By systematically integrating gene expression profiles and genomic imbalances in the tumors using comparative genomic hybridization, we could show that DNA copy number gains of recurrently aneuploid chromosome arms 7p, 8q, 13q, 18q, 20pand 20q correlated significantly with their average chromosome arm expression profile. Taken together, our results demonstrate that both the high-level, significant transcriptional deregulation of specific genes and general modification of the average transcriptional activity of genes residing on aneuploid chromosomes coexist in rectal adenocarcinomas. In order to characterize patterns of global transcriptional deregulation in primary colon carcinomas, we performed gene expression profiling of some 300 tumors, including colon and rectum using oligonucleotide microarrays. For most of these tumors, expression profiles were compared to those from matched normal mucosa samples. We identified a set of 1,950 genes with highly significant deregulation between tumors and mucosa samples (P1e-7). A significant proportion of these genes mapped to chromosome 20 (P=0.01), which is frequently gained in colorectal tumors. Finally, we established a relationship between specific genomic imbalances, which were mapped for many 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/b-catenin signaling cascade, suggesting similar pathogenic pathways. Nuclear topography of aneuploid chromosomes and their consequences on transcriptional activity The 3D-position of chromosome territories in interphase nuclei is non-random and is conserved in evolution. Gene rich chromosomes, such as human chromosome 19, are located towards the center of the nucleus, whereas gene poor chromosomes, such as chromosome 18, are predominantly peripheral in many different cell types. We were eager to explore to which extent this remarkable conservation is maintained in cells containing chromosomal aneuploidies, in particular because we have previously established that these chromosomes are transcriptionally active. Therefore, a relationship could exist between nuclear position and transcriptional activity. We investigated, first, whether aneuploid chromosomes assume a nuclear position similar to that of their endogenous homologues. Using 3D-FISH and confocal laser scanning microscopy, we showed that human chromosomes 7, 18, or 19 introduced via microcell mediated chromosome transfer (MMCT) into the parental diploid colon cancer cell line DLD-1 maintain their conserved position. Our data is thus consistent with the model that each chromosome has an associated zip code that determines its nuclear localization. Whether the nuclear localization determines, or is determined by, the transcriptional activity of resident genes has yet to be ascertained. Similar analyses were extended to the murine donor cells used for MMCT which carried an individual human chromosome. Here we investigated (i) whether mouse nuclei recognize such a zip code of nuclear position on human chromosomes to facilitate their distinct partitioning and (ii) if chromosome positioning and transcriptional activity remain coupled under these trans-species conditions. We showed (i) that gene poor and gene rich human chromosomes do in fact maintain their divergent but conserved positions in mouse-human hybrid nuclei and (ii) that a foreign human chromosome is actively transcribed in mouse nuclei. The results suggest a species-independent conserved mechanism for the non-random positioning of chromosomes in the 3D interphase nucleus. Aneuploidy, oncogene amplification, and epithelial/mesenchymal transition govern transformation of murine epithelial cells: models for human cancer. We extended these analyses and designed a novel mouse model for colon cancer. Normal primary murine epithelial colon cells were selectively isolated from the large intestine of eight different isogenic C57BL/6 mice. A portion of the primary colon cells were grown in culture under reduced serum conditions while another fraction of these normal cells were cryopreserved for further analysis. The cells were sequentially recovered from culture based on distinct morphological changes: first, when the cells were in the pre-immortal stage as cells were actively growing;second, as the cells bypassed crisis, formed colonies and became immortal;third, at early transformation, as the cells first formed foci;fourth, mid-transformed (higher proliferation rates, later passages);and finally, as cells became less adherent, and exhibiting multiple foci. The cells were analyzed at each stage using molecular cytogenetic methods and microarray techniques. SKY identified recurrent structural and numerical aberrations in all eight of the late transformed mouse colon cell lines. Array CGH revealed regions of gene amplifications and deletions. Global gene expression profiling identified deregulated genes specific to each stage of transformation. Three of the late transformed mouse colon cell lines produced tumors in nude mouse assays. This is a unique murine model that reproduces a stage-specific progression of alterations which is common in human colon tumorigenesis. The similarity of the cytogenetic and genetic aberration profiles suggest that this model can serve as a preclinical system for the validation of therapeutic targets. As in human tumors, chromosomal aneuploidies resulted in the transcriptional deregulation of resident genes;in other words, genomic copy number is positively and linearly correlated with transcription levels.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC010835-07
Application #
8763244
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
7
Fiscal Year
2013
Total Cost
$999,975
Indirect Cost
Name
National Cancer Institute Division of Basic Sciences
Department
Type
DUNS #
City
State
Country
Zip Code
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