title: The need for vigilance: false-negative screening for adenylosuccinate lyase deficiency caused by deribosylation of urinary biomarkers.
abstract: OBJECTIVE
  Adenylosuccinate lyase deficiency (dADSL) is a rare inherited metabolic disorder. Biochemical diagnosis of the disease is based on the determination of enormously elevated urinary levels of succinylaminoimidazole carboxamide riboside (SAICA-riboside) and succinyladenosine (SAdo). We report a case of false negative screening for dADSL caused by deribosylation of the urinary biomarkers SAICA-riboside and SAdo.
  
  
  METHODS
  A thin-layer chromatography (TLC) method with Pauly reagent detection of SAICA-riboside was used as a screening method. High-performance liquid chromatography with diode-array detection (HPLC-DAD) and LC-MS/MS methods were used for the identification and quantitative determination of SAICA-riboside, SAdo, succinylaminoimidazole carboxamide (SAICA) and succinyladenine (SA).
  
  
  RESULTS
  Following a negative TLC screening in a known case of dADSL, we analyzed urine using HPLC-DAD. The concentration of SAICA-riboside was 2.7mmol/mol creatinine (below the TLC detection limit), and we detected the two abnormal metabolites identified by LC-MS/MS as SAICA and SA. We showed that SAICA and SA were produced by deribosylation of SAICA-riboside and SAdo in the patient's urine. Studies performed by monitoring the production of SAICA and SA after the addition of SAICA-riboside and SAdo to the patient's urine and to urine samples from patients with urinary tract infections suggested that deribosylation is facilitated by bacterial enzymes.
  
  
  CONCLUSIONS
  Screening methods for the diagnosis of dADSL may be falsely negative due to bacteria-mediated deribosylation of SAICA-riboside and SAdo. HPLC-DAD or LC-MS/MS analyses allowing for simultaneous detection of SAICA-riboside, SAdo and their deribosylation products SAICA and SA should be preferentially used for the diagnosis of dADSL in urine.
authors.affiliation: Institute of Inherited Metabolic Disorders, First Faculty of Medicine, Charles University in Prague and General University Hospital in Prague, Ke Karlovu 2, 128 08 Praha 2, Czech Republic. Electronic address: jkrijt@lf1.cuni.cz.
authors.forename: Jakub
authors.initials: J
authors.lastname: Krijt
authors.forename: Vaclava
authors.initials: V
authors.lastname: Skopova
authors.forename: Vaclava
authors.initials: V
authors.lastname: Adamkova
authors.forename: Renata
authors.initials: R
authors.lastname: Cermakova
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Jurecka
authors.forename: Stanislav
authors.initials: S
authors.lastname: Kmoch
authors.forename: Marie
authors.initials: M
authors.lastname: Zikanova
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives / Q000652:urine; D000264:Adenylosuccinate Lyase / Q000172:deficiency* / Q000652:urine; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000378:metabolism / Q000652:urine; D001321:Autistic Disorder; D001426:Bacterial Proteins / Q000378:metabolism; D002675:Child, Preschool; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer / Q000379:methods; D013293:Enterococcus faecalis; D004798:Enzymes / Q000378:metabolism; D005188:False Negative Reactions; D006801:Humans; D007711:Klebsiella pneumoniae; D011686:Purine-Pyrimidine Metabolism, Inborn Errors / Q000175:diagnosis* / Q000652:urine*; D012263:Ribonucleosides / Q000378:metabolism / Q000652:urine*; D053719:Tandem Mass Spectrometry / Q000379:methods; D014556:Urine / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; D004798:Enzymes; D012263:Ribonucleosides; C043566:succinylaminoimidazole carboxamide riboside; D000620:Aminoimidazole Carboxamide; C043565:succinyladenosine; D000264:Adenylosuccinate Lyase; D000241:Adenosine
keywords: ADSL; Adenylosuccinate lyase deficiency; HPLC-DAD; LC–MS/MS; Purine metabolism; SA; SAICA; SAICA-riboside; SAdo; Succinyladenosine; Succinylaminoimidazole carboxamide riboside; Succinylpurines; TLC; adenylosuccinate lyase; adenylosuccinate lyase deficiency; dADSL; high-performance liquid chromatography with diode array detection; liquid chromatography–tandem mass spectrometry; succinyladenine; succinyladenosine; succinylaminoimidazole carboxamide; succinylaminoimidazole carboxamide riboside; thin-layer chromatography