck via upregulation of Pdx1, and Foxa2 leading to the activation of the glucose stimulated-insulin secretion (GSIS) pathway. Conclusion: This study identified a novel candidate gene, ADCY7, to cause CHI via activation of the GSIS pathway.

Transient neonatal hyperinsulinism: early predictors of duration.

OBJECTIVES: Hyperinsulinism is the most common cause of recurrent hypoglycemia in infants, with transient and permanent forms. Currently, there are no effective tools to predict severity and time to resolution in infants with transient hyperinsulinism (tHI). Therefore, our objective was to assess whether early glucose trends predict disease duration in tHI. METHODS: A retrospective, pilot cohort of infants admitted with tHI was phenotyped for clinical and laboratory parameters. Blood glucose (BG) values were collected from the first documented hypoglycemia for 120 h (five days). RESULTS: In 27 neonates with tHI, the presence of fetal distress (p=0.001) and higher mean daily BG (p=0.035) were associated with shorter time to resolution of hypoglycemia. In a further sensitivity analysis that grouped the cohort by the presence or absence of fetal distress, we found that in neonates without fetal distress, lower mean daily glucose was associated with longer disease duration (R2=0.53, p=0.01). CONCLUSIONS: Our pilot data suggests that predictors for disease duration of tHI may be elicited in the first week of life, and that tHI associated with fetal distress may represent a distinct clinical entity with a shorter time course.

Biphasic dynamics of beta cell mass in a mouse model of congenital hyperinsulinism: implications for type 2 diabetes.

AIMS/HYPOTHESIS: Acute hyperglycaemia stimulates pancreatic beta cell proliferation in the mouse whereas chronic hyperglycaemia appears to be toxic. We hypothesise that this toxic effect is mediated by increased beta cell workload, unrelated to hyperglycaemia per se. METHODS: To test this hypothesis, we developed a novel mouse model of cell-autonomous increased beta cell glycolytic flux caused by a conditional heterozygous beta cell-specific mutation that activates glucokinase (GCK), mimicking key aspects of the rare human genetic disease GCK-congenital hyperinsulinism. RESULTS: In the mutant mice, we observed random and fasting hypoglycaemia (random 4.5-5.4 mmol/l and fasting 3.6 mmol/l) that persisted for 15 months. GCK activation led to increased beta cell proliferation as measured by Ki67 expression (2.7% vs 1.5%, mutant and wild-type (WT), respectively, p < 0.01) that resulted in a 62% increase in beta cell mass in young mice. However, by 8 months of age, mutant mice developed impaired glucose tolerance, which was associated with decreased absolute beta cell mass from 2.9 mg at 1.5 months to 1.8 mg at 8 months of age, with preservation of individual beta cell function. Impaired glucose tolerance was further exacerbated by a high-fat/high-sucrose diet (AUC 1796 vs 966 mmol/l × min, mutant and WT, respectively, p < 0.05). Activation of GCK was associated with an increased DNA damage response and an elevated expression of Chop, suggesting metabolic stress as a contributor to beta cell death. CONCLUSIONS/INTERPRETATION: We propose that increased workload-driven biphasic beta cell dynamics contribute to decreased beta cell function observed in long-standing congenital hyperinsulinism and type 2 diabetes.

SUR1-mutant iPS cell-derived islets recapitulate the pathophysiology of congenital hyperinsulinism.

