Mutations In GCK Gene Research Articles

Epidemiological and clinical features of newly diagnosed diabetes cases in hospitalized children in a single center from 2010 to 2016

Objective To analyze the epidemiological features, clinical classification, clinical manifestation and the detection of islet autoantibody of the children with diabetes hospitalized in a single center. Methods A retrospective analysis was performed on the 431 patients with diabetes newly diagnosed at the Department of Endocrinology, Capital Institute of Pediatrics-Peking University Teaching Hospital from January 2010 to December 2016.The medical records, including the number of children per year, the proportion of various types of diabetes, children′s general situation, family history, blood sugar, urine and urine ketone body, blood gas analysis, fasting C peptide, and islet autoantibody test results, were analyzed. Results Of the 431 patients, there were 309 cases (71.7%) of type 1 diabetes mellitus (T1DM), 86 cases (20.0%) of type 2 diabetes mellitus (T2DM), 22 cases (5.1%)of special type of diabetes and 14 cases (3.2%) of undetermined type of diabetes.Maturity onset diabetes in young (MODY) was found in 16 cases out of 22 special type of diabetes.Nine MODY children underwent gene detection, and 2 children were detected with mutation in the GCK gene(c.370G>C, p.D124H; c.707A>G, p.E236G) and the types were MODY2.The annual number of children with diabetes from 2010 to 2016 were 43 cases, 50 cases, 65 cases, 65 cases, 68 cases, 75 cases and 65 cases respectively, and the growth rate increased 51.2% from 2010 to 2016.The median age of T1DM patients at diagnosis was 7.0 years old, compared with 12.7 years of T2DM patients; 94.5% (292 cases) of T1DM patients had typical three polys and one little symptoms of diabetes, compared with 75.6% (65 cases) of T2DM patients; 41.1%(128 cases) of T1DM patients had ketoacidosis at the onset of diabetes, compared with 16.2%(14 cases) of T2DM patients; 33.0%(102 cases) of T1DM patients had family history of diabetes, compared with 74.4% (64 cases)of T2DM patients; the median of fasting C peptide of 302 T1DM patients was 0.20 μg/L, compared with 2.14 μg/L of 85 T2DM patients; the differences above were all statistically significant(Z=-11.127, P=0.00; χ2=27.689, P=0.000; χ2=18.473, P=0.000 017; χ2=47.348, P=0.000; Z=-12.865, P=0.00). Two hundred and seventy-nine T1DM patients were tested for islet cell antibody (ICA), insulin autoantibody (IAA), glutamic acid decarboxylase antibody (GADA) at the same time, and the positive rate was 2.9%(8/279 cases), 14.7%(41/279 cases), and 9.3%(26/279 cases) respectively, and there were 65 cases (23.3%) with at least one positive result.One hundred and three T1DM patients were detected by the combination of 5 islet autoantibodies (ICA, IAA, GADA, islet antigen-2 antibody, Zinc transporter type 8 autoantibodies), and the positive rate of at least 1 antibody was 53.4%(55/103 cases), while 23 T2DM patients were tested for these five auto-antibodies, and the positive rate of at least 1 antibody was 13.0%(3/23 cases), and the difference between 2 groups was statistically significant(χ2=12.325, P=0.000 447). Conclusions (1)The number of hospitalized diabetes of children and adolescents in our center was increasing gradually.(2)In the center′s hospitalized diabetes patients, T1DM accounted for the most, T2DM took the second place, and there were minority of special type of diabetes.MODY accounted for the most part of special type of diabetes, and the main gene mutation was GCK gene mutation with type of MODY2.(3)T1DM had the characteristics of early onset age, typical three polys and one little symptoms of diabetes, more ketoacidosis at onset, low fasting C peptide, fewer family history of diabetes compared with T2DM.(4)The combined detection of islet autoantibodies could improve the detection rate of T1DM. Key words: Diabetes mellitus; Child; Epidemiological features; Clinical classification; Clinical features; Autoantibodies

Analysis of the GCK gene in 79 MODY type 2 patients: A multicenter Turkish study, mutation profile and description of twenty novel mutations

Maturity onset diabetes is a genetic form of diabetes mellitus characterized by an early age at onset and several etiologic genes for this form of diabetes have been identified in many patients. Maturity onset diabetes type 2 [MODY2 (#125851)] caused by mutations in the glucokinase gene (GCK). Although its prevalence is not clear, it is estimated that 1%–2% of patients with diabetes have the monogenic form. The aim of this study was to evaluate the molecular spectrum of GCK gene mutations in 177 Turkish MODY type 2 patients. Mutations in the GCK gene were identified in 79 out of 177. All mutant alleles were identified, including 45 different GCK mutations, 20 of which were novel.

