Direct Infusion Based Metabolomics Identifies Metabolic Disease in Patients' Dried Blood Spots and Plasma.

Hanneke Haijes,Nanda Verhoeven-Duif,Maria Van Der Ham,Marcel Willemsen,Hubertus Prinsen,Mia Pras-Raves,Monique De Sain-Van Der Velden,Johan Gerrits,Peter Van Hasselt,Judith Jans

doi:10.3390/metabo9010012

Abstract

In metabolic diagnostics, there is an emerging need for a comprehensive test to acquire a complete view of metabolite status. Here, we describe a non-quantitative direct-infusion high-resolution mass spectrometry (DI-HRMS) based metabolomics method and evaluate the method for both dried blood spots (DBS) and plasma. 110 DBS of 42 patients harboring 23 different inborn errors of metabolism (IEM) and 86 plasma samples of 38 patients harboring 21 different IEM were analyzed using DI-HRMS. A peak calling pipeline developed in R programming language provided Z-scores for ~1875 mass peaks corresponding to ~3835 metabolite annotations (including isomers) per sample. Based on metabolite Z-scores, patients were assigned a ‘most probable diagnosis’ by an investigator blinded for the known diagnoses of the patients. Based on DBS sample analysis, 37/42 of the patients, corresponding to 22/23 IEM, could be correctly assigned a ‘most probable diagnosis’. Plasma sample analysis, resulted in a correct ‘most probable diagnosis’ in 32/38 of the patients, corresponding to 19/21 IEM. The added clinical value of the method was illustrated by a case wherein DI-HRMS metabolomics aided interpretation of a variant of unknown significance (VUS) identified by whole-exome sequencing. In summary, non-quantitative DI-HRMS metabolomics in DBS and plasma is a very consistent, high-throughput and nonselective method for investigating the metabolome in genetic disease.

Highlights

With unprecedented pace, the number of known inborn errors of metabolism (IEM) is expanding.It is a challenge to diagnose this diverse spectrum of diseases in a timely manner
We evaluate the use of a non-quantitative metabolomics method for both dried blood spots (DBS) and plasma by analyzing samples of patients with a broad range of IEM
The samples of patients and controls were analyzed in seven batches by direct-infusion high-resolution mass spectrometry (DI-HRMS) (Figure 1)

Summary

Introduction

The number of known inborn errors of metabolism (IEM) is expanding.It is a challenge to diagnose this diverse spectrum of diseases in a timely manner. Metabolites 2019, 9, 12 this need, metabolomics—the parallel determination of thousands of small-molecule metabolites—is regarded as the way forward. All intermediates and final products of metabolic pathways in the body can potentially be measured. Metabolomics with exact quantification of a subset of predefined metabolites has been used for small-scale diagnostic screening [2,3,4] and for biomarker identification in predefined patient groups [5]. While broad quantitative metabolomics assays are already a big step forward compared to the focused targeted assays currently used in metabolic diagnostics, semi- and non-quantitative metabolomics can potentially detect an even larger range of metabolites. In the IEM field, both combined approaches of quantitative and non-quantitative metabolomics [6,7,8,9,10] as well as exclusively non-quantitative metabolomics have been used to study predefined patient groups [11,12,13,14,15,16,17,18,19,20]

Methods

Results

Discussion

Conclusion