AIMS/HYPOTHESIS: Congenital hyperinsulinism caused by mutations in the KATP-channel-encoding genes (KATPHI) is a potentially life-threatening disorder of the pancreatic beta cells. No optimal medical treatment is available for patients with diazoxide-unresponsive diffuse KATPHI. Therefore, we aimed to create a model of KATPHI using patient induced pluripotent stem cell (iPSC)-derived islets. METHODS: We derived iPSCs from a patient carrying a homozygous ABCC8V187D mutation, which inactivates the sulfonylurea receptor 1 (SUR1) subunit of the KATP-channel. CRISPR-Cas9 mutation-corrected iPSCs were used as controls. Both were differentiated to stem cell-derived islet-like clusters (SC-islets) and implanted into NOD-SCID gamma mice. RESULTS: SUR1-mutant and -corrected iPSC lines both differentiated towards the endocrine lineage, but SUR1-mutant stem cells generated 32% more beta-like cells (SC-beta cells) (64.6% vs 49.0%, p = 0.02) and 26% fewer alpha-like cells (16.1% vs 21.8% p = 0.01). SUR1-mutant SC-beta cells were 61% more proliferative (1.23% vs 0.76%, p = 0.006), and this phenotype could be induced in SUR1-corrected cells with pharmacological KATP-channel inactivation. The SUR1-mutant SC-islets secreted 3.2-fold more insulin in low glucose conditions (0.0174% vs 0.0054%/min, p = 0.0021) and did not respond to KATP-channel-acting drugs in vitro. Mice carrying grafts of SUR1-mutant SC-islets presented with 38% lower fasting blood glucose (4.8 vs 7.7 mmol/l, p = 0.009) and their grafts failed to efficiently shut down insulin secretion during induced hypoglycaemia. Explanted SUR1-mutant grafts displayed an increase in SC-beta cell proportion and SC-beta cell nucleomegaly, which was independent of proliferation. CONCLUSIONS/INTERPRETATION: We have created a model recapitulating the known pathophysiology of KATPHI both in vitro and in vivo. We have also identified a novel role for KATP-channel activity during human islet development. This model will enable further studies for the improved understanding and clinical management of KATPHI without the need for primary patient tissue.

Possible New Strategies for the Treatment of Congenital Hyperinsulinism.

Objective: Congenital hyperinsulinism (CHI) is a rare disease characterized by persistent hypoglycemia as a result of inappropriate insulin secretion, which can lead to irreversible neurological defects in infants. Poor efficacy and strong adverse effects of the current medications impede successful treatment. The aim of the study was to investigate new approaches to silence ß-cells and thus attenuate insulin secretion. Research Design and Methods: In the scope of our research, we tested substances more selective and more potent than the gold standard diazoxide that also interact with neuroendocrine ATP-sensitive K+ (KATP) channels. Additionally, KATP channel-independent targets as Ca2+-activated K+ channels of intermediate conductance (KCa3.1) and L-type Ca2+ channels were investigated. Experiments were performed using human islet cell clusters isolated from tissue of CHI patients (histologically classified as pathological) and islet cell clusters obtained from C57BL/6N (WT) or SUR1 knockout (SUR1-/-) mice. The cytosolic Ca2+ concentration ([Ca2+]c) was used as a parameter for the pathway regulated by electrical activity and was determined by fura-2 fluorescence. The mitochondrial membrane potential (ΔΨ) was determined by rhodamine 123 fluorescence and single channel currents were measured by the patch-clamp technique. Results: The selective KATP channel opener NN414 (5 µM) diminished [Ca2+]c in isolated human CHI islet cell clusters and WT mouse islet cell clusters stimulated with 10 mM glucose. In islet cell clusters lacking functional KATP channels (SUR1-/-) the drug was without effect. VU0071063 (30 µM), another KATP channel opener considered to be selective, lowered [Ca2+]c in human CHI islet cell clusters. The compound was also effective in islet cell clusters from SUR1-/- mice, showing that [Ca2+]c is influenced by additional effects besides KATP channels. Contrasting to NN414, the drug depolarized ΔΨ in murine islet cell clusters pointing to severe interference with mitochondrial metabolism. An opener of KCa3.1 channels, DCEBIO (100 µM), significantly decreased [Ca2+]c in SUR1-/- and human CHI islet cell clusters. To target L-type Ca2+ channels we tested two already approved drugs, dextromethorphan (DXM) and simvastatin. DXM (100 µM) efficiently diminished [Ca2+]c in stimulated human CHI islet cell clusters as well as in stimulated SUR1-/- islet cell clusters. Similar effects on [Ca2+]c were observed in experiments with simvastatin (7.2 µM). Conclusions: NN414 seems to provide a good alternative to the currently used KATP channel opener diazoxide. Targeting KCa3.1 channels by channel openers or L-type Ca2+ channels by DXM or simvastatin might be valuable approaches for treatment of CHI caused by mutations of KATP channels not sensitive to KATP channel openers.

Activation of Protein Kinase A (PKA) signaling mitigates congenital hyperinsulinism associated hypoglycemia in the Sur1-/- mouse model.