The molecular diagnosis and their detailed clinical presentations in 5 cases of maturity-onset diabetes of the young

Objective To analyze the genetic changes and detailed clinical presentations of 5 maturity-onset diabetes of the young (MODY) cases in order to enhance the knowledge about MODY in children. Methods Seventy-eight patients initially diagnosed as diabetes mellitus between January 1 and December 31, 2015 in Capital Institute of Pediatrics were retrospectively studied.Nine of them were suspected of MODY, and 5 patients were diagnosed as MODY through gene test.Clinical informations were collected including age, gender, main complaint, family history, body mass index(BMI), fasting blood glucose, fasting blood insulin, 2-hour blood glucose and insulin after oral glucose tolerance test and glycosylated hemoglobin.The blood glucose was monitored dynamically in 2 patients.Targeted capture panel was designed to capture the 16 genes related to MODY, including 12 genes from MODY1 to MODY13 type and 4 genes with weak evidence of MODY according to Human Gene Mutation Database Exome capture, and Next-Generation sequencing on a HiSeq2000(Illumina) was performed.After bioinformatics analysis, all prioritized variants detected in patients were validated by Sanger sequencing, including the probands and their parents. Results Five patients were confirmed as MODY by molecular diagnosis, accounting for 6.4% of all the 78 patients in 2015.The ratio of male to female was 23.The ages at diagnosis ranged from 2 to 11 years old, and the median age was 3 years old.Two cases were found to have abnormal blood glucose in physical examination.The rest 3 cases were discovered with abnormal blood glucose during hospitalization because of pneumonia (1 case)or diarrhea(2 cases). In 4 cases, their mothers had gestational diabetes history, in 1 case the father suffering from diabetes.BMI ranged 15.68-23.40 kg/m2.Fasting blood glucose was 6.3-7.2 mmol/L.Fasting blood insulin was 0.5-8.0 IU/L.Glucose tolerance test results showed that blood glucose of the patients was 8.6-10.8 mmol/L after 2 hours.The level of glycosylated hemoglobin was 5.5%-6.7%.Blood glucose was 3.9-13.0 mmol/L.All the 5 confirmed patients were caused by GCK gene mutation (MODY2 type). The mutations detected were located at Exon7(2 cases), Exon4(1 case), Exon5(1 case), and Exon10(1 case). Conclusions All the confirmed MODY patients were identified either through medical exam or infectious disease, and all had positive family history.Their BMI ranged widely.Fasting blood glucose was slightly elevated and glycosylated hemoglobin was normal or slightly elevated, but fasting blood insulin was normal in all the patients.Abnormal glucose tolerance test results were found in all 5 patients.Glycosylated hemoglobin was normal or slightly elevated.MODY2 was the only subtype detected in this group, which indicated that the common type in children was different from that in adults. Key words: Maturity-onset diabetes of the young; Glucokinase; Next-generation sequencing

GCK gene mutations are a common cause of childhood-onset MODY (maturity-onset diabetes of the young) in Turkey.

