The presence of
gangliosides containing de-N-acetylated
sialic acids in human tissues has been so far shown by using mouse
monoclonal antibodies specific for the de-N-acetylated forms, but the isolation and chemical characterization of such compounds have not yet been performed. Since indirect evidence suggested that de-N-acetylGD3
ganglioside could be present in human
melanoma tumors, we analyzed the
gangliosides purified from a 500-g pool of those
tumors. The de-N-acetylGD3 that was found to migrate just below GD2 in thin-layer chromatography was isolated from the
disialogangliosides by high-pressure liquid chromatography using the specific antibody SGR37 to monitor the elution. The amount of
antigen was found to be 320 ng per gram of fresh
tumor or 0.1% of total
gangliosides. Gas chromatography-mass spectrometry analysis of the antibody-positive
ganglioside showed that
sialic acids were formed of one molecule of
N-acetylneuraminic acid and one molecule of neuraminic
acid. Radioactive re-N-acetylation of the
antigen yielded a GD3-like
ganglioside with the radioactive label on the external
sialic acid. The constitutive
fatty acids were found to differ markedly from those of GD3 and 9-O-acetylGD3 isolated from the same pool of
tumors. The major
fatty acids were C16:0 and C18:0 in de-N-acetylGD3, whereas GD3 and its 9-O-acetylated derivative contained a large amount of C24:1. These data show that de-N-acetylGD3
ganglioside is indeed present in human
melanoma tumors, and the
fatty acid content suggests the existence of a de-N-
acetylase mostly active on the molecular species of
gangliosides with
short-chain fatty acids.