There is a significant unmet need for a safe and effective therapy for the treatment of children with congenital hyperinsulinism. We hypothesized that amplification of the glucagon signaling pathway could ameliorate hyperinsulinism associated hypoglycemia. In order to test this we evaluated the effects of loss of Prkar1a, a negative regulator of Protein Kinase A in the context of hyperinsulinemic conditions. With reduction of Prkar1a expression, we observed a significant upregulation of hepatic gluconeogenic genes. In wild type mice receiving a continuous infusion of insulin by mini-osmotic pump, we observed a 2-fold increase in the level of circulating ketones and a more than 40-fold increase in Kiss1 expression with reduction of Prkar1a. Loss of Prkar1a in the Sur1-/- mouse model of KATP hyperinsulinism significantly attenuated fasting induced hypoglycemia, decreased the insulin/glucose ratio, and also increased the hepatic expression of Kiss1 by more than 10-fold. Together these data demonstrate that amplification of the hepatic glucagon signaling pathway is able to rescue hypoglycemia caused by hyperinsulinism.

Hyperinsulinemic hypoglycemia in children and adolescents: Recent advances in understanding of pathophysiology and management.

Hyperinsulinemic hypoglycemia (HH) is characterized by unregulated insulin release, leading to persistently low blood glucose concentrations with lack of alternative fuels, which increases the risk of neurological damage in these patients. It is the most common cause of persistent and recurrent hypoglycemia in the neonatal period. HH may be primary, Congenital HH (CHH), when it is associated with variants in a number of genes implicated in pancreatic development and function. Alterations in fifteen genes have been recognized to date, being some of the most recently identified mutations in genes HK1, PGM1, PMM2, CACNA1D, FOXA2 and EIF2S3. Alternatively, HH can be secondary when associated with syndromes, intra-uterine growth restriction, maternal diabetes, birth asphyxia, following gastrointestinal surgery, amongst other causes. CHH can be histologically characterized into three groups: diffuse, focal or atypical. Diffuse and focal forms can be determined by scanning using fluorine-18 dihydroxyphenylalanine-positron emission tomography. Newer and improved isotopes are currently in development to provide increased diagnostic accuracy in identifying lesions and performing successful surgical resection with the ultimate aim of curing the condition. Rapid diagnostics and innovative methods of management, including a wider range of treatment options, have resulted in a reduction in co-morbidities associated with HH with improved quality of life and long-term outcomes. Potential future developments in the management of this condition as well as pathways to transition of the care of these highly vulnerable children into adulthood will also be discussed.

The Effectiveness of Sirolimus Treatment in Two Rare Disorders with Nonketotic Hypoinsulinemic Hypoglycemia: The Role of mTOR Pathway

Nonketotic-hypoinsulinemic hypoglycemia (NkHH) is a very rare problem charcterized by increase in glucose consumption without hyperinsulinism. This disorder has mainly been reported in cases with AKT2 mutation and rarely in cases with PTEN mutation. In cases with PTEN or AKT2 mutation, there is no effective therapy other than frequent feeding to counter hypoglycemia. The mammalian target of rapamicin (mTOR) inhibitor, sirolimus, has been used in hyperinsulinemic hypoglycemia that was unresponsive to other medical treatment. In the insulin signaling pathway, both AKT2 and PTEN function upstream of mTOR. However, the role of Sirolimus on hypoglycemia in AKT2 and PTEN mutations is unknown. Case 1: Six month-old female with AKT2 mutation [c.49G>A (p.E17K)] and evidence of NkHH. Frequent feeding was unsuccesful in correcting hypoglycemia and her proptosis continued to worsen. Sirolimus treatment was started at three years of age. Subsequently, blood glucose (BG) levels increased to normal levels. Case 2: In a male with PTEN mutation (p.G132V (c.395G>T), persistent NkHH started at 16 years of age (fasting BG: 27 mg/dL, fasting insulin 1.5 mmol/L, while ketone negative). Sirolimus treatment was started and hypoglycemia was succesfully controlled. NkHH is a very rare and serious disorder which is challenging, both for diagnosis and treatment. Additionally, AKT2 and PTEN mutations may result in NkHH. Sirolimus treatment, through mTOR inhibition, appeared to be effectively controlling the peristent hypoglycemia and may be a life-saving therapy in this NkHH due to AKT2 and PTEN mutations.

Chronic stimulation induces adaptive potassium channel activity that restores calcium oscillations in pancreatic islets in vitro.

