Six novel mutations in the GCK gene in MODY patients

To the Editor: Maturity-onset diabetes of the young (MODY; MIM# 606391) is a genetically and clinically heterogeneous form of diabetes mellitus, characterized by an autosomal dominant inheritance, early-onset non-insulin-dependent diabetes mellitus and by a primary defect in the pancreatic beta-cell function (1). Until now, six types of MODY diabetes have been identified, depending on the gene causing the disease (2). Screening for glucokinase (GCK) mutations in subjects with clinical characteristics of MODY allows distinguishing between patients with a benign metabolic condition (GCK mutation positive, clinical diagnosis MODY2) and those with a higher risk of progressive hyperglycemia associated with more prevalent and severe diabetic complications (GCK mutation negative). The first mutation in the GCK gene was reported in 1992 (3). Up to now, 195 mutations in GCK have been described, in 285 families (4). Diabetic complications are rare in GCK–MODY, thus GCK–MODY patients only need to be followed by annual HbA1c examination. Also, screening of GCK for heterozygous inactivating mutations allows to determine the subtype of MODY diabetes and to predict the lifelong prognosis. All 12 exons (exons 1a, 1b, 1c and 2–10), the intron–exon boundaries and promotor region of GCK (GenBank accession number, AF04101222) were screened; in 92 Czech probands fulfilling classical MODY criteria, using denaturing highperformance liquid chromatography as previously described (5). The nature of identified mutations was established by direct nucleotide sequencing using BigDye Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems, Foster City, USA) according to manufacture’s instructions. Mutations were confirmed using a second, independent amplification of the affected part of GCK and re-sequenced the following day. The probands were recruited from pediatricians and endocrinologists from the entire Czech Republic. Fifteen different missense mutations were identified in 27 patients. Of these, six were novel missense mutations R250C (exon 7, c.748C.T), L315H (exon 8, c.944T.A), F316V (exon 8, c.946T.G), F419L (exon 10, c.1255T.C), I436N (exon 10, c.1307T.A) and A454E (exon 10, c.1361C.A). Some of the identified GCK missense mutations are located near putative functional domains: R250C was found in the close vicinity of a putative glucose binding site, while F419L was detected near a putative MgATP binding site and could thus affect binding kinetics (6). Five of these mutations co-segregated with hyperglycemia in the family, suggesting that the variants are new diseasecausing mutations. For the novel R250C variant, family members were not available for cosegregation studies. All codons, which are changed by the six novel mutations, are conserved in the human, mouse, rat and chimpanzee genomes and we found none of these mutations in 50 unrelated healthy Czech Caucasian subjects. Therefore, we assume that the mutations are probably novel disease-causing mutations. We also compared the clinical characteristics of patients with GCK mutations and those without mutation in GCK (data not shown in details). In short – the treatment of hyperglycemia with diet was more frequent (p , 0.001) in the group of probands with mutations in GCK and they had a significantly lower frequency of diabetic complications (p 1⁄4 0.02). None of the patients with mutations in GCK was treated with insulin (p , 0.001). Moreover, GCK mutation carriers had a lower level of glycosylated hemoglobin (p1⁄4 0.02). ThemeanHbA1c (%) inGCK-positive probands vs negative was 5.7 0.2 vs 6.5 0.2. In conclusion, we identified 29% of GCK mutation carriers among Czech MODY probands, confirming that mutations in GCK are a common cause of MODY in the Czech population. The present high relative prevalence of GCK–MODY, compared with some other European studies, might reflect not only a specific genetic background, but also the mode of recruitment, because most of the probands in the present investigation were recruited by