Approach for Profiling of Glycosphingolipid Glycosylation by Multiplexed Capillary Gel Electrophoresis Coupled to Laser-Induced Fluorescence Detection To Identify Cell-Surface Markers of Human Pluripotent Stem Cells and Derived Cardiomyocytes

Rossdam, Charlotte; Konze, Sarah A.; Oberbeck, Astrid; Rapp, Erdmann; Gerardy‐Schahn, Rita; Itzstein, Mark von; Buettner, Falk F. R.

doi:10.1021/acs.analchem.9b01114

Cited by 32 publications

(40 citation statements)

References 37 publications

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“…While immunodetection is based on in situ detection, mass spectrometry requires the extraction of gangliosides from biological samples before the analysis. Recently, the combination of these two detection methods has allowed a lot of progress regarding ganglioside profiling in sera [23,24], tissues [25], or cellular extracts [26]. Besides, O-acetylation of the sialic acid residues is one of the most common modifications of gangliosides and is sufficient to induce dramatic changes of physiopathological properties carried by the native non-O-acetylated gangliosides.…”

Section: Discussionmentioning

confidence: 99%

Profiling of O-acetylated Gangliosides Expressed in Neuroectoderm Derived Cells

Cavdarli

Yamakawa

Clarisse

et al. 2020

IJMS

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The expression and biological functions of oncofetal markers GD2 and GD3 were extensively studied in neuroectoderm-derived cancers in order to characterize their potential as therapeutic targets. Using immunological approaches, we previously identified GD3, GD2, and OAcGD2 expression in breast cancer (BC) cell lines. However, antibodies specific for O-acetylated gangliosides are not exempt of limitations, as they only provide information on the expression of a limited set of O-acetylated ganglioside species. Consequently, the aim of the present study was to use structural approaches in order to apprehend ganglioside diversity in melanoma, neuroblastoma, and breast cancer cells, focusing on O-acetylated species that are usually lost under alkaline conditions and require specific analytical procedures. We used purification and extraction methods that preserve the O-acetyl modification for the analysis of native gangliosides by MALDI-TOF. We identified the expression of GM1, GM2, GM3, GD2, GD3, GT2, and GT3 in SK-Mel28 (melanoma), LAN-1 (neuroblastoma), Hs 578T, SUM 159PT, MDA-MB-231, MCF-7 (BC), and BC cell lines over-expressing GD3 synthase. Among O-acetylated gangliosides, we characterized the expression of OAcGM1, OAcGD3, OAcGD2, OAcGT2, and OAcGT3. Furthermore, the experimental procedure allowed us to clearly identify the position of the sialic acid residue that carries the O-acetyl group on b- and c-series gangliosides by MS/MS fragmentation. These results show that ganglioside O-acetylation occurs on both inner and terminal sialic acid residue in a cell type-dependent manner, suggesting different O-acetylation pathways for gangliosides. They also highlight the limitation of immuno-detection for the complete identification of O-acetylated ganglioside profiles in cancer cells.

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Section: Discussionmentioning

confidence: 99%

Profiling of O-acetylated Gangliosides Expressed in Neuroectoderm Derived Cells

Cavdarli

Yamakawa

Clarisse

et al. 2020

IJMS

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“…For glycolipid analysis, HT29 and PaCa5061 cells were cultivated in the presence or absence of GalNAc-α- O -benzyl as described above and equal cell numbers harvested for glycolipid extraction. Glycolipid extraction and subsequent xCGE-LIF analysis was performed as previously described by Rossdam et al ( Rossdam et al 2019 ). Briefly, the cells were lysed by sonication and glycolipids were extracted in chloroform/methanol (1:2 (v/v)).…”

Section: Methodsmentioning

confidence: 99%

Systematic analysis of the human tumor cell binding to human vs. murine E- and P-selectin under static vs. dynamic conditions

