Comprehensive serial analysis of gene expression of the cervical transcriptome

<p>Abstract</p> <p>Background</p> <p>More than half of the approximately 500,000 women diagnosed with cervical cancer worldwide each year will die from this disease. Investigation of genes expressed in precancer lesions compared to those expressed in normal cervical epithelium will yield insight into the early stages of disease. As such, establishing a baseline from which to compare to, is critical in elucidating the abnormal biology of disease. In this study we examine the normal cervical tissue transcriptome and investigate the similarities and differences in relation to CIN III by Long-SAGE (L-SAGE).</p> <p>Results</p> <p>We have sequenced 691,390 tags from four L-SAGE libraries increasing the existing gene expression data on cervical tissue by 20 fold. One-hundred and eighteen unique tags were highly expressed in normal cervical tissue and 107 of them mapped to unique genes, most belong to the ribosomal, calcium-binding and keratinizing gene families. We assessed these genes for aberrant expression in CIN III and five genes showed altered expression. In addition, we have identified twelve unique HPV 16 SAGE tags in the CIN III libraries absent in the normal libraries.</p> <p>Conclusion</p> <p>Establishing a baseline of gene expression in normal cervical tissue is key for identifying changes in cancer. We demonstrate the utility of this baseline data by identifying genes with aberrant expression in CIN III when compared to normal tissue.</p

Up regulation in gene expression of chromatin remodelling factors in cervical intraepithelial neoplasia

<p>Abstract</p> <p>Background</p> <p>The highest rates of cervical cancer are found in developing countries. Frontline monitoring has reduced these rates in developed countries and present day screening programs primarily identify precancerous lesions termed cervical intraepithelial neoplasias (CIN). CIN lesions described as mild dysplasia (CIN I) are likely to spontaneously regress while CIN III lesions (severe dysplasia) are likely to progress if untreated. Thoughtful consideration of gene expression changes paralleling the progressive pre invasive neoplastic development will yield insight into the key casual events involved in cervical cancer development.</p> <p>Results</p> <p>In this study, we have identified gene expression changes across 16 cervical cases (CIN I, CIN II, CIN III and normal cervical epithelium) using the unbiased long serial analysis of gene expression (L-SAGE) method. The 16 L-SAGE libraries were sequenced to the level of 2,481,387 tags, creating the largest SAGE data collection for cervical tissue worldwide. We have identified 222 genes differentially expressed between normal cervical tissue and CIN III. Many of these genes influence biological functions characteristic of cancer, such as cell death, cell growth/proliferation and cellular movement. Evaluation of these genes through network interactions identified multiple candidates that influence regulation of cellular transcription through chromatin remodelling (<it>SMARCC1</it>, <it>NCOR1</it>, <it>MRFAP1 </it>and <it>MORF4L2</it>). Further, these expression events are focused at the critical junction in disease development of moderate dysplasia (CIN II) indicating a role for chromatin remodelling as part of cervical cancer development.</p> <p>Conclusion</p> <p>We have created a valuable publically available resource for the study of gene expression in precancerous cervical lesions. Our results indicate deregulation of the chromatin remodelling complex components and its influencing factors occur in the development of CIN lesions. The increase in SWI/SNF stabilizing molecule <it>SMARCC1 </it>and other novel genes has not been previously illustrated as events in the early stages of dysplasia development and thus not only provides novel candidate markers for screening but a biological function for targeting treatment.</p

Global approaches to identifying aberrations in early staged disease in cancers affecting women

INTRODUCTION: Breast and cervical cancer are the most common cancers in women worldwide. Widely implemented screening programs have allowed for the detection of precancerous lesions in the breast and cervix and have provided a valuable opportunity to study the earliest events in disease development with the goal of distinguishing cases that are likely to progress from those that are self limited or would spontaneously regress. OBJECTIVE: The overall objective of this thesis is to identify genes in biological pathways or networks altered in pre cancerous lesions of the breast and cervix using global analysis tools. HYPOTHESIS: I hypothesize that global transcriptome and high resolution genome analysis will identify altered genes that are shared in pre cancer lesions of both the cervix and breast. METHODS: A comprehensive Serial Analysis of Gene Expression method was used for the unbiased analysis of well characterized, frozen samples of normal cervix, CIN I, CIN II and CIN III. Tiling resolution whole genome array comparative hybridization was used for the detailed investigation of lobular carcinoma in situ (LCIS) and atypical lobular hyperplasia (ALH). The efficacy of this tool was first confirmed in commonly used breast cancer cell lines and then in archival breast cancer tissue. RESULTS: This work has led to the identification of aberrations in a chromatin remodelling gene network not previously implicated in cervical intraepithelial neoplasia. Genomic copy number analysis revealed novel features not previously described including aberrations to multiple components of a single biological pathway (epidermal growth factor receptor), the delineation of alteration boundaries and the identification of a novel amplicon of prognostic significance in breast cancer. We identified novel copy number changes in several genes in LCIS and ALH (including HOXB cluster genes) that were used to elucidate a genomic signature. Aberrations in HoxB7 were identified in pre cancer lesions of both breast and cervix (LCIS and CIN III) tissue. CONCLUSION: Collectively, this work demonstrates that through whole genome approach of assessment of genomic copy number and expression we can identify novel genes and gene networks that are altered during the development of pre cancer lesions.Medicine, Faculty ofPathology and Laboratory Medicine, Department ofGraduat

Epidermal growth factor receptor (EGFR) is transcriptionally induced by the Y-box binding protein-1 (YB-1) and can be inhibited with Iressa in basal-like breast cancer, providing a potential target for therapy

Introduction Basal-like breast cancers (BLBCs) are very aggressive, and present serious clinical challenges as there are currently no targeted therapies available. We determined the regulatory role of Y-box binding protein-1 (YB-1) on epidermal growth factor receptor (EGFR) overexpression in BLBC, and the therapeutic potential of inhibiting EGFR. We pursued this in light of our recent work showing that YB-1 induces the expression of EGFR, a new BLBC marker. Methods Primary tumour tissues were evaluated for YB1 protein expression by immunostaining tissue microarrays, while copy number changes were assessed by comparative genomic hybridization (CGH). The ability of YB-1 to regulate EGFR was evaluated using luciferase reporter, chromatin immunoprecipitation (ChIP) and gel shift assays. The impact of Iressa on monolayer cell growth was measured using an ArrayScan VTI high-throughput analyser to count cell number, and colony formation in soft agar was used to measure anchorage-independent growth. Results YB-1 (27/37 or 73% of cases, P = 3.899 × 10-⁴) and EGFR (20/37 or 57.1% of cases, P = 9.206 × 10-¹²) are expressed in most cases of BLBC. However, they are not typically amplified in primary BLBC, suggesting overexpression owing to transcriptional activation. In support of this, we demonstrate that YB-1 promotes EGFR reporter activity. YB-1 specifically binds the EGFR promoter at two different YB-1-responsive elements (YREs) located at -940 and -968 using ChIP and gel shift assays in a manner that is dependent on the phosphorylation of S102 on YB-1. Inhibiting EGFR with Iressa suppressed the growth of SUM149 cells by ~40% in monolayer, independent of mutations in the receptor. More importantly anchorage-independent growth of BLBC cell lines was inhibited with combinations of Iressa and YB-1 suppression. Conclusion We have identified for the first time a causal link for the expression of EGFR in BLBC through the induction by YB-1 where it binds specifically to two distinguished YREs. Finally, inhibition of EGFR in combination with suppression of YB-1 presents a potential opportunity for therapy in BLBC.Medicine, Faculty ofPediatrics, Department ofOther UBCNon UBCReviewedFacult