SummaryObjectiveInactivating heterozygous mutations in the GCK gene are a common cause of MODY and result in mild fasting hyperglycaemia, which does not require treatment. We aimed to identify the frequency, clinical and molecular features of GCK mutations in a Turkish paediatric cohort.Design and PatientsFifty‐four unrelated probands were selected based on the following criteria: age of diagnosis ≤17 years, family history of diabetes in at least two generations, anti‐GAD/ICA negative, BMI<95.p and follow‐up with diet, oral antidiabetic drug or low‐dose insulin treatment (≤0·5U/kg/d). A MODY probability score (www.diabetesgenes.org) was calculated and 21 patients with a score ≥75%, HbA1c levels ≤7·5% (58·5 mmol/mol) and fasting blood glucose (FBG) levels 99–145 mg/dl (5·5–8·0 mmol/l) were selected for Sanger sequencing of the GCK gene. Targeted next‐generation sequencing for all known monogenic diabetes genes was undertaken for any patient without a GCK gene mutation.Results GCK gene mutations (pathogenic or likely pathogenic variants) and a novel intronic variant of uncertain significance (c.208 + 3A>T) were identified in 13/54 probands (24%). Twelve of these patients had a MODY probability score ≥75%. FBG level and 2‐h glucose level in OGTT were 123 ± 14 mg/dl (6·8 ± 0·7 mmol/l) (107–157 mg/dl) and 181 ± 30 mg/dl (10·1 ± 1·6 mmol/l) (136–247 mg/dl), respectively. Average of glucose increment in OGTT was 58 ± 27 mg/dl (3·2 ± 1·5 mmol/l) (19–120 mg/dl), and mean HbA1c level was 6·5 ± 0·5% (47·5 ± 5·5 mmol/mol) (5·9–7·6%). Five novel missense mutations were identified (p.F123S, p.L58P, p.G246A, p.F419C, and p.S151C). Two patients treated with low‐dose insulin before the molecular analysis were able to stop treatment.ConclusionsApproximately 1 in 4 MODY cases in this Turkish paediatric cohort have a GCK mutation. Selection of patients for GCK gene analysis using the MODY probability score was an effective way of identifying most (11/12) patients with a GCK mutation.

Genetic counseling in monogenic diabetes GCK MODY.

Genetic testing in families with monogenic GCK MODY has predictive, diagnostic, and preventive utility. Predictive tests relate to people who have no features of the disorder themselves at the time of testing. Diagnostic tests relate to family members who have been previously diagnosed with diabetes mellitus or glucose metabolism disturbances. The preventive value of genetic testing for families is to raise awareness of the circumstances leading to glucose metabolism disorders. The detection of mutation carriers among family members of patients with GCK MODY and the determination of the clinical significance of the genetic test result. The study group included 27 families of adolescent patients with GCK MODY (39 (75%) of parents and 19 (73.08%) of siblings) monitored in the Department of Pediatrics, Endocrinology and Diabetes and in the Diabetes Clinic of John Paul II Upper Silesian Child Health Centre in Katowice in the years 2007-2012. Subjects underwent a blood sample drawing for genetic and biochemical testing. Through the genetic diagnostics we diagnosed GCK MODY in 14 (63.64%) mothers, 6 (35.29%) fathers and in 7 (36,84%) siblings. Genetic testing has contributed to the detection of 7 (26.92%) asymptomatic carriers of GCK gene mutation among parents and 3 (15,79%) asymptomatic carriers among siblings declaring no carbohydrate metabolism disturbances (before genetic testing there were no indications suggesting carbohydrate metabolism disturbances; OGTT were performed after positive genetic testing). Each case of mutation detection, which is the cause of monogenic diabetes in a patient, justifies the genetic testing in other members of his/her family. Awareness of the genetic status may allow sick family member to confirm the diagnosis, while asymptomatic mutation carriers could benefit from an early clinical observation. Consequently, in each case it gives an opportunity to take diagnostic and therapeutic measures in accordance with the current state of knowledge.

A new compound heterozygosis for inactivating mutations in the glucokinase gene as cause of permanent neonatal diabetes mellitus (PNDM) in double-first cousins.

BackgroundPermanent neonatal diabetes mellitus (PNDM) is a rare disorder, characterized by uncontrolled hyperglycemia diagnosed during the first 6 months of life. In general, PNDM has a genetic origin and most frequently it results from heterozygous mutations in KCNJ11, INS and ABCC8 genes. Homozygous or compound heterozygous inactivating mutations in GCK gene as cause of PNDM are rare. In contrast, heterozygosis for GCK inactivating mutations is frequent and results in the maturity-onset diabetes of young (MODY), manifested by a mild fasting hyperglycemia usually detected later in life. Therefore, as an autosomal recessive disorder, GCK-PNDM should be considered in families with history of glucose intolerance or MODY in first relatives, especially when consanguinity is suspected.ResultsHere we describe two patients born from non-consanguineous parents within a family. They presented low birth weight with persistent hyperglycemia during the first month of life. Molecular analyses for KCNJ11,INS, ABCC8 did not show any mutation. GCK gene sequencing, however, revealed that both patients were compound heterozygous for two missense combined in a novel GCK-PNDM genotype. The p.Asn254His and p.Arg447Gly mutations had been inherited from their mothers and fathers, respectively, as their mothers are sisters and their fathers are brothers. Parents had been later diagnosed as having GCK-MODY.ConclusionsMutations’ in silico analysis was carried out to elucidate the role of the amino acid changes on the enzyme structure. Both p.Asn254His and p.Arg447Gly mutations appeared to be quite damaging. This is the first report of GCK-PNDM in a Brazilian family.

