Comment[ArrayExpressAccession] E-GEOD-50767 MAGE-TAB Version 1.1 Public Release Date 2013-10-10 Investigation Title Anaplastic thyroid carcinoma cell lines (HTH83 and TL3) infected with LAMC2 shRNA Comment[Submitted Name] Anaplastic thyroid carcinoma cell lines (HTH83 and TL3) infected with LAMC2 shRNA Experiment Description Laminin-5 gamma-2 (LAMC2) is highly expressed in anaplastic thyroid carcinoma and associated with tumor progression, migration and invasion by modulating signaling of EGFR LAMC2 was highly expressed in ATC samples and cell lines compared to normal thyroid tissues. Silencing LAMC2 by shRNA in ATC cells moderately inhibited cell growth in liquid culture and dramatically decreased growth in soft agar and in xenografts growing in immunodeficient mice. Silencing LAMC2 caused cell cycle arrest and significantly suppressed migration, invasion and wound healing of ATC cells. Rescue experiments by overexpressing LAMC2 in LAMC2 knockdown cells, reversed the inhibitory effects as shown by increased cell proliferation and colony formation. Microarray data demonstrated that LAMC2 shRNA significantly altered expression of genes associated with migration, invasion, proliferation and survival. Immunoprecipitation studies showed that LAMC2 was bound to EGFR in ATC cells. Silencing of LAMC2 partially blocked EGF-mediated activation of EGFR and its downstream pathway. Anaplastic thyroid carcinoma (ATC) is an aggressive malignancy having no effective treatment. Laminin subunit gamma-2 (LAMC2) is an epithelial basement membrane protein involved in cell migration and tumour invasion and might represent an ideal target for the development of novel therapeutic approaches for ATC. LAMC2 was highly expressed in ATC samples and cell lines compared to normal thyroid tissues. Silencing LAMC2 by shRNA in ATC cells moderately inhibited cell growth in liquid culture and dramatically decreased growth in soft agar and in xenografts growing in immunodeficient mice. Silencing LAMC2 caused cell cycle arrest and significantly suppressed migration, invasion and wound healing of ATC cells. Rescue experiments by overexpressing LAMC2 in LAMC2 knockdown cells, reversed the inhibitory effects as shown by increased cell proliferation and colony formation. Microarray data demonstrated that LAMC2 shRNA significantly altered expression of genes associated with migration, invasion, proliferation and survival. Immunoprecipitation studies showed that LAMC2 was bound to EGFR in ATC cells. Silencing of LAMC2 partially blocked EGF-mediated activation of EGFR and its downstream pathway. LAMC2 was highly expressed in ATC samples and cell lines compared to normal thyroid tissues. Silencing LAMC2 by shRNA in ATC cells moderately inhibited cell growth in liquid culture and dramatically decreased growth in soft agar and in xenografts growing in immunodeficient mice. Silencing LAMC2 caused cell cycle arrest and significantly suppressed migration, invasion and wound healing of ATC cells. Rescue experiments by overexpressing LAMC2 in LAMC2 knockdown cells, reversed the inhibitory effects as shown by increased cell proliferation and colony formation. Microarray data demonstrated that LAMC2 shRNA significantly altered expression of genes associated with migration, invasion, proliferation and survival. Immunoprecipitation studies showed that LAMC2 was bound to EGFR in ATC cells. Silencing of LAMC2 partially blocked EGF-mediated activation of EGFR and its downstream pathway. Anaplastic thyroid carcinoma cell lines (HTH83 and TL3) were infected with scrambled shRNA and LAMC2 shRNA and stable clones from each cell line were generated and used for RNA extraction and hybridization on Illumina Microarray. We compared scrambled shRNA stable cells with LAMC2 shRNA stable cells. Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Liu Garg Kanojia Okamoto Jain Madan Chien Sampath Ding Xuan Said Doan Liu Yang Gery Braunstein Koeffler Person First Name Lizhen Manoj Deepika Ryoko Saket Vikas Wenwen Abhishek Ling-Wen Meng Jonathan Ngan Li-Zhen Henry Sigal Glenn H Person Mid Initials W B D P Person Email csiliul@nus.edu.sg Person Affiliation Cancer Science Institute Person Address Cancer Science Institute, 14 Medical Drive, Singapore, Singapore Person Roles submitter Protocol Name P-GSE50767-1 P-GSE50767-5 P-GSE50767-6 P-GSE50767-2 P-GSE50767-3 P-GSE50767-4 P-GSE50767-7 Protocol Description The data were analyzed using RMA in the bioconductor and normalized using the Cross Correlation method. The trimmed mean target intensity of each array was set to 8. ID_REF = VALUE = normalized intensity cRNA were synthesized using Illumina Total Prep RNA Amplification Kit (Illumina) Bitinylated cRNA were fragmented and hybridised to an illumina whole genome expression chip, Human HT-12 v4.0 for 18 h at 58M-0C. Beadchips were then washed and stained and subsequently scanned to obtain fluorescence intensities. Three different clones from TL3 and HTH 83 cell lines stably expressing scramble shRNA and LAMC2 shRNA were used Cells were grown under normal culture conditions for 3 day Total RNA was isolated from cultured cells with RNeasy mini kit (Qiagen GmbH, Hilden, Germany) according to the manufacturerM-bM-^@M-^Ys protocol. Gene Chips were scanned using an Illumina Bead Array Reader Protocol Type normalization data transformation protocol labelling protocol hybridization protocol sample treatment protocol growth protocol nucleic acid extraction protocol array scanning protocol Experimental Factor Name CELL LINE EXPRESSION Experimental Factor Type cell line expression Comment[SecondaryAccession] GSE50767 Comment[GEOReleaseDate] 2013-10-10 Comment[ArrayExpressSubmissionDate] 2013-09-11 Comment[GEOLastUpdateDate] 2013-10-10 Comment[AEExperimentType] transcription profiling by array Comment[AdditionalFile:Data1] GSE50767_non-normalized.txt SDRF File E-GEOD-50767.sdrf.txt