JIMD Reports, Volume 37

Lisette M. Berends, Eduard A. Struys, Birthe Roos, Ulbe Holwerda, Erwin E. W. Jansen, Gajja S. Salomons, Mirjam M. C. Wamelink, Berends, Lisette M., Struys, Eduard A., Roos, Birthe, Holwerda, Ulbe, Jansen, Erwin E. W., Salomons, Gajja S., Wamelink, Mirjam M. C.

Guanidinoacetate methyltransferase (GAMT) deficiency is an inborn error of metabolism (IEM), clinically characterized by intellectual disability, developmental delay, seizures, and movement disorders. Biochemical diagnosis of GAMT deficiency is based on the measurement of creatine and guanidinoacetate in urine, plasma, or CSF and is confirmed genetically by DNA analysis or by enzyme assay in lymphoblasts or fibroblasts. To obtain enough cells, these cells need to be cultured for at least 1 month. A less time-consuming diagnostic functional test is needed, since GAMT deficiency is a candidate for newborn screening (NBS) programs, to be able to confirm or rule out this IEM after an initial positive result in the NBS. Stable-isotope-labeled (13)C2-guanidinoacetate and (2)H3-S-adenosylmethionine (SAM) were used, which are converted by GAMT present in lymphocyte extracts into (2)H3-(13)C2-creatine. The formed (2)H3-(13)C2-creatine was butylated and subsequently measured by liquid chromatography tandem mass-spectrometry (LC-MS/MS). We measured GAMT enzyme activity in lymphocyte extracts of 24 controls, 3 GAMT deficient patients and of 2 parents proven to be carrier. Because GAMT activity decreases when isolation time after venipuncture increases, reference values were obtained for 2 control groups: isolation on the day of venipuncture (27-130 pmol/h/mg) and 1 day afterwards (15-146 pmol/h/mg). Deficient patients had no detectable GAMT activity. The two carriers had GAMT activity within the normal range. We designed a fast, less invasive, and valid method to measure GAMT activity in lymphocytes using LC-MS/MS analysis without the need of time-consuming and laborious cell culture.