Genetic testing for monogenic diabetes using targeted next-generation sequencing in patients with maturity-onset diabetes of the young

Molecular diagnosis of monogenic diabetes mellitus is important for individualized patient care. Next-generation sequencing (NGS) enables a simultaneous analysis of multiple genes in a single test. We aimed to assess the feasibility of using NGS for detecting mutations in a set of known monogenic diabetes gene mutations in a cohort of Polish patients with maturity-onset diabetes of the young (MODY) with earlier negative Sanger sequencing results for HNF1A-MODY or GCK-MODY. We selected a panel of 28 chromosomal genes in which mutations have been reported to cause monogenic diabetes. The MiSeq platform was used for NGS. An exon-capture assay was designed to include coding regions and splice sites. A total of 54 patients with existing negative Sanger sequencing screening results for HNF1A or GCK gene mutations were selected for the study. NGS results were generated for all 54 patients and 9 positive controls with previously identified HNF1A or GCK gene mutation. All selected positive controls were confirmed by NGS. Among 28 genes, mutations were detected in 16. The type of the analyzed genetic changes was described in the NGS study as high (n = 3) or moderate (n = 76). Among the detected mutations, there were 4 known GCK gene mutations that had been previously missed in Sanger sequencing. So far, Sanger sequencing allowed us to confirm 21 gene mutations detected by NGS, and segregation with diabetes in 14 pedigrees. Our pilot study using NGS for monogenic diabetes screening in the MODY cohort confirmed that it improves the detection of diabetes-related sequence differences. The screening with NGS should also include diabetic patients for whom Sanger-based screening for particular subtypes of MODY provided negative results.

Recognition and Management of Individuals With Hyperglycemia Because of a Heterozygous Glucokinase Mutation.

Glucokinase-maturity-onset diabetes of the young (GCK-MODY), also known as MODY2, is caused by heterozygous inactivating mutations in the GCK gene. GCK gene mutations are present in ∼1 in 1,000 of the population, but most are not diagnosed. They are common causes of MODY (10-60%): persistent incidental childhood hyperglycemia (10-60%) and gestational diabetes mellitus (1-2%). GCK-MODY has a unique pathophysiology and clinical characteristics, so it is best considered as a discrete genetic subgroup. People with GCK-MODY have a defect in glucose sensing; hence, glucose homeostasis is maintained at a higher set point resulting in mild, asymptomatic fasting hyperglycemia (5.4-8.3 mmol/L, HbA1c range 5.8-7.6% [40-60 mmol/mol]), which is present from birth and shows slight deterioration with age. Even after 50 years of mild hyperglycemia, people with GCK-MODY do not develop significant microvascular complications, and the prevalence of macrovascular complications is probably similar to that in the general population. Treatment is not recommended outside pregnancy because glucose-lowering therapy is ineffective in people with GCK-MODY and there is a lack of long-term complications. In pregnancy, fetal growth is primarily determined by whether the fetus inherits the GCK gene mutation from their mother. Insulin treatment of the mother is only appropriate when increased fetal abdominal growth on scanning suggests the fetus is unaffected. The impact on outcome of maternal insulin treatment is limited owing to the difficulty in altering maternal glycemia in these patients. Making the diagnosis of GCK-MODY through genetic testing is essential to avoid unnecessary treatment and investigations, especially when patients are misdiagnosed with type 1 or type 2 diabetes.

