Inter-laboratory analytical improvement of succinylacetone and nitisinone quantification from dried blood spot samples.

Hilde Laeremans,M Rebecca Heiner-Fokkema,Giancarlo La Marca,Jose Angel Cocho De Juan,Diran Herebian,Tommy Andersson,Adam Gerrard,Mattias Rudebeck,Nils Janzen,Charles Turner

doi:10.1002/jmd2.12112

Abstract

BackgroundNitisinone is used to treat hereditary tyrosinemia type 1 (HT‐1) by preventing accumulation of toxic metabolites, including succinylacetone (SA). Accurate quantification of SA during newborn screening is essential, as is quantification of both SA and nitisinone for disease monitoring and optimization of treatment. Analysis of dried blood spots (DBS) rather than plasma samples is a convenient method, but interlaboratory differences and comparability of DBS to serum/plasma may be issues to consider.MethodsEight laboratories with experience in newborn screening and/or monitoring of patients with HT‐1 across Europe participated in this study to assess variability and improve SA and nitisinone concentration measurements from DBS by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS). Quantification of nitisinone from both DBS and plasma was performed to assess sample comparability. In addition, efforts to harmonize laboratory procedures of SA and nitisinone quantifications during 5 rounds of analysis are described.ResultsNitisinone levels measured from DBS and plasma strongly correlated (R 2 = 0.93). Due to partitioning of nitisinone to the plasma, levels were higher in plasma by a factor of 2.34. In the initial assessment of laboratory performance, all had linear calibrations of SA and nitisinone although there was large inter‐laboratory variability in actual concentration measurements. Subsequent analytical rounds demonstrated markedly improved spread and precision over previous rounds, an outcome confirmed in a final re‐test round.ConclusionThe study provides guidance for the determination of nitisinone and SA from DBS and the interpretation of results in the clinic. Inter‐laboratory analytical harmonization was demonstrated through calibration improvements.

Highlights

Hereditary tyrosinemia type 1 (HT-1; OMIM reference 276700) is a rare autosomal recessive disorder with an incidence of one in 100 000 worldwide.[1]
To test if nitisinone levels measured from plasma samples were comparable to samples prepared as dried blood spots (DBS), concentrations in 27 blood samples were analysed by LC-MS/MS at two different laboratories in the study (Figure 1)
DBS is an attractive alternative to plasma to use for the clinical follow-up of patients

Summary

| INTRODUCTION

Hereditary tyrosinemia type 1 (HT-1; OMIM reference 276700) is a rare autosomal recessive disorder with an incidence of one in 100 000 worldwide.[1]. While it is essential to keep SA levels as low as possible, the dose of nitisinone should not be unnecessarily high since clinical data on long-term usage are sparse and effects of nitisinone during pregnancy are not fully known.[11] Nitisinone has a presumed half-life of 54 hours and is typically dosed once or twice daily at 1 mg/kg/ day.[12,13,14] Monitoring nitisinone levels in combination with SA serve as a tool facilitating disease management, dose optimization, detection of inter-individual variability, and monitoring treatment compliance. Harmonized results allow comparable information to be generated regardless of methodology or site of analysis, improving patient information and outcomes.[20]

| Study design

| RESULTS

| DISCUSSION