title: Elevated plasma dihydroorotate in Miller syndrome: Biochemical, diagnostic and clinical implications, and treatment with uridine.
abstract: Miller syndrome (post-axial acrofacial dysostosis) arises from gene mutations for the mitochondrial enzyme dihydroorotate dehydrogenase (DHODH). Nonetheless, despite demonstrated loss of enzyme activity dihydroorotate (DHO) has not been shown to accumulate, but paradoxically urine orotate has been reported to be raised, confusing the metabolic diagnosis.
  
  
  
  We analysed plasma and urine from a 4-year-old male Miller syndrome patient. DHODH mutations were determined by PCR and Sanger sequencing. Analysis of DHO and orotic acid (OA) in urine, plasma and blood-spot cards was performed using liquid chromatography-tandem mass spectrometry. In vitro stability of DHO in distilled water and control urine was assessed for up to 60h. The patient received a 3-month trial of oral uridine for behavioural problems.
  
  
  
  The patient had early liver complications that are atypical of Miller syndrome. DHODH genotyping demonstrated compound-heterozygosity for frameshift and missense mutations. DHO was grossly raised in urine and plasma, and was detectable in dried spots of blood and plasma. OA was raised in urine but undetectable in plasma. DHO did not spontaneously degrade to OA. Uridine therapy did not appear to resolve behavioural problems during treatment, but it lowered plasma DHO.
  
  
  
  This case with grossly raised plasma DHO represents the first biochemical confirmation of functional DHODH deficiency. DHO was also easily detectable in dried plasma and blood spots. We concluded that DHO oxidation to OA must occur enzymatically during renal secretion. This case resolved the biochemical conundrum in previous reports of Miller syndrome patients, and opened the possibility of rapid biochemical screening.
authors.affiliation: School of Pharmacy and Mater Research Institute, The University of Queensland, Brisbane, QLD 4102, Australia. Electronic address: jduley@pharmacy.uq.edu.au.
authors.forename: John A
authors.initials: JA
authors.lastname: Duley
authors.affiliation: Department of Pathology, Mater Health Services, Brisbane, QLD 4101, Australia. Electronic address: michael.henman@mater.org.au.
authors.forename: Michael G
authors.initials: MG
authors.lastname: Henman
authors.affiliation: NSW Biochemical Genetics Service, The Children's Hospital at Westmead, Disciplines of Genetic Medicine & Child and Adolescent Health, The University of Sydney, NSW 2145, Australia. Electronic address: kevin.carpenter@health.nsw.gov.au.
authors.forename: Kevin H
authors.initials: KH
authors.lastname: Carpenter
authors.affiliation: Department of Pediatrics, University of Washington, Division of Genetic Medicine at Seattle Children's Hospital, Seattle, WA 98195, USA. Electronic address: mbamshad@u.washington.edu.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Bamshad
authors.affiliation: Department of Pathology, Mater Health Services, Brisbane, QLD 4101, Australia. Electronic address: george.marshall@mater.org.au.
authors.forename: George A
authors.initials: GA
authors.lastname: Marshall
authors.affiliation: School of Women's and Children's Health, University of NSW, Sydney Children's Hospital, Sydney, NSW 2031, Australia; School of Medicine, University of NSW, Sydney Children's Hospital, Sydney, NSW, 2031, Australia. Electronic address: keith.ooi@unsw.edu.au.
authors.forename: Chee Y
authors.initials: CY
authors.lastname: Ooi
authors.affiliation: Department of Medical Genetics, Sydney Children's Hospital, University of Sydney, NSW 2031, Australia. Electronic address: bridget.wilcken@health.nsw.gov.au.
authors.forename: Bridget
authors.initials: B
authors.lastname: Wilcken
authors.affiliation: Department of Medical Genomics, Royal Prince Alfred Hospital, The University of Sydney, NSW 2050, Australia. Electronic address: jason.pinner@sswahs.nsw.gov.au.
authors.forename: Jason R
authors.initials: JR
authors.lastname: Pinner
mesh_terms: D000015:Abnormalities, Multiple / Q000097:blood / Q000235:genetics* / Q000503:physiopathology / Q000652:urine; D002675:Child, Preschool; D000090722:Dihydroorotate Dehydrogenase; D005838:Genotype; D006801:Humans; D017880:Limb Deformities, Congenital / Q000097:blood / Q000235:genetics* / Q000503:physiopathology / Q000652:urine; D008297:Male; D008342:Mandibulofacial Dysostosis / Q000097:blood / Q000235:genetics* / Q000503:physiopathology / Q000652:urine; D008844:Micrognathism / Q000097:blood / Q000235:genetics* / Q000503:physiopathology / Q000652:urine; D009154:Mutation; D009963:Orotic Acid / Q000031:analogs & derivatives* / Q000097:blood / Q000652:urine; D010084:Oxidation-Reduction; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000097:blood / Q000235:genetics* / Q000652:urine; D014529:Uridine / Q000097:blood / Q000652:urine
chemical_list: D000090722:Dihydroorotate Dehydrogenase; C004768:4,5-dihydroorotic acid; D009963:Orotic Acid; D044925:Oxidoreductases Acting on CH-CH Group Donors; D014529:Uridine
keywords: Dihydroorotate-dehydrogenase; Miller syndrome; Mitochondria; POADS; Screening