Analysis of the glucokinase gene in Iranian families with maturity onset diabetes of the young

Non insulin dependent diabetes mellitus (NIDDM) as a most common form of diabetes is a major public health problem; there is a subgroup of NIDDM patients who develop the disease at an early age and show a dominant mode of inheritance. This type is nominates Maturity onset diabetes of the young (MODY). The prevalence of MODY is difficult to access, and patients with MODY genes mutations are often identified during routine screening for other purposes. MODY2 was linked to glucokinase gene (GCK) mutations, and accounted for 8% to 56% of MODY, with the highest prevalence found in the southern Europe. The aim of this study was to examine the prevalence and nature of mutations in GCK gene in Iranian paients. We have screened GCK mutations by polymerase chain reaction (PCR); single stranded conformation polymorphism (SSCP) technique in 12 Iranian families with clinical diagnosis of MODY, included 30 patients (8 males and 22 females) and their 21 family members. PCR products with abnormal mobility in denaturing gradient gel electrophoresis (DGGE) were directly sequenced. We identified 6 novel mutations in GCK gene in Iranian families (corresponding to 36.6% prevalence). Our findings and the last study on MODY1 highlight that in addition to GCK, other MODY genes such as MODY3 and MODYX may play a significant role in diabetes characterized by monogenic autosomal dominant transmission. There is an important point that the genetic recognation can be used to pre-symptomatically identify family members at risk for developing MODY.

Clinical follow-up of two Brazilian subjects with glucokinase-MODY (MODY2) with description of a novel mutation

Mutations in the glucokinase gene (GCK) account for many cases of monogenic diabetes featuring maturity-onset diabetes of the young type 2 (MODY2). The clinical pattern of this form of hyperglycemia is rather stable, with a slight elevation in blood glucose, which is usually not progressive. Patients rarely require pharmacological interventions and microvascular complications related to diabetes are unusual. We describe the clinical follow-up of two cases of MODY2 with two different mutations in GCK gene, one in exon 7, p.Glu265Lys (c.793 G> A), which has been previously described, and a novel one, in exon 2, p.Ser69Stop (c. 206C> G). The clinical course of both cases shows similarity in metabolic control of this form of diabetes over the years.

Functional Characterization of MODY2 Mutations Highlights the Importance of the Fine-Tuning of Glucokinase and Its Role in Glucose Sensing

Glucokinase (GK) acts as a glucose sensor in the pancreatic beta-cell and regulates insulin secretion. Heterozygous mutations in the human GK-encoding GCK gene that reduce the activity index increase the glucose-stimulated insulin secretion threshold and cause familial, mild fasting hyperglycaemia, also known as Maturity Onset Diabetes of the Young type 2 (MODY2). Here we describe the biochemical characterization of five missense GK mutations: p.Ile130Thr, p.Asp205His, p.Gly223Ser, p.His416Arg and p.Ala449Thr. The enzymatic analysis of the corresponding bacterially expressed GST-GK mutant proteins show that all of them impair the kinetic characteristics of the enzyme. In keeping with their position within the protein, mutations p.Ile130Thr, p.Asp205His, p.Gly223Ser, and p.His416Arg strongly decrease the activity index of GK, affecting to one or more kinetic parameters. In contrast, the p.Ala449Thr mutation, which is located in the allosteric activator site, does not affect significantly the activity index of GK, but dramatically modifies the main kinetic parameters responsible for the function of this enzyme as a glucose sensor. The reduced Kcat of the mutant (3.21±0.28 s−1 vs 47.86±2.78 s−1) is balanced by an increased glucose affinity (S0.5 = 1.33±0.08 mM vs 7.86±0.09 mM) and loss of cooperativity for this substrate. We further studied the mechanism by which this mutation impaired GK kinetics by measuring the differential effects of several competitive inhibitors and one allosteric activator on the mutant protein. Our results suggest that this mutation alters the equilibrium between the conformational states of glucokinase and highlights the importance of the fine-tuning of GK and its role in glucose sensing.

