Liquid chromatography/electrospray ionisation-tandem mass spectrometry quantification of GM2 gangliosides in human peripheral cells and plasma.
GM2 gangliosidosis is a group of inherited neurodegenerative disorders resulting primarily from the excessive accumulation of GM2 gangliosides (GM2) in neuronal cells. As biomarkers for categorising patients and monitoring the effectiveness of developing therapies are lacking for this group of disorders, we sought to develop methodology to quantify GM2 levels in more readily attainable patient samples such as plasma, leukocytes, and cultured skin fibroblasts. Following organic extraction, gangliosides were partitioned into the aqueous phase and isolated using C18 solid-phase extraction columns. Relative quantification of three species of GM2 was achieved using LC/ESI-MS/MS with d35GM1 18:1/18:0 as an internal standard. The assay was linear over the biological range, and all GM2 gangliosidosis patients were demarcated from controls by elevated GM2 in cultured skin fibroblast extracts. However, in leukocytes only some molecular species could be used for differentiation and in plasma only one was informative. A reduction in GM2 was easily detected in patient skin fibroblasts after a short treatment with media from normal cells enriched in secreted β-hexosaminidase. This method may show promise for measuring the effectiveness of experimental therapies for GM2 gangliosidosis by allowing quantification of a reduction in the primary storage burden.
Genetics and Molecular Pathology, SA Pathology, Adelaide, SA 5006, Australia; Department of Paediatrics, The University of Adelaide, Adelaide, SA 5005, Australia. Electronic address: maria.fuller@adelaide.edu.au.
Maria
M
Fuller
Lysosomal Diseases Research Unit, SAHMRI, Adelaide, SA 5000, Australia.
Stephen
S
Duplock
Lysosomal Diseases Research Unit, SAHMRI, Adelaide, SA 5000, Australia.
Leanne K
LK
Hein
Genetics and Genome Biology, The Hospital for Sick Children, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada.
Brigitte A
BA
Rigat
Genetics and Genome Biology, The Hospital for Sick Children, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario M5G 2C4, Canada.
Don J
DJ
Mahuran
D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D005347:Fibroblasts / Q000737:chemistry; D005678:G(M2) Ganglioside / Q000032:analysis* / Q000097:blood / Q000302:isolation & purification; D006801:Humans; D007962:Leukocytes / Q000737:chemistry; D052616:Solid Phase Extraction; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
D005678:G(M2) Ganglioside; D001619:beta-N-Acetylhexosaminidases
Ganglioside; Mass spectrometry; Sandhoff disease; Tay-Sachs disease