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Series GSE12422 Query DataSets for GSE12422
Status Public on Aug 18, 2008
Title O-glycan inhibitors generate aryl-glycans, induce apoptosis, and inhibit growth in colorectal cancer cell lines
Organism Homo sapiens
Experiment type Expression profiling by array
Summary Our studies provide direct evidence that O-glycosylation pathways play a role in the regulation of cell growth through apoptosis and proliferation pathways. Eight small molecular weight analogues of the GalNAc-alpha-1-O-serine/threonine structure based on 1-benzyl-2-acetamido-2- deoxy-alpha-O-D-galactopyranoside have been synthesised and tested in 5 human colorectal cancer cell lines. Three inhibitors, 1-benzyl-2-acetamido-2-deoxy-alpha-O-D-galactopyranoside and the corresponding 2-azido- and C-glycoside analogues, were screened in two colorectal cancer cell lines at 0.5mM and showed induction of apoptosis. Proliferation was down regulated in the same two cell lines with all three inhibitors, as detected by Ki67 staining and gene array. Treatment both cell lines with inhibitors led to changes in glycosylation detected with peanut lectin. The competitive action of the inhibitors resulted in the intracellular formation of 28 aryl-glycan products which were identified by MALDI and electrospray mass spectroscopy. The structures found map onto known O-glycosylation biosynthetic pathways and showed a differential pattern for each of the inhibitors in both cell lines. Gene array analysis of the glycogenes illustrated a pattern of glycosytransferases that matched the glycan structures found in glycoproteins and aryl-glycans formed in the PC/AA/C1/SB10C cells, however there was no action of the three inhibitors on glycogene transcript levels. The inhibitors act at both intermediary metabolic and genomic levels, resulting in altered protein glycosylation and arylglycan formation. These events may play a part in growth arrest.

Keywords: Response to inhibitors, Apoptosis, Aryl-glycans, Benzyl-O-GalNAc, Growth Inhibition, O-Glycans
Overall design The effects of the O-glycan inhibitors Benzyl 2-azido-2-deoxy-alpha-D-galactopyranoside (alpha-OBn GalN3), Benzyl 2-acetamido-2-deoxy-alpha-D-galactopyranoside (alpha-OBn GalNAc), and 2-(2-acetamido-2-deoxy-alpha-D-galactopyranosyl)-1-phenylethane (alpha-CBn GalNAc) on the colorectal cancer cell line PC/AA/C1/SB10C were examined.
The PC/AA/C1/SB10C cells were treated with with 0.5 mM of the inhibitors for four days. In control experiments cells were cultivated without inhibitor. All experiments were performed twice. Total RNA was extracted with RNAzol reagent in accordance to the manufacturer instructions (PeQLab, Biotechnology Fareham, UK), treated with RNase-free DNase (DNA-free, Ambion,Warrington, UK) and integrity verified using the Agilent Bioanalyzer (Agilent Technologies, West Lothian, UK), by the presence of the 28S and 18S rRNA on agarose gels and an A260/280 ratio in the range of 1.9–2.1. A sample (5 μg) of total RNA was used for production of biotinylated cRNA as described in the Affymetrix GeneChip analysis instruction manual (Affymetrix UK, High Wycombe, UK). The human genome 133A 2.0 array was then hybridized with the biotin-labeled cRNA fragments for 16 h at 45°C. Washing steps for the chip, staining with streptavidin-phycoerythrin, signal amplification and scanning were performed according to the manufacturer’s instructions (Affymetrix UK, High Wycombe, UK). Signal values were exported with the GeneChip operating software (GCOS, Affymetrix). Further analyses were performed with the software “CorrXpression”, which is described in detail elsewhere (Klein et al. 2005, J Mol Med. 83:362-376; Wessel et al. 2006, In Silico Biol.6:61-70).
Web link
Contributor(s) Patsos G, Klein A, Corfield A
Citation(s) 19122213
Submission date Aug 12, 2008
Last update date Dec 06, 2018
Contact name Andreas Klein
Phone +49 30 450 528087
Fax +49 30 450 52945
Organization name Charité (CCO)
Department Institute of Biochemistry
Street address Charitéplatz 1
City Berlin
ZIP/Postal code 10117
Country Germany
Platforms (1)
GPL571 [HG-U133A_2] Affymetrix Human Genome U133A 2.0 Array
Samples (8)
GSM312043 SB10C_Control_1
GSM312044 SB10C_alphaOBnGalN3_1
GSM312045 SB10C_alphaOBnGalNAc_1
BioProject PRJNA113747

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Supplementary file Size Download File type/resource
GSE12422_RAW.tar 14.6 Mb (http)(custom) TAR (of CEL)
Processed data included within Sample table

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