Identification of eight new mutations in the GCK gene by DHPLC screening in a Spanish population

Maturity onset diabetes of the young (MODY) is a genetically heterogeneous disorder characterized by autosomal dominant inheritance, altered function of pancreatic beta cells and early onset diabetes mellitus, usually before 25 years old. The prevalence of specific mutations of MODY genes differs considerably among different countries. In this study we analyzed 53 index cases from unrelated MODY families who are potential carriers of mutations in GCK gene. In addition, 122 relatives were also studied. We have identified eight new mutations in the GCK gene. One of them is a non-frameshift deletion involving Lysine 143. This amino acid is part of the conserved stretch of basic residues (KHKKL) which spans from residue 140 to 144. The non-frameshit deletion might implicate the affinity of GCK for GCKRP, and potentially the abnormal nuclear localization of GCK. Additional studies should be performed to confirm this possibility.

Glucokinase Gene Mutation Screening in Argentinean Clinically Characterized MODY Patients

Mutations in the glucokinase gene (GCK) produce a subtype of Maturity onset diabetes in the young (MODY), named MODY 2. To date over than 190 different mutations have been identified, distributed over the coding regions and the exon-intron boundaries of the gene. The aim of this work was to study the nature and frequency of mutations in the GCK gene, in a MODY clinically characterized Argentinean population. Seventy unrelated individuals were selected based on MODY clinical features. The study methodology consisted in PCR amplification of the coding regions of the GCK gene, SSCP electrophoresis analysis of the amplified fragments and direct sequencing of the fragments with abnormal electrophoresis pattern. We identified a total of six patients with mutations in the GCK gene. This included two novel mutations: g.1831C>A, g.3792T>A, one already reported by our group, g.168fsdelC (same mutation in two non-related patients) and two already reported: p.Gln138Pro and p.Gly261Glu. With that data, we could establish the prevalence of MODY 2 among the patients in study reaching to 8.6%. The main contribution of this study is to inform about two novel mutations not described to date and to make an approach to the establishment of the prevalence of MODY 2 in the population under study. These findings contribute to confirm the allelic heterogeneity of GCK gene mutations and may provide an insight into the structure-function relationship of the GCK.

Screening of mutations in the GCK gene in Jordanian maturity-onset diabetes of the young type 2 (MODY2) patients

Maturity-onset diabetes of the young type 2 (MODY2) is a genetic form of diabetes mellitus caused by mutations in the glucokinase gene (GCK). We assessed the frequency of GCK gene mutations in Jordanian suspected MODY2 patients. We screened exons 7, 8 and 9, which are specific for pancreatic glucokinase, for mutations at positions 682A>G, p.T228A; 895G>C, p.G299R, and 1148C>A, p.S383X, respectively, in 250 subjects (100 patients suspected to have MODY2 and 150 healthy controls without family history of diabetes mellitus). We did not find any association of these mutations in Jordanian suspected MODY2 patients or in healthy controls, different from data on Caucasian Italian patients screened for the same mutations.

Genetic and clinical characteristics of Korean maturity-onset diabetes of the young (MODY) patients

Maturity-onset diabetes of the young (MODY) is mostly caused by mutations of the hepatocyte nuclear factor (HNF)-1α (MODY3) and glucokinase (MODY2) genes in Caucasians. But most Japanese and Chinese MODY patients are not linked to known MODY genes. In this study, we examined the genetic and clinical characteristics of Korean subjects with MODY and early onset type 2 diabetes who had been diagnosed before 15 years of age. The study included 23 unrelated subjects fulfilling the criteria for MODY (three consecutive generations of type 2 diabetes with at least one member diagnosed under the age of 25 year) and 17 unrelated subjects diagnosed with early onset type 2 DM under the age of 15 years. The HNF-4α (MODY1), glucokinase (MODY2) and HNF-1α (MODY3) genes were analysed by direct sequencing. Mutations in the HNF-1α gene were found in two patients (5%). One of these, P393fsdelC, was novel, and was found in a patient classified in the MODY group. The GCK gene mutation, R191W, was identified in one patient classified as early-onset type 2 DM (2.5%). No mutations were found in the HNF-4α gene, except the T130I variant, which is a known rare polymorphism. In conclusion, the mutations in the HNF-1α gene and GCK account for a small proportion, about 5% and 2.5%, respectively, in Korean MODY and early onset type 2 patients. The majority of MODY cases in the Korean population are due to defects in unknown genes.

Effects of novel maturity-onset diabetes of the young (MODY)-associated mutations on glucokinase activity and protein stability.

