Patients with type Ia glycogen storage disease (GSD) have been surviving well into adulthood since continuous glucose therapy was introduced in the 1970s, and there have been many documented successful pregnancies in women with this condition. Historically, few individuals with type Ib GSD, however, survived into adulthood prior to the introduction of granulocyte colony stimulating factor (G-CSF) in the late 1980s. There are no previously published reports of pregnancies in GSD type Ib. In this case report we describe the course and management of five successful pregnancies in three patients with GSD type Ib. Patient 1 experienced an increase in glucose requirement in all three of her pregnancies, starting from the second trimester onwards. There were no major complications related to neutropenia except for oral ulcers. The infants did well, except for respiratory distress in two of them at birth. Patient 2 used cornstarch to maintain euglycemia, but precise dosing was not part of her regimen, and, hence, an increase in metabolic demands was difficult to demonstrate. She developed a renal calculus and urinary tract infection during her pregnancy and had chronic iron deficiency anemia but no neutropenia. The neonate did well after delivery. Patient 3 had poor follow-up during pregnancy. Increasing glucose requirements, worsening lipid profile, neutropenia associated with multiple infections, and anemia were noted. The newborn infant did well after delivery. In addition to the case reports, the challenges of the usage of G-CSF, the treatment of enterocolitis, and comparisons with the management of GSD Ia are discussed.
Infants with congenital hyperinsulinism owing to inactivating mutations in the KATP channel (KATPHI) who are unresponsive to medical therapy will require pancreatectomy to control the hypoglycemia. In preclinical studies, we showed that the GLP-1 receptor antagonist exendin-(9-39) suppresses insulin secretion and corrects fasting hypoglycemia in SUR-1−/− mice. The aim of this study was to examine the effects of exendin-(9-39) on fasting blood glucose in subjects with KATPHI. This was a randomized, open-label, two-period crossover pilot clinical study. Nine subjects with KATPHI received either exendin-(9-39) or vehicle on two different days. The primary outcome was blood glucose; secondary outcomes were insulin, glucagon, and GLP-1. In all subjects, mean nadir blood glucose and glucose area under the curve were significantly increased by exendin-(9-39). Insulin-to-glucose ratios were significantly lower during exendin-(9-39) infusion compared with vehicle. Fasting glucagon and intact GLP-1 were not affected by treatment. In addition, exendin-(9-39) significantly inhibited amino acid–stimulated insulin secretion in pancreatic islets isolated from neonates with KATPHI. Our findings have two important implications: 1) GLP-1 and its receptor play a role in the regulation of fasting glycemia in KATPHI; and 2) the GLP-1 receptor may be a therapeutic target for the treatment of children with KATPHI.
Hypertrophic cardiomyopathy (HCM) is a well-recognised complication in infants of diabetic mothers and is attributed to a compensatory increase in fetal insulin secretion. Infants with congenital hyperinsulinism have excessive prenatal and postnatal insulin secretion due to defects in pathways of insulin secretion (most commonly the KATP channel). HCM has been reported in a few neonates with hyperinsulinism, but its extent and risk factors for its development have not been evaluated.
Retrospective chart review of infants, age <3 months, with congenital hyperinsulinism managed by Children’s Hospital of Philadelphia over a 3.5-year period.
Gestational age, birth weight, hyperinsulinism form and treatments, echocardiogram results, cardiac/ respiratory complications.
68 infants were included, 58 requiring pancreatectomy for diffuse (n=28) or focal (n=30) disease, 10 were diazoxide-sensitive. Twenty-five had echocardiograms performed. Ten had HCM, all of whom required pancreatectomy and eight of whom had confirmed ATP-sensitive potassium-hyperinsulinism. Subjects with HCM had younger gestational age 36(32, 38) than their surgical counterparts without HCM 38 (31.6, 43), p=0.02.
HCM appears common in infants with severe hyperinsulinism. Routine echocardiogram and EKG of at-risk newborns should be considered. Fetal hyperinsulinism is the likely mediating factor for HCM in HI infants.
