T1717 The Combined Analysis of miRNA and mRNA Expression Levels in Normal and Neoplastic Colorectal Tissues Allows the Identification of Putative miRNA Target Transcripts

Abstract

G A A b st ra ct s RNA was isolated from 105 specimens and evaluated on Affymetrics GeneChip HG-U133 plus 2.0 microarrays. For proteomic and kinomic analyses, the total tissue protein was extracted from pulled 5-7 samples of each study group. The iTRAQ tagging technology and isoelectrofocusing combined with tandem mass spectrometry (MS) analysis were applied to estimate the relative protein/peptide abundance. A kinase activity was defined as the relative level of phosphorylation of peptide substrates provided by PepChip Kinomics array (Pepscan Presto B.V.). The data were evaluated using pair-wise comparisons, clustering analyses and data decomposition into SVD modes. Results: From 25410 probe sets which passed the filtering procedure according to GCRMA+LVS algorithm, 824 probe sets represented 424 PK genes. From 6162 proteins identified in MS analyses (peptides score ≥ 40), 177 represented PKs. Of them, 157 were also identified by microarray analysis. Altogether, 444 PK transcripts and/or PK proteins were identified. Expression of 64% of transcripts, the amount of 26% of proteins and phosphorylation level of 34% of peptide substrates were changed in Ad, CRC or both tissues, compared to NM. The highest level of transcript, protein and phosphorylation changes were observed in the NM-CRC, Ad-CRC, and NM-Ad paircomparisons, respectively. Low level of correlation between transcript and protein levels was observed, pointing the utility of combined approach for a kinome study. Conclusion: The combination of multiple approaches defined by integrative genomics offers a powerful tool to elucidate changes in the kinome and cell signaling during neoplastic progression to CRC. Supported by PBZ-MNiI-2/1/2005 grant.