Liquid chromatography/electrospray ionisation-tandem mass spectrometry quantification of G_M2 gangliosides in human peripheral cells and plasma

G_M2 gangliosidosis is a group of inherited neurodegenerative disorders resulting primarily from the excessive accumulation of G_M2 gangliosides (G_M2) 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 G _M2 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 G_M2 was achieved using LC/ESI-MS/MS with d₃₅G_M1 18:1/18:0 as an internal standard. The assay was linear over the biological range, and all G_M2 gangliosidosis patients were demarcated from controls by elevated G_M2 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 G _M2 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 G_M2 gangliosidosis by allowing quantification of a reduction in the primary storage burden.