Browsing by Author "Hoermann, Patrick"

Genes that are highly overexpressed in tumor cells can be required for tumor cell survival and have the potential to be selective therapeutic targets. In an attempt to identify such targets, we combined a functional genomics and a systems biology approach to assess the consequences of RNAi-mediated silencing of overexpressed genes that were selected from 140 gene expression profiles from colorectal cancers (CRCs) and matched normal mucosa. In order to identify credible models for in-depth functional analysis, we first confirmed the overexpression of these genes in 25 different CRC cell lines. We then identified five candidate genes that profoundly reduced the viability of CRC cell lines when silenced with either siRNAs or short-hairpin RNAs (shRNAs), i.e., HMGA1, TACSTD2, RRM2, RPS2 and NOL5A. These genes were further studied by systematic analysis of comprehensive gene expression profiles generated following siRNA-mediated silencing. Exploration of these RNAi-specific gene expression signatures allowed the identification of the functional space in which the five genes operate and showed enrichment for cancer-specific signaling pathways, some known to be involved in CRC. By comparing the expression of the RNAi signature genes with their respective expression levels in an independent set of primary rectal carcinomas, we could recapitulate these defined RNAi signatures, therefore, establishing the biological relevance of our observations. This strategy identified the signaling pathways that are affected by the prominent oncogenes HMGAI and TACSTD2, established a yet unknown link between RRM2 and PLK1 and identified RPS2 and NOL5A as promising potential therapeutic targets in CRC.

Genomic aberrations on chromosome 8 are common in colon cancer, and are associated with lymph node and distant metastases as well as with disease susceptibility. This prompted us to generate a high-resolution map of genomic imbalances of chromosome 8 in 51 primary colon carcinomas using a custom-designed genomic array consisting of a tiling path of BAC clones. This analysis confirmed the dominant role of this chromosome. Unexpectedly, the position of the breakpoints suggested colocalization with structural variants in the human genome. In order to map these sites with increased resolution and to extend the analysis to the entire genome, we analyzed a subset of these tumors (n = 32) by comparative genomic hybridization on a 185K oligonucleotide array platform. Our comprehensive map of the colon cancer genome confirmed recurrent and specific low-level copy number changes of chromosomes 7, 8, 13, 18, and 20, and unveiled additional, novel sites of genomic imbalances including amplification of a histone gene cluster on chromosome 6p21.1-21.33 and deletions on chromosome 4q34-35. The systematic comparison of segments of copy number change with gene expression profiles showed that genomic imbalances directly affect average expression levels. Strikingly, we observed a significant association of chromosomal breakpoints with structural variants in the human genome: 41% of all copy number changes occurred at sites of such copy number variants (P < 2.2e(-16)). Such an association has not been previously described and reveals a yet underappreciated plasticity of the colon cancer genome; it also points to potential mechanisms for the induction of chromosomal breakage in cancer cells.

To characterize patterns of global transcriptional deregulation in primary colon carcinomas, we did gene expression profiling of 73 tumors [Unio Internationale Contra Cancrum stage II (n = 33) and stage III (n = 40)] using oligonucleotide microarrays. For 30 of the tumors, expression profiles were compared with those from matched normal mucosa samples. We identified a set of 1,950 genes with highly significant deregulation between tumors and mucosa samples (P < 1e-7). A significant proportion of these genes mapped to chromosome 20 (P = 0.01). Seventeen genes had a > 5-fold average expression difference between normal colon mucosa and carcinomas, including up-regulation of MYC and of HMGA1, a putative oncogene. Furthermore, we identified 68 genes that were significantly differentially expressed between lymph node-negative and lymph node-positive tumors (P < 0.001), the functional annotation of which revealed a preponderance of genes that play a role in cellular immune response and surveillance. The microarray-derived gene expression levels of 20 deregulated genes were validated using quantitative real-time reverse transcription-PCR in > 40 tumor and normal mucosa samples with good concordance between the techniques. Finally, we established a relationship between specific genomic imbalances, which were mapped for 32 of the analyzed colon tumors by comparative genomic hybridization, and alterations of global transcriptional activity. Previously, we had conducted a similar analysis of primary rectal carcinomas. The systematic comparison of colon and rectal carcinomas revealed a significant overlap of genomic imbalances and transcriptional deregulation, including activation of the Wnt/beta-catenin signaling cascade, suggesting similar pathogenic pathways.