Detection of HNF1A MODY Diabetes with Glycan Biomarker

Session: 
PS1 Poster session 1 Odd numbers
Code: 
P47
Location (hall): 
Foyer
Start/end time: 
Monday, July 1, 2019 - 15:45 to 17:15
Daniel
Demus

Daniel Demus1,2, Paulina Urbanowicz1, Richard Gardner1, Bas Jansen1, Manfred Wuhrer2, Katharine Owen3,4, Daniel Spencer1

1Ludger Ltd, Culham Science Centre, UK, 2LUMC, Leiden, Netherlands, 3Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Oxford, UK, 4Oxford NIHR Biomedical Research Centre, Oxford Hospitals NHS Foundation Trust, Oxford, UK

BACKGROUND:
Maturity onset diabetes of the young HNF1A (MODY 3) is the most frequent form of autosomal dominant monogenic diabetes and is caused by mutations in HNF1A gene. MODY is estimated to account for 1–2% of all diabetes cases. The most common form of testing for MODY is genetic testing, however, the cost and its unavailability in some countries lead to over 80% of MODY subjects remain misdiagnosed as either type 1 or type 2 diabetes [1]. As genome-wide association studies (GWAS) showed, N-glycan outer arm fucosylation levels are significantly altered by certain mutations in HNF1A gene [2]. Therefore, MODY 3 is an example of a disease where a glycan biomarker could be used in identification and stratification of patients.

METHODS:
LC-MS method was used to analyze procainamide-labeled N-glycans released from a cohort of 346 diabetes patient plasma samples including 30 subjects with three differently classified mutations in HNF1A gene - damaging, likely-damaging and benign. Improved LC glycan separation was achieved using HALO 2 Penta- HILIC column. Glycan structures were identified and quantitation of antennary fucosylated glycans was performed in a high-throughput manner using an open source HappyTools software. A receiver operating characteristic (ROC) curves were used to estimate the diagnostic accuracy of antennary fucosylated glycan structures for discrimination of certain types of HNF1A mutation. 

RESULTS:
46 glycan peaks were assigned and amongst them, 7 antennary fucosylated glycan structures were identified. A combination of two (A3FG3S2_A4FG4S4) and three (A3FG3S2_A4FG4S4_A4F2G4S4) glycan structures showed the best discriminative power between subjects with damaging HNF1A mutation and a group of subjects with benign or no HNF1A mutation. The best performing single glycan for discrimination between these subject groups was A3FG3S2. 
For discrimination between subjects with likely-damaging HNF1A mutation and subjects with benign or no HNF1A mutation, A3FG3S2_A4F2G4S4 and A3FG3S3_A3F2G3S3_A4F2G4S4 showed the best performing power as glycan combinations and A4F2G4S4 as a single glycan.

CONCLUSIONS:
Most cases with damaging mutations in HNF1A gene are manifested phenotypically. These subjects represent a decreased level of N-glycans with outer-arm fucosylation. Therefore, HPLC methods for determining antennary fucosylated glycan structures as a biomarker are a suitable tool to discriminate MODY subjects.

We have developed and presented a workflow which focuses not only on highly repeatable HPLC analysis but also on high-throughput, systematic data processing. As a result, diagnostic performance of a single glycan biomarker and a combination of biomarkers were evaluated against a set of subjects with different types of mutation in the HNF1A gene. 

In this study, we have presented an innovative way to test glycan features for potential biomarkers in discrimination of subjects with damaging mutations in HNF1A gene. It was shown that a combination of glycan structures perform better than a single glycan as a diagnostic biomarker.

Acknowlegements

This project has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No 722095.

References: 
  1. Shields BM, Hicks S, Shepherd MH, et al. “Maturity-onset diabetes of the young (MODY): how many cases are we missing?” Diabetologia. 2010;53(12):2504–2508
  2. Lauc G, et al. “Genomics meets glycomics-the first GWAS study of human N-Glycome identifies HNF1αas a master regulator of plasma protein fucosylation.” PLoS genetics vol. 6,12 e1001256. 23 Dec. 2010

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