Insulin pulsatility is important to hepatic response in regulating blood glucose. Growing evidence suggests that insulin-secreting pancreatic ß-cells can adapt to chronic disruptions of pulsatility to rescue this physiologically important behavior. We determined the time scale for adaptation and examined potential ion channels underlying it. We induced the adaptation both by chronic application of the ATP-sensitive K+ [K(ATP)] channel blocker tolbutamide and by application of the depolarizing agent potassium chloride (KCl). Acute application of tolbutamide without pretreatment results in elevated Ca2+ as measured by fura-2AM and the loss of endogenous pulsatility. We show that after chronic exposure to tolbutamide (12-24 h), Ca2+ oscillations occur with subsequent acute tolbutamide application. The same experiment was conducted with potassium chloride (KCl) to directly depolarize the ß-cells. Once again, following chronic exposure to the cell stimulator, the islets produced Ca2+ oscillations when subsequently exposed to tolbutamide. These experiments suggest that it is the chronic stimulation, and not tolbutamide desensitization, that is responsible for the adaptation that rescues oscillatory ß-cell activity. This compensatory response also causes islet glucose sensitivity to shift rightward following chronic tolbutamide treatment. Mathematical modeling shows that a small increase in the number of K(ATP) channels in the membrane is one adaptation mechanism that is compatible with the data. To examine other compensatory mechanisms, pharmacological studies provide support that Kir2.1 and TEA-sensitive channels play some role. Overall, this investigation demonstrates ß-cell adaptability to overstimulation, which is likely an important mechanism for maintaining glucose homeostasis in the face of chronic stimulation.

Congenital hyperinsulinism disorders: Genetic and clinical characteristics.

Congenital hyperinsulinism (HI) is the most frequent cause of persistent hypoglycemia in infants and children. Delays in diagnosis and initiation of appropriate treatment contribute to a high risk of neurocognitive impairment. HI represents a heterogeneous group of disorders characterized by dysregulated insulin secretion by the pancreatic beta cells, which in utero, may result in somatic overgrowth. There are at least nine known monogenic forms of HI as well as several syndromic forms. Molecular diagnosis allows for prediction of responsiveness to medical treatment and likelihood of surgically-curable focal hyperinsulinism. Timely genetic mutation analysis has thus become standard of care. However, despite significant advances in our understanding of the molecular basis of this disorder, the number of patients without an identified genetic diagnosis remains high, suggesting that there are likely additional genetic loci that have yet to be discovered.

Clinical characteristics and phenotype-genotype review of 25 Omani children with congenital hyperinsulinism in infancy. A one-decade single-center experience.

OBJECTIVES: To report the genotype-phenotype characteristics, demographic features and clinical outcome of Omani patients with congenital hyperinsulinism (CHI). Methods: We retrospectively analyzed the clinical, biochemical, genotypical, phenotypical characteristics and outcomes of  children with CHI who were presented to the pediatric endocrine team in the Royal Hospital, Muscat, Oman between January 2007 and December 2016. Results: Analysis of 25 patients with CHI genetically revealed homozygous mutation in ABCC8 in 23 (92%) patients and 2 patients (8%) with compound heterozygous mutation in ABCC8. Fifteen (60%) patients underwent subtotal pancreatectomy as medical therapy failed and 2 (8%) patients showed response to medical therapy. Three patients expired during the neonatal period, 2 had cardiomyopathy and sepsis, and one had sepsis and severe metabolic acidosis. Out of the 15 patients who underwent pancreatectomy, 6 developed diabetes mellitus, 6 continued to have hypoglycemia and required medical therapy and one had pancreatic exocrine dysfunction post-pancreatectomy, following up with gastroenterology clinic and was placed on pancreatic enzyme supplements, while 2 patients continued to have hypoglycemia and both had abdominal MRI and 18-F-fluoro-L-DOPA positron emission tomography scan (PET-scan), that showed  persistent of the disease and started on medical therapy. Conclusion:  Mutation in ABCC8 is the most common cause of CHI and reflects the early age of presentation. There is a need for early diagnosis and appropriate therapeutic strategy.

Clinical and enzymatic phenotypes in congenital hyperinsulinemic hypoglycemia due to glucokinase-activating mutations: A report of two cases and a brief overview of the literature.