Glucokinase acts as the pancreatic glucose sensor and plays a critical role in the regulation of insulin secretion by the beta-cell. Heterozygous mutations in the glucokinase-encoding GCK gene, which result in a reduction of the enzymatic activity, cause the monogenic form of diabetes, MODY2 (maturity-onset diabetes of the young 2). We have identified and functionally characterized missense mutations in the GCK gene in diabetic families that result in protein mutations Leu165-->Phe, Glu265-->Lys and Thr206-->Met. The first two are novel GCK mutations that co-segregate with the diabetes phenotype in their respective families and are not found in more than 50 healthy control individuals. In order to measure the biochemical effects of these missense mutations on glucokinase activity, we bacterially expressed and affinity-purified islet human glucokinase proteins carrying the respective mutations and fused to GST (glutathione S-transferase). Enzymatic assays on the recombinant proteins revealed that mutations Thr206-->Met and Leu165-->Phe strongly affect the kinetic parameters of glucokinase, in agreement with the localization of both residues close to the active site of the enzyme. In contrast, mutation Glu265-->Lys, which has a weaker effect on the kinetics of glucokinase, strongly affects the protein stability, suggesting a possible structural defect of this mutant protein. Finally, none of the mutations tested appears to affect the interaction of gluco-kinase with the glucokinase regulatory protein in the yeast two-hybrid system.

Clinical characteristics of mutation carriers in a large family with glucokinase diabetes (MODY2)

To investigate the prevalence and clinical characteristics of heterozygotes of the glucokinase gene mutations G264S and IVS8+2 in the extended pedigree of two patients with permanent neonatal diabetes as a result of glucokinase deficiency (IVS8+2 homozygosity and IVS8+2/G264S compound heterozygosity). Eighty-eight first, second and third degree family members of the two patients with permanent neonatal diabetes were genotyped. Clinical, laboratory and historical data were collected via chart reviews. Thirty-one IVS8+2 and three G264S heterozygotes were identified. Of these, 18/34 (52.9%) had diabetes and 9/34 (26.5%) impaired fasting glucose (IFG), compared with 1/54 (1.9%) with diabetes and 2/54 (3.7%) with IFG in the non-carrier group. Odds ratio for heterozygotes was 70.4 (95% CI 16.9-293.5 and P < 0.001). Mean body mass index of heterozygotes (> 18 years of age) who had diabetes was 27.1 +/- 2.66, compared with 23.18 +/- 4.72 for heterozygotes with normal glucose levels (P < 0.05). While none of the non-carrier women had gestational diabetes, eight of the 10 heterozygotes developed gestational diabetes. Inheritance of a glucokinase mutation by the fetus from a carrier mother resulted in a significant reduction in birthweight (3600 +/- 570 vs. 2970 +/- 390 grams, P < 0.05). These data support the association between carriage of GCK gene mutations, G264S and IVS8+2, and the development of diabetes, impaired fasting glucose and reduced birthweight. Moreover, in heterozygotes, a clear correlation between body mass index and the development of diabetes was observed. These findings underline the need for surveillance and prevention in individuals at risk.

Diagnosis of MODY in the offspring of parents with insulin-dependent and non-insulin-dependent diabetes mellitus.

Maturity Onset Diabetes of the Young (MODY) is an autosomal dominant monogenic form of type 2 diabetes mellitus (DM) representing 5% of youth-onset DM in the Caucasian population. In young adults the disease can be present as either non-insulin dependent or insulin-dependent DM. The diagnosis of this genetic disorder in children and adolescents is rare because of the mild glucose metabolism disorder at this time. We performed a metabolic, autoimmune and genetic study in 40 offspring of young parents affected by insulin-dependent DM (Group A) and in 35 offspring of young parents affected by early-onset non-insulin-dependent DM (Group B). Two children of Group A (5%) were found to be affected by fasting hyperglycemia and carry a GCK gene mutation that in one case was present also in the diabetic father. Eighteen offspring of Group B (51%) were positive for GCK or HNF-1alpha gene mutations present in the affected parents. All but two of these young patients had impaired fasting glucose (IFG) or impaired glucose tolerance (IGT). Eleven of them were younger than 16 years. We conclude that screening for DM in youth should be extended to MODY in young families with both non-insulin-dependent and insulin-dependent DM. The sensitivity of the metabolic tests will precede the genetic diagnosis.