Congenital hyperinsulinism (CHI) occurs as a consequence of unregulated insulin secretion from the pancreatic beta-cells. Severe recessive mutations and milder dominant mutations have been described in the ABCC8 and KCNJ11 genes encoding SUR1 and Kir6.2 subunits of the beta-cell ATP-sensitive K(+) channel. Here we report two patients with CHI unresponsive to medical therapy with diazoxide. Sequencing analysis identified a compound heterozygous mutation in ABCC8 in both patients. The first one is a carrier for the known mild dominant mutation p.Glu1506Lys jointly with the novel mutation p.Glu1323Lys. The second carries the p.Glu1323Lys mutation and a second novel mutation, p.Met1394Arg. Functional studies of both novel alleles showed reduced or null cell surface expression, typical of recessive mutations. Compound heterozygous mutations in congenital hyperinsulinism result in complex interactions. Studying these mechanisms can improve the knowledge of this disease and modify its therapy.
Congenital hyperinsulinism; Mutation; ABCC8; Functional study
The primary accumulating metabolites in fatty acid oxidation defects are intramitochondrial acyl-CoAs. Typically, secondary metabolites such as acylcarnitines, acylglycines and dicarboxylic acids are measured to study these disorders. Methods have not been adapted for tissue acyl-CoA measurement in defects with primarily acyl-CoA accumulation. Our objective was to develop a method to measure fatty acyl-CoA species that are present in tissues of mice with fatty acid oxidation defects using flow-injection tandem mass spectrometry.
Following the addition of internal standards of [13C2] acetyl-CoA, [13C8] octanoyl-CoA, and [C17] heptadecanoic CoA, acyl-CoA’s are extracted from tissue samples and are injected directly into the mass spectrometer. Data is acquired using a 506.9 neutral loss scan and multiple reaction-monitoring (MRM).
This method can identify all long, medium and short-chain acyl-CoA species in wild type mouse liver including predicted 3-hydroxyacyl-CoA species. We validated the method using liver of the short-chain-acyl-CoA dehydrogenase (SCAD) knock-out mice. As expected, there is a significant increase in [C4] butyryl-CoA species in the SCAD −/− mouse liver compared to wild type. We then tested the assay in liver from the short-chain 3-hydroxyacyl-CoA dehydrogenase (SCHAD) deficient mice to determine the profile of acyl-CoA accumulation in this less predictable model. There was more modest accumulation of medium chain species including 3-hydroxyacyl-CoA’s consistent with the known chain-length specificity of the SCHAD enzyme.
Acyl-coenzyme A’s; Fatty acid oxidation defects; Biomarkers; SCAD deficiency; SCHAD deficiency
Glutamate dehydrogenase (GDH) is a homohexameric enzyme that catalyzes the reversible oxidative deamination of L-glutamate to 2-oxoglutarate. Only in the animal kingdom is this enzyme heavily allosterically regulated by a wide array of metabolites. The major activators are ADP and leucine, while the most important inhibitors include GTP, palmitoyl CoA, and ATP. Recently, spontaneous mutations in the GTP inhibitory site that lead to the hyperinsulinism/hyperammonemia (HHS) syndrome have shed light as to why mammalian GDH is so tightly regulated. Patients with HHS exhibit hypersecretion of insulin upon consumption of protein and concomitantly extremely high levels of ammonium in the serum. The atomic structures of four new inhibitors complexed with GDH complexes have identified three different allosteric binding sites. Using a transgenic mouse model expressing the human HHS form of GDH, at least three of these compounds were found to block the dysregulated form of GDH in pancreatic tissue. EGCG from green tea prevented the hyper-response to amino acids in whole animals and improved basal serum glucose levels. The atomic structure of the ECG-GDH complex and mutagenesis studies is directing structure-based drug design using these polyphenols as a base scaffold. In addition, all of these allosteric inhibitors are elucidating the atomic mechanisms of allostery in this complex enzyme.
glutamate dehydrogenase; insulin regulation; polyphenols; allostery; subunit communication; protein dynamics
The purpose of the study was to determine the sensitivity of the 18fluoro-dihydroxyphenylalanine positron emission tomography/computed tomography scan (18 F-PET/CT) in the diagnosis of focal congenital hyperinsulinism (HI).
A retrospective review of children with HI who underwent a preoperative 18 F-PET/CT scan was performed.