AIMS/INTRODUCTION: The principal aim of this study was to investigate the clinical, genetic and functional characteristics of two cases of congenital hyperinsulinism (CHI) caused by glucokinase (GCK) mutations in young patients. MATERIALS AND METHODS: Novel mutations were detected by CHI next-generation sequencing, and the kinetic parameters and thermal stability of recombinant wild-type and mutant glucokinase were determined in vitro. In addition, 18 naturally occurring GCK-CHI mutations reported previously were also summarized. RESULTS: A de novo mutation (M197V) was found in a 17-year-old male with an epilepsy history, whereas an autosomal dominant mutation (K90R) was found in a 20-year-old female with inherited asymptomatic hypoglycemia. Kinetic analysis showed increased enzyme activity for both mutants (RAI 4.7 for M197V and 1.6 for K90R) and enhanced thermal stability for the M197V mutant. However, of all the GCK-CHI mutants, the increase in enzyme activity (RAI between 1.6 and 130) did not correlate strongly with the severity of hypoglycemia. The de novo group (7/19) showed distinctive phenotypes from the autosomal dominant group (12/19), such as a higher proportion of diazoxide unresponsiveness (28.6% vs 0%), a higher incidence of macrosomia (85.7% vs 40%) and a rarer incidence of adulthood onset (0% vs 25%). CONCLUSIONS: The clinical phenotypes of GCK-CHIs were highly heterogeneous. We have identified two novel GCK-CHI mutations in young patients and investigated their pathogenicity by enzyme kinetic analysis, which expanded the spectrum of this rare disease.

Continuous Intragastric Dextrose: A Therapeutic Option for Refractory Hypoglycemia in Congenital Hyperinsulinism.

Feeding problems are frequent in infants with congenital hyperinsulinism (HI) and may be exacerbated by continuous enteral nutrition (EN) used to maintain euglycemia. Our center's HI team uses dextrose solution given continuously via gastric tube (intrasgastric dextrose, IGD) for infants not fully responsive to conventional medical therapy or pancreatectomy. Here, we describe our practice as well as growth, feeding, and adverse events in infants with HI exposed to IGD. METHODS: This was a retrospective cohort of infants with HI treated with IGD from 2009-2017. Primary outcomes were weight-for-length and body mass index Z-scores (WFL-Z and BMI-Z) in the year following IGD initiation. Secondary outcomes included EN use and adverse events. We used multivariable regression to assess covariates of interest. RESULTS: We studied 32 subjects (13 female) with a median age at IGD initiation of 73 days (range 17-367); median follow-up was 11.2 months (range 5.0-14.2). WFL-Z did not change significantly over time (p > 0.05). EN use decreased significantly over time, i.e., at 0 months: 72% (95% CI 53-85) vs. at 12 months 39% (95% CI 22-59). No potential adverse events led to discontinuation of IGD. CONCLUSIONS: Over a median follow-up of nearly 1 year, IGD was well-tolerated, with no change in WFL-Z or BMI-Z from baseline.

Short- and long-term consequences for offspring exposed to maternal diabetes: a review.

The prevalence of gestational diabetes mellitus is increasing, as is the worldwide prevalence of type 2 diabetes and obesity, even in children and adolescents. Exposure in utero to maternal diabetes carries several short-term consequences due mainly to maternal hyperglycemia, and consequent fetal hyperinsulinemia. Current evidence also supports the hypothesis that adult health and disease have developmental origins, and that disorders in early-life environments prompt metabolic imprinting that results in a greater risk of negative metabolic outcomes later in life. In particular, exposure in utero to maternal diabetes seems to influence long-term metabolic outcomes, carrying a higher risk of obesity and type 2 diabetes, and thus creating a vicious cycle for future generations. In this paper, the short- and long-term consequences of exposure in utero to hyperglycemia are reviewed, focusing particularly on the long-term metabolic consequences, and investigating the possible pathogenic mechanisms involved.

Congenital Hyperinsulinism in Infants with Turner Syndrome: Possible Association with Monosomy X and KDM6A Haploinsufficiency.

