Monogenic disorders of glucose-stimulated insulin secretion: massively parallel sequencing approaches

Abstract

Maturity-onset diabetes of the young (MODY) and congenital hyperinsulinism (CHI) are monogenic disorders of glucose-stimulated insulin secretion. MODY is the commonest form of monogenic diabetes and results from a heterozygous mutation in one of thirteen identified genes. It is estimated that up to 90% of cases of MODY are not diagnosed, possibly due to the insufficient awareness of or access to genetic screening. CHI is the commonest cause of persistent hypoglycemia in infancy and childhood, for which there are ten identified genes (mutation-positive disease found in 45% of all CHI, and 88% of diazoxide-unresponsive cases).Genetic confirmation of MODY and CHI traditionally involved Sanger sequencing of each exon of each gene individually; however, the development of massively parallel sequencing (MPS) allows simultaneous sequencing of multiple target regions efficiently and cost-effectively. This thesis assessed two forms of MPS - whole exome sequencing and targeted sequencing, in population and small family cohorts.An entire paediatric diabetes clinic underwent targeted MPS for MODY genes, which showed the prevalence of pathogenic/likely pathogenic MODY variants to be 2.1%. Based on these results, a cost-effectiveness analysis was performed, demonstrating that routine MPS screening for MODY in all paediatric cases with presumed type 1 diabetes was cost-saving. The utility of WES was evaluated in MODY and CHI in a proof-of-concept study. Subjects with known mutations in MODY and CHI genes were re-sequenced using two “off-the-shelf” WES capture technologies. All mutations were confirmed; however, the study showed the importance of careful selection of appropriate capture technology to ensure targeting of relevant genes and the use of bioinformatics tools appropriate to mutation type. Seven probands diagnosed with MODY but without a previously identified mutation were also assessed using WES. Three novel variants in known MODY genes were identified by WES, which variants segregated with diabetes in the families. A potential novel MODY gene was also identified through sequencing of one extended family. Six probands with CHI but without a known mutation were also sequenced. Two probands and their affected family members had the same variant in a potential novel CHI gene. MPS is an extremely useful addition to the diagnostic workup for paediatric cases of diabetes overall, in addition to its utility in screening clinically diagnosed MODY/CHI cases. Additionally, MPS can lead to novel gene identification for these conditions.