Between 1/2008 and 2/2012 we performed 105 consecutive 18 F-PET/CT scans on infants with HI. Fifty-three patients had focal HI. Of those fifty-three patients, eight had a preoperative 18 F-PET/CT scan read as “diffuse disease”. The sensitivity of the study in the diagnosis of focal HI was 85%. The location of the eight missed focal lesions was: head (3), body (2), and tail (3). The 18 F-PET/ CT of the missed head lesions showed homogeneous tracer uptake (n =2) or heterogeneous uptake throughout the pancreas (n=1). The 18 F-PET/CT of the 2 missed body lesions and 1 missed tail lesion showed heterogeneous uptake throughout the pancreas. The 18 F-PET/CT of the other 2 missed tail lesions showed lesions adjacent to and obscured by the signal of the upper renal pole, identified retrospectively by closer observation. Fifty-two of the 105 patients had diffuse HI. Two of them had 18 F-PET/CT studies read as “focal disease”. Therefore, the specificity of the study was 96%. Of the forty-seven 18 F-PET/CT studies read as “focal disease”, forty-five had true focal HI. Therefore, the positive predictive value of the study in the diagnosis of focal HI was 96%.
The sensitivity and specificity of 18 F-PET/CT can be affected by certain anatomic features of the pancreas, by the location of the lesion, and by the reader’s experience.
Congenital hyperinsulinism; Positron Emission Tomography; Pancreatectomy; Hypoglycemia
To determine the outcome of patients who underwent pancreatic head resection and Roux-en-Y pancreaticojejunostomy to the remaining normal pancreatic body and tail for the treatment of a focal lesion in the pancreatic head causing congenital hyperinsulinism (HI).
One hundred thirty-eight patients underwent pancreatic resection for focal HI between 1998 and 2010. Twenty-three patients in the group underwent pancreatic head resection and Roux-en-Y pancreaticojejunostomy.
There were 13 females and 10 males. Median age and weight at surgery were 8 weeks and 5.8 kg, respectively. Twenty-one patients had a near-total pancreatic head resection, and 2 patients had a pylorus-preserving Whipple procedure. The pancreaticojejunostomy anastomosis was performed with interrupted fine monofilament sutures such that the transected end of the pancreatic body was tucked within the end of the Roux-en-Y jejunal limb. Median hospital stay was 22 days. All patients were cured of HI.
We conclude that pancreatic head resection with Roux-en-Y pancreaticojejunostomy is a safe and effective procedure for the treatment of the HI patient with a large focal lesion in the pancreatic head that is not amenable to local resection alone.
Congenital hyperinsulinism; Pancreatectomy; Pancreaticojejunostomy; Roux-en-Y
Glutamate dehydrogenase (GDH) has recently been shown to be involved in two genetic disorders of hyperinsulinemic hypoglycemia in children. These include the Hyperinsulinism / Hyperammonemia Syndrome caused by dominant activating mutations of GLUD1 which interfere with inhibitory regulation by GTP and hyperinsulinism due to recessive deficiency of short-chain 3-hydroxy-acyl-CoA dehydrogenase (SCHAD, encoded by HADH1). The clinical manifestations of the abnormalities in pancreatic ϐ-cell insulin regulation include fasting hypoglycemia, as well as protein-sensitive hypoglycemia. The latter is due to abnormally increased sensitivity of affected children to stimulation of insulin secretion by the amino acid, leucine. In patients with GDH activating mutations, mild hyperammonemia occurs in both the basal and protein-fed state, possibly due to increased renal ammoniagenesis. Some patients with GDH activating mutations appear to be at unusual risk of developmental delay and generalized epilepsy, perhaps reflecting consequences of increased GDH activity in the brain. Studies of these two disorders have been carried out in mouse models to define the mechanisms of insulin dysregulation. In SCHAD deficiency, the activation of GDH is due to loss of a direct inhibitory protein-protein interaction between SCHAD and GDH. These two novel human disorders demonstrate the important role of GDH in insulin regulation and illustrate unexpectedly important reasons for the unusually complex allosteric regulation of GDH.
hypoglycemia; hyperinsulinism; beta-cell; insulin; leucine-sensitive; genetic disorders; children
This systematic review and meta-analysis aimed to quantify the diagnostic performance of pancreatic venous sampling (PVS), selective pancreatic arterial calcium stimulation with hepatic venous sampling (ASVS), and 18F-DOPA positron emission tomography (PET) in diagnosing and localizing focal congenital hyperinsulinism (CHI).
This systematic review and meta-analysis was conducted according to the PRISMA statement. PubMed, EMBASE, SCOPUS and Web of Science electronic databases were systematically searched from their inception to November 1, 2011. Using predefined inclusion and exclusion criteria, two blinded reviewers selected articles. Critical appraisal ranked the retrieved articles according to relevance and validity by means of the QUADAS-2 criteria. Pooled data of homogeneous study results estimated the sensitivity, specificity, likelihood ratios and diagnostic odds ratio (DOR).