BACKGROUND: Previous case reports have suggested a possible association of congenital hyperinsulinism with Turner syndrome. OBJECTIVE: We examined the clinical and molecular features in girls with both congenital hyperinsulinism and Turner syndrome seen at The Children's Hospital of Philadelphia (CHOP) between 1974 and 2017. METHODS: Records of girls with hyperinsulinism and Turner syndrome were reviewed. Insulin secretion was studied in pancreatic islets and in mouse islets treated with an inhibitor of KDM6A, an X chromosome gene associated with hyperinsulinism in Kabuki syndrome. RESULTS: Hyperinsulinism was diagnosed in 12 girls with Turner syndrome. Six were diazoxide-unresponsive; 3 had pancreatectomies. The incidence of Turner syndrome among CHOP patients with hyperinsulinism (10 of 1,050 from 1997 to 2017) was 48 times more frequent than expected. The only consistent chromosomal anomaly in these girls was the presence of a 45,X cell line. Studies of isolated islets from 1 case showed abnormal elevated cytosolic calcium and heightened sensitivity to amino acid-stimulated insulin release; similar alterations were demonstrated in mouse islets treated with a KDM6A inhibitor. CONCLUSION: These results demonstrate a higher than expected frequency of Turner syndrome among children with hyperinsulinism. Our data suggest that haploinsufficiency for KDM6A due to mosaic X chromosome monosomy may be responsible for hyperinsulinism in Turner syndrome.

Whole-exome sequencing for mutation detection in pediatric disorders of insulin secretion: Maturity onset diabetes of the young and congenital hyperinsulinism.

BACKGROUND: To assess the utility of whole-exome sequencing (WES) for mutation detection in maturity-onset diabetes of the young (MODY) and congenital hyperinsulinism (CHI). MODY and CHI are the two commonest monogenic disorders of glucose-regulated insulin secretion in childhood, with 13 causative genes known for MODY and 10 causative genes identified for CHI. The large number of potential genes makes comprehensive screening using traditional methods expensive and time-consuming. METHODS: Ten subjects with MODY and five with CHI with known mutations underwent WES using two different exome capture kits (Nimblegen SeqCap EZ Human v3.0 Exome Enrichment Kit, Nextera Rapid Capture Exome Kit). Analysis was blinded to previously identified mutations, and included assessment for large deletions. The target capture of five exome capture technologies was also analyzed using sequencing data from >2800 unrelated samples. RESULTS: Four of five MODY mutations were identified using Nimblegen (including a large deletion in HNF1B). Although targeted, one mutation (in INS) had insufficient coverage for detection. Eleven of eleven mutations (six MODY, five CHI) were identified using Nextera Rapid (including the previously missed mutation). On reconciliation, all mutations concorded with previous data and no additional variants in MODY genes were detected. There were marked differences in the performance of the capture technologies. CONCLUSIONS: WES can be useful for screening for MODY/CHI mutations, detecting both point mutations and large deletions. However, capture technologies require careful selection.

Regulation of KATP Channel Trafficking in Pancreatic ß-Cells by Protein Histidine Phosphorylation.

Protein histidine phosphatase 1 (PHPT-1) is an evolutionarily conserved 14-kDa protein that dephosphorylates phosphohistidine. PHPT-1-/- mice were generated to gain insight into the role of PHPT-1 and histidine phosphorylation/dephosphorylation in mammalian biology. PHPT-1-/- mice exhibited neonatal hyperinsulinemic hypoglycemia due to impaired trafficking of KATP channels to the plasma membrane in pancreatic ß-cells in response to low glucose and leptin and resembled patients with congenital hyperinsulinism (CHI). The defect in KATP channel trafficking in PHPT-1-/- ß-cells was due to the failure of PHPT-1 to directly activate transient receptor potential channel 4 (TRPC4), resulting in decreased Ca2+ influx and impaired downstream activation of AMPK. Thus, these studies demonstrate a critical role for PHPT-1 in normal pancreatic ß-cell function and raise the possibility that mutations in PHPT-1 and/or TRPC4 may account for yet to be defined cases of CHI.

Morphoproteomics and biomedical analytics coincide with clinical outcomes in supporting a constant but variable role for the mTOR pathway in the biology of congenital hyperinsulinism of infancy.

We first introduced the concept of the mTOR pathway's involvement in congenital hyperinsulinism of infancy (CHI), based largely on morphoproteomic observations and clinical outcomes using sirolimus (rapamycin) as a therapeutic agent in infants refractory to octreotide and diazoxide treatment. Subsequent publications have verified the efficacy of such treatment in some cases but limited and variable in others. We present further evidence of a constant but variable role for the mTOR pathway in the biology of CHI and provide a strategy that allows for the short-term testing of sirolimus in individual CHI patients.