Identification of Genetic and Epigenetic Alterations in Gynecologic Cancers and Their Clinical Implications

This dissertation, "Identification of Genetic and Epigenetic Alterations in Gynecologic Cancers and Their Clinical Implications" by Huijuan, Yang, 楊慧娟, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Abstract of thesis entitled Identification of Genetic and Epigenetic Alterations in Gynecologic Cancers and Their Clinical Implications Submitted by Yang Hui Juan for the degree of Doctor of Philosophy at The University of Hong Kong in Aug, 2004 Cervical, endometrial and ovarian cancers are relatively common causes of morbidity and mortality in women worldwide. A series of genetic and epigenetic changes contribute to the initiation and progression of human cancers. This study attempted to identify somatic mutations of mitochondrial DNA (mtDNA), nuclear microsatellite instability (nMSI), human papillomavirus (HPV) infection, and CpG