A novel tandem mass spectrometry method for rapid confirmation of medium- and very long-chain acyl-CoA dehydrogenase deficiency in newborns.
BACKGROUND
Newborn screening for medium- and very long-chain acyl-CoA dehydrogenase (MCAD and VLCAD, respectively) deficiency, using acylcarnitine profiling with tandem mass spectrometry, has increased the number of patients with fatty acid oxidation disorders due to the identification of additional milder, and so far silent, phenotypes. However, especially for VLCADD, the acylcarnitine profile can not constitute the sole parameter in order to reliably confirm disease. Therefore, we developed a new liquid chromatography tandem mass spectrometry (LC-MS/MS) method to rapidly determine both MCAD- and/or VLCAD-activity in human lymphocytes in order to confirm diagnosis.


METHODS
LC-MS/MS was used to measure MCAD- or VLCAD-catalyzed production of enoyl-CoA and hydroxyacyl-CoA, in human lymphocytes.


RESULTS
VLCAD activity in controls was 6.95+/-0.42 mU/mg (range 1.95 to 11.91 mU/mg). Residual VLCAD activity of 4 patients with confirmed VLCAD-deficiency was between 0.3 and 1.1%. Heterozygous ACADVL mutation carriers showed residual VLCAD activities of 23.7 to 54.2%. MCAD activity in controls was 2.38+/-0.18 mU/mg. In total, 28 patients with suspected MCAD-deficiency were assayed. Nearly all patients with residual MCAD activities below 2.5% were homozygous 985A>G carriers. MCAD-deficient patients with one other than the 985A>G mutation had higher MCAD residual activities, ranging from 5.7 to 13.9%. All patients with the 199T>C mutation had residual activities above 10%.


CONCLUSIONS
Our newly developed LC-MS/MS method is able to provide ample sensitivity to correctly and rapidly determine MCAD and VLCAD residual activity in human lymphocytes. Importantly, based on measured MCAD residual activities in correlation with genotype, new insights were obtained on the expected clinical phenotype.
ter Veld
Frank
F
Department of General Pediatrics, Heinrich- Heine -University, Düsseldorf, Germany. frank.terveld@uni-duesseldorf.de
Mueller
Martina
M
Kramer
Simone
S
Haussmann
Ulrike
U
Herebian
Diran
D
Mayatepek
Ertan
E
Laryea
Maurice D
MD
Primassin
Sonja
S
Spiekerkoetter
Ute
U
D042964:Acyl-CoA Dehydrogenase / Q000172:deficiency*; D044942:Acyl-CoA Dehydrogenase, Long-Chain / Q000172:deficiency*; D002851:Chromatography, High Pressure Liquid; D006580:Genetic Carrier Screening; D005838:Genotype; D006801:Humans; D007231:Infant, Newborn; D008052:Lipid Metabolism, Inborn Errors / Q000175:diagnosis* / Q000235:genetics; D009154:Mutation; D053719:Tandem Mass Spectrometry / Q000379:methods*
D042964:Acyl-CoA Dehydrogenase; D044942:Acyl-CoA Dehydrogenase, Long-Chain