et al. 2020

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Endothelial E- and P-selectins promote metastasis formation by interacting with sialyl-Lewis X and A (sLeX/sLeA) on circulating tumor cells. This interaction precedes extravasation and can take place under dynamic and static conditions. Metastasis formation is often studied in xenograft models. However, it is unclear whether species differences exist in the ligand specificity of human (h) vs. murine (m) selectins and whether different ligands are functional under dynamic vs. static conditions. We systematically compared the h vs. m E- and P-selectin (ESel/PSel) binding of a range of human tumor cells under dynamic vs. static conditions. The tumor cells were categorized by their sLeA/X status (sLeA+/sLeX+, sLeA−/sLeX+ and sLeA−/sLeX−). The general biological nature of the tumor–selectin interaction was analyzed by applying several tumor cell treatments (anti-sLeA/X blockade, neuraminidase, pronase and inhibition of O/N-glycosylation). We observed remarkable differences in the static vs. dynamic interaction of tumor cells with h vs. m ESel/PSel depending on their sLeA/X status. The tumor cell treatments mostly affected either static or dynamic as well as either h- or m-selectin interaction. mESel showed a higher diversity of potential ligands than hESel. Inhibition of O-GalNAc-glycosylation also affected glycosphingolipid synthesis. Summarized, different ligands on human tumor cells are functional under static vs. dynamic conditions and for the interaction with human vs. murine ESel/PSel. Non-canonical selectin ligands lacking the sLeA/X glycan epitopes exist on human tumor cells. These findings have important implications for the current development of glycomimetic, antimetastatic drugs and encourage the development of immunodeficient mice with humanized selectins.

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“…This method is based on the building of a database using migration times for commercially available glycan species, and on the comparison of the retention times of biological samples to this library. xCGE-LIF allowed the characterization of GM3 and GD3 at the surface of stem-cell-derived cardiomyocytes [63].…”

Section: Analysis Of O-acetylated and Non-o-acetylated Gangliosides Smentioning

confidence: 99%

O-acetylated Gangliosides as Targets for Cancer Immunotherapy

Cavdarli

Delannoy

Groux-Degroote

2020

Cells

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O-acetylation of sialic acid residues is one of the main modifications of gangliosides, and modulates ganglioside functions.O-acetylation of gangliosides is dependent on sialyl-O-acetyltransferases and sialyl-O-acetyl-esterase activities. CAS1 Domain-Containing Protein 1 (CASD1) is the only human sialyl-O-acetyltransferases (SOAT) described until now. O-acetylated ganglioside species are mainly expressed during embryonic development and in the central nervous system in healthy adults, but are re-expressed during cancer development and are considered as markers of cancers of neuroectodermal origin. However, the specific biological roles of O-acetylated gangliosides in developing and malignant tissues have not been extensively studied, mostly because of the requirement of specific approaches and tools for sample preparation and analysis. In this review, we summarize our current knowledge of ganglioside biosynthesis and expression in normal and pathological conditions, of ganglioside O-acetylation analysis and expression in cancers, and of the possible use of O-acetylated gangliosides as targets for cancer immunotherapy. a potential functional significance of ganglioside O-acetylation in cancer. The use of O-acetylated gangliosides as targets for cancer immunotherapy is also discussed. Ganglioside Structures and BiosynthesisGangliosides are acidic GSL containing from one to five sialic acids residues. They are mainly, but not exclusively, derived from ganglio-series GSL (GgCer) that result from the substitution of a ceramide (Cer) by the tetrasaccharide core Galβ1-3GalNAcβ1-4Galβ1-4Glcβ (Gg4). In different cell types and tissues, gangliosides are usually found as a mixture of di-, tri-and tetrasaccharide cores substituted by one or more sialic acid residues [1,5]. The biosynthesis of gangliosides takes place in the Golgi apparatus and starts by the transfer of sialic acid residues to lactosylceramide (LacCer) by specific sialyltransferases, i.e., the CMP-Neu5Ac: LacCer α2,3-sialyltransferase ST3Gal V (GM3 synthase) [6], the CMP-Neu5Ac: GM3 α2,8-sialyltransferase ST8Sia I (GD3 synthase) [7] and the CMP-Neu5Ac: GD3 α2,8-sialyltransferase ST8Sia V [8], that show high specificity toward glycolipid substrates [9]. Thus, LacCer, GM3, GD3 and GT3 are the precursors for 0-, a-, b-and c-series gangliosides, respectively (Figure 1).

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Approach for Profiling of Glycosphingolipid Glycosylation by Multiplexed Capillary Gel Electrophoresis Coupled to Laser-Induced Fluorescence Detection To Identify Cell-Surface Markers of Human Pluripotent Stem Cells and Derived Cardiomyocytes

Cited by 32 publications

References 37 publications

Profiling of O-acetylated Gangliosides Expressed in Neuroectoderm Derived Cells

Profiling of O-acetylated Gangliosides Expressed in Neuroectoderm Derived Cells

Systematic analysis of the human tumor cell binding to human vs. murine E- and P-selectin under static vs. dynamic conditions

O-acetylated Gangliosides as Targets for Cancer Immunotherapy

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