18F-DOPA PET was superior in distinguishing focal from diffuse CHI (summary DOR, 73.2) compared to PVS (summary DOR, 23.5) and ASVS (summary DOR, 4.3). Furthermore, it localized focal CHI in the pancreas more accurately than PVS and ASVS (pooled accuracy, 0.82 vs. 0.76, and 0.64, respectively). Important limitations comprised the inclusion of studies with small sample sizes, high probability of bias and heterogeneity among their results. Studies with small sample sizes and high probability of bias tended to overestimate the diagnostic accuracy.
This systematic review and meta-analysis found evidence for the superiority of 18F-DOPA PET in diagnosing and localizing focal CHI in patients requiring surgery for this disease.
Congenital hyperinsulinism; Pancreatic venous sampling; Arterial stimulation venous sampling; 18F-DOPA PET; Positron emission tomography; Diagnosis
Congenital hyperinsulinemic hypoglycemia is a group of genetic disorders of insulin secretion most commonly associated with inactivating mutations of the β-cell ATP-sensitive K+ channel (KATP channel) genes ABCC8 (SUR1) and KCNJ11 (Kir6.2). Recessive mutations of these genes cause hyperinsulinism that is unresponsive to treatment with diazoxide, a channel agonist. Dominant KATP mutations have been associated with diazoxide-responsive disease. We hypothesized that some medically uncontrollable cases with only one KATP mutation might have dominant, diazoxide-unresponsive disease.
RESEARCH DESIGN AND METHODS
Mutations of the KATP genes were identified by sequencing genomic DNA. Effects of mutations on KATP channel function in vitro were studied by expression in COSm6 cells.
In 15 families with diazoxide-unresponsive diffuse hyperinsulism, we found 17 patients with a monoallelic missense mutation of SUR1. Nine probands had de novo mutations, two had an affected sibling or parent, and four had an asymptomatic carrier parent. Of the 13 different mutations, 12 were novel. Expression of mutations revealed normal trafficking of channels but severely impaired responses to diazoxide or MgADP. Responses were significantly lower compared with nine SUR1 mutations associated with dominant, diazoxide-responsive hyperinsulinism.
These results demonstrate that some dominant mutations of SUR1 can cause diazoxide-unresponsive hyperinsulinism. In vitro expression studies may be helpful in distinguishing such mutations from dominant mutations of SUR1 associated with diazoxide-responsive disease.
Proteins involved in mitochondrial metabolic pathways engage in functionally relevant multi-enzyme complexes. We previously described an interaction between short-chain 3-hydroxyacyl-coenzyme A dehydrogenase (SCHAD) and glutamate dehydrogenase (GDH) explaining the clinical phenotype of hyperinsulinism in SCHAD-deficient patients and adding SCHAD to the list of mitochondrial proteins capable of forming functional, multi-pathway complexes. In this work, we provide evidence of SCHAD's involvement in additional interactions forming tissue-specific metabolic super complexes involving both membrane-associated and matrix-dwelling enzymes and spanning multiple metabolic pathways. As an example, in murine liver, we find SCHAD interaction with aspartate transaminase (AST) and GDH from amino acid metabolic pathways, carbamoyl phosphate synthase I (CPS-1) from ureagenesis, other fatty acid oxidation and ketogenesis enzymes and fructose-bisphosphate aldolase, an extra-mitochondrial enzyme of the glycolytic pathway. Most of the interactions appear to be independent of SCHAD's role in the penultimate step of fatty acid oxidation suggesting an organizational, structural or non-enzymatic role for the SCHAD protein.
Mutations in the high-affinity sulfonylurea receptor (SUR)-1 cause one of the severe recessively inherited diffuse forms of congenital hyperinsulinism or, when associated with loss of heterozygosity, focal adenomatosis. We hypothesized that SUR1 mutations would render the β-cell insensitive to sulfonylureas and to glucose. Stimulated insulin responses were compared among eight patients with diffuse hyperinsulinism (two mutations), six carrier parents, and ten normal adults. In the patients with diffuse hyperinsulinism, the acute insulin response to intravenous tolbutamide was absent and did not overlap with the responses seen in either adult group. There was positive, albeit significantly blunted, acute insulin response to intravenous dextrose in the patients with diffuse hyperinsulinism. Graded infusions of glucose, to raise and then lower plasma glucose concentrations over 4 h, caused similar rises in blood glucose but lower peak insulin levels in the hyperinsulinemic patients. Loss of acute insulin response to tolbutamide can identify children with diffuse SUR1 defects. The greater response to glucose than to tolbutamide indicates that ATP-sensitive potassium (KATP) channel–independent pathways are involved in glucose-mediated insulin release in patients with diffuse SUR1 defects. The diminished glucose responsiveness suggests that SUR1 mutations and lack of KATP channel activity may contribute to the late development of diabetes in patients with hyperinsulinism independently of subtotal pancreatectomy.
Mutations of glutamate dehydrogenase cause the hyperinsulinism/hyperammonemia syndrome by desensitizing glutamate dehydrogenase to allosteric inhibition by GTP. Normal allosteric activation of glutamate dehydrogenase by leucine is thus uninhibited, leading us to propose that children with hyperinsulinism/hyperammonemia syndrome will have exaggerated acute insulin responses to leucine in the postabsorptive state. As hyperglycemia increases β-cell GTP, we also postulated that high glucose concentrations would extinguish abnormal responsiveness to leucine in hyperinsulinism/hyperammonemia syndrome patients. After an overnight fast, seven hyperinsulinism/hyperammonemia syndrome patients (aged 9 months to 29 yr) had acute insulin responses to leucine performed using an iv bolus of l-leucine (15 mg/kg) administered over 1 min and plasma insulin measurements obtained at −10, −5, 0, 1, 3, and 5 min. The acute insulin response to leucine was defined as the mean increase in insulin from baseline at 1 and 3 min after an iv leucine bolus. The hyperinsulinism/hyperammonemia syndrome group had excessively increased insulin responses to leucine (mean ± SEM, 73 ± 21 μIU/ml) compared with the control children and adults (n = 17) who had no response to leucine (1.9 ± 2.7 μU/ml; P < 0.05). Four hyperinsulinism/hyperammonemia syndrome patients then had acute insulin responses to leucine repeated at hyperglycemia (blood glucose, 150–180 mg/dl). High blood glucose suppressed their abnormal baseline acute insulin responses to leucine of 180, 98, 47, and 28 μU/ml to 73, 0, 6, and 19 μU/ml, respectively. This suppression suggests that protein-induced hypoglycemia in hyperinsulinism/hyperammonemia syndrome patients may be prevented by carbohydrate loading before protein consumption.
Inflammatory bowel disease, including Crohn's disease (CD) and ulcerative colitis (UC), and type 1 diabetes (T1D) are autoimmune diseases that may share common susceptibility pathways. We examined known susceptibility loci for these diseases in a cohort of 1689 CD cases, 777 UC cases, 989 T1D cases and 6197 shared control subjects of European ancestry, who were genotyped by the Illumina HumanHap550 SNP arrays. We identified multiple previously unreported or unconfirmed disease associations, including known CD loci (ICOSLG and TNFSF15) and T1D loci (TNFAIP3) that confer UC risk, known UC loci (HERC2 and IL26) that confer T1D risk and known UC loci (IL10 and CCNY) that confer CD risk. Additionally, we show that T1D risk alleles residing at the PTPN22, IL27, IL18RAP and IL10 loci protect against CD. Furthermore, the strongest risk alleles for T1D within the major histocompatibility complex (MHC) confer strong protection against CD and UC; however, given the multi-allelic nature of the MHC haplotypes, sequencing of the MHC locus will be required to interpret this observation. These results extend our current knowledge on genetic variants that predispose to autoimmunity, and suggest that many loci involved in autoimmunity may be under a balancing selection due to antagonistic pleiotropic effect. Our analysis implies that variants with opposite effects on different diseases may facilitate the maintenance of common susceptibility alleles in human populations, making autoimmune diseases especially amenable to genetic dissection by genome-wide association studies.
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.
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).
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.
OBJECTIVE—Two recent genome-wide association (GWA) studies have revealed novel loci for type 1 diabetes, a common multifactorial disease with a strong genetic component. To fully utilize the GWA data that we had obtained by genotyping 563 type 1 diabetes probands and 1,146 control subjects, as well as 483 case subject–parent trios, using the Illumina HumanHap550 BeadChip, we designed a full stage 2 study to capture other possible association signals.
RESEARCH DESIGN AND METHODS—From our existing datasets, we selected 982 markers with P < 0.05 in both GWA cohorts. Genotyping these in an independent set of 636 nuclear families with 974 affected offspring revealed 75 markers that also had P < 0.05 in this third cohort. Among these, six single nucleotide polymorphisms in five novel loci also had P < 0.05 in the Wellcome Trust Case-Control Consortium dataset and were further tested in 1,303 type 1 diabetes probands from the Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications (DCCT/EDIC) plus 1,673 control subjects.
RESULTS—Two markers (rs9976767 and rs3757247) remained significant after adjusting for the number of tests in this last cohort; they reside in UBASH3A (OR 1.16; combined P = 2.33 × 10−8) and BACH2 (1.13; combined P = 1.25 × 10−6).
CONCLUSIONS—Evaluation of a large number of statistical GWA candidates in several independent cohorts has revealed additional loci that are associated with type 1 diabetes. The two genes at these respective loci, UBASH3A and BACH2, are both biologically relevant to autoimmunity.
Genome-wide association studies (GWAS) have been fruitful in identifying disease susceptibility loci for common and complex diseases. A remaining question is whether we can quantify individual disease risk based on genotype data, in order to facilitate personalized prevention and treatment for complex diseases. Previous studies have typically failed to achieve satisfactory performance, primarily due to the use of only a limited number of confirmed susceptibility loci. Here we propose that sophisticated machine-learning approaches with a large ensemble of markers may improve the performance of disease risk assessment. We applied a Support Vector Machine (SVM) algorithm on a GWAS dataset generated on the Affymetrix genotyping platform for type 1 diabetes (T1D) and optimized a risk assessment model with hundreds of markers. We subsequently tested this model on an independent Illumina-genotyped dataset with imputed genotypes (1,008 cases and 1,000 controls), as well as a separate Affymetrix-genotyped dataset (1,529 cases and 1,458 controls), resulting in area under ROC curve (AUC) of ∼0.84 in both datasets. In contrast, poor performance was achieved when limited to dozens of known susceptibility loci in the SVM model or logistic regression model. Our study suggests that improved disease risk assessment can be achieved by using algorithms that take into account interactions between a large ensemble of markers. We are optimistic that genotype-based disease risk assessment may be feasible for diseases where a notable proportion of the risk has already been captured by SNP arrays.
An often touted utility of genome-wide association studies (GWAS) is that the resulting discoveries can facilitate implementation of personalized medicine, in which preventive and therapeutic interventions for complex diseases can be tailored to individual genetic profiles. However, recent studies using whole-genome SNP genotype data for disease risk assessment have generally failed to achieve satisfactory results, leading to a pessimistic view of the utility of genotype data for such purposes. Here we propose that sophisticated machine-learning approaches on a large ensemble of markers, which contain both confirmed and as yet unconfirmed disease susceptibility variants, may improve the performance of disease risk assessment. We tested an algorithm called Support Vector Machine (SVM) on three large-scale datasets for type 1 diabetes and demonstrated that risk assessment can be highly accurate for the disease. Our results suggest that individualized disease risk assessment using whole-genome data may be more successful for some diseases (such as T1D) than other diseases. However, the predictive accuracy will be dependent on the heritability of the disease under study, the proportion of the genetic risk that is known, and that the right set of markers and right algorithms are being used.
Microbial transformation studies of the marine diterpene cyanthiwigin B (1), isolated from the Jamaican sponge Myrmekioderma styx, were accomplished. Two actinomycetes cultures Streptomyces NRRL 5690 and Streptomyces spheroides, significantly metabolized cyanthiwigin B to new metabolites. Streptomyces NRRL 5690 transformed cyanthiwigin B to three new compounds cyanthiwigins AE (2), AF (3), AG (4), and the known cyanthiwigin R (5). S. spheroides transformed cyanthiwigin B to cyanthiwigins S (6), E (7), and AE (2). All microbial metabolized derivatives (2 ~ 7) of cyanthiwigin B exhibited the ability to increase the antimicrobial activity of curcuphenol, the major antimicrobial sesquiterpene isolated from M. styx.
Congenital hyperinsulinism (CHI) is a disease characterized by persistent insulin secretion despite severe hypoglycemia. Mutations in the pancreatic ATP-sensitive K+ (KATP) channel proteins sulfonylurea receptor 1 (SUR1) and Kir6.2, encoded by ABCC8 and KCNJ11, respectively, is the most common cause of the disease. Many mutations in SUR1 render the channel unable to traffic to the cell surface, thereby reducing channel function. Previous studies have shown that for some SUR1 trafficking mutants, the defects could be corrected by treating cells with sulfonylureas or diazoxide. The purpose of this study is to identify additional mutations that cause channel biogenesis/trafficking defects and those that are amenable to rescue by pharmacological chaperones. Fifteen previously uncharacterized CHI-associated missense SUR1 mutations were examined for their biogenesis/trafficking defects and responses to pharmacological chaperones, using a combination of immunological and functional assays. Twelve of the 15 mutations analyzed cause reduction in cell surface expression of KATP channels by >50%. Sulfonylureas rescued a subset of the trafficking mutants. By contrast, diazoxide failed to rescue any of the mutants. Strikingly, the mutations rescued by sulfonylureas are all located in the first transmembrane domain of SUR1, designated as TMD0. All TMD0 mutants rescued to the cell surface by the sulfonylurea tolbutamide could be subsequently activated by metabolic inhibition on tolbutamide removal. Our study identifies a group of CHI-causing SUR1 mutations for which the resulting KATP channel trafficking and expression defects may be corrected pharmacologically to restore channel function.
Congenital hyperinsulinism is a condition of dysregulated insulin secretion often caused by inactivating mutations of the ATP-sensitive K+ (KATP) channel in the pancreatic β cell. Though most disease-causing mutations of the 2 genes encoding KATP subunits, ABCC8 (SUR1) and KCNJ11 (Kir6.2), are recessively inherited, some cases of dominantly inherited inactivating mutations have been reported. To better understand the differences between dominantly and recessively inherited inactivating KATP mutations, we have identified and characterized 16 families with 14 different dominantly inherited KATP mutations, including a total of 33 affected individuals. The 16 probands presented with hypoglycemia at ages from birth to 3.3 years, and 15 of 16 were well controlled on diazoxide, a KATP channel agonist. Of 29 adults with mutations, 14 were asymptomatic. In contrast to a previous report of increased diabetes risk in dominant KATP hyperinsulinism, only 4 of 29 adults had diabetes. Unlike recessive mutations, dominantly inherited KATP mutant subunits trafficked normally to the plasma membrane when expressed in COSm6 cells. Dominant mutations also resulted in different channel-gating defects, as dominant ABCC8 mutations diminished channel responses to magnesium adenosine diphosphate or diazoxide, while dominant KCNJ11 mutations impaired channel opening, even in the absence of nucleotides. These data highlight distinctive features of dominant KATP hyperinsulinism relative to the more common and more severe recessive form, including retention of normal subunit trafficking, impaired channel activity, and a milder hypoglycemia phenotype that may escape detection in infancy and is often responsive to diazoxide medical therapy, without the need for surgical pancreatectomy.
The β-cell ATP-sensitive potassium (KATP) channel controls insulin secretion by linking glucose metabolism to membrane excitability. Loss of KATP channel function due to mutations in ABCC8 or KCNJ11, genes that encode the sulfonylurea receptor 1 or the inward rectifier Kir6.2 subunit of the channel, is a major cause of congenital hyperinsulinism. Here, we report identification of a novel KCNJ11 mutation associated with the disease that renders a missense mutation, F55L, in the Kir6.2 protein. Mutant channels reconstituted in COS cells exhibited wild-type like surface expression level and normal sensitivity to ATP, MgADP, and diazoxide. However, the intrinsic open probability of the mutant channel was greatly reduced, by ~10-fold. This low open probability defect could be reversed by application of phosphatidylinositol 4,5-bisphosphates or oleoyl CoA to the cytoplasmic face of the channel, indicating that reduced channel response to membrane phospholipids and/or long chain acyl CoAs underlies the low intrinsic open probability in the mutant. Our findings reveal a novel molecular mechanism for loss of KATP channel function and congenital hyperinsulinism, and support the importance of phospholipids and/or long chain acyl CoAs in setting the physiological activity of β-cell KATP channels. The F55L mutation is located in the slide helix of Kir6.2. Several permanent neonatal diabetes-associated mutations found in the same structure have the opposite effect of increasing intrinsic channel open probability. Our results also highlight the critical role of the Kir6.2 slide helix in determining the intrinsic open probability of KATP channels.
KATP channel; KCNJ11; inward rectifiers; gating; PIP2; long chain-CoA; congenital hyperinsulinism