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Kidney Week

Abstract: TH-PO720

Development of a 2D-UPLC-MS/MS Assay for Therapeutic Monitoring in Patients with APRT Deficiency

Session Information

Category: Genetic Diseases of the Kidney

  • 1002 Genetic Diseases of the Kidney: Non-Cystic


  • Edvardsson, Vidar O., Landspital - The National University Hospital of Iceland, Reykjavik, Iceland
  • Thorsteinsdottir, Unnur A., University of Iceland, Reykjavik, Iceland
  • Runolfsdottir, Hrafnhildur L., Landspital - The National University Hospital of Iceland, Reykjavik, Iceland
  • Bjarnason, Thorsteinn Hjörtur, University of Iceland, Reykjavik, Iceland
  • Eiriksson, Finnur Freyr, ArcticMass, Reykjavik, Iceland
  • Thorsteinsdottir, Margret, University of Iceland, Reykjavik, Iceland
  • Palsson, Runolfur, Landspital - The National University Hospital of Iceland, Reykjavik, Iceland

Group or Team Name

  • The APRT Deficiency Program of the Rare Kidney Stone Consortium

Adenine phosphoribosyltransferase deficiency (APRTd) is a rare hereditary purine metabolism disorder resulting in accumulation and renal excretion of 2,8-dihydroxyadenine (DHA), causing kidney stones and chronic kidney disease (CKD). The drugs allopurinol and febuxostat reduce urinary DHA excretion and ameliorate the disease manifestations. We developed a two-dimensional ultra-performance liquid chromatography tandem mass spectrometry (2D-UPLC-MS/MS) assay for simultaneous measurement of DHA and the pharmacological agents allopurinol, its active metabolite oxypurinol, and febuxostat in human plasma for monitoring of pharmacotherapy.


A design of experiments was used to define the optimum 2D-UPLC-MS/MS conditions based on the minimum number of experiments. Experimental screening of the variables was performed by D-optimal design to identify factors influencing retention time, peak height and peak area for all compounds. Variables were optimized with central composite face design and related to sensitivity and retention time using partial least square regression. Protein precipitation was carried out with 1% formic acid in methanol as a crash solvent. Plasma samples from APRTd patients and control specimens from laboratory personnel were used for the study.


The plasma concentration of DHA, allopurinol, oxypurinol, and febuxostat was reliably achieved with the 2D-UPLC-MS/MS assay. The DHA concentrations in plasma from 3 APRTd patients were 456, 459 and 741 ng/mL off drug treatment and 130, 61 and 27 ng/mL while on allopurinol or febuxostat therapy. By contrast, DHA was not detected in plasma from healthy controls. The plasma allopurinol and oxypurinol concentration during drug treatment in the 3 patients was 802, 2391, 5089 and 6810, 7076 and 12,510 ng/mL, respectively, and the febuxostat concentration was 1276 ng/mL in 1 patient.


Our data suggest that the 2D-UPLC-MS/MS assay may be used to monitor the efficacy of pharmacotherapy and treatment adherence among APRTd patients. This method may prove useful in determining the plasma DHA concentration that must be attained in order to prevent new kidney stone growth and CKD progression in these patients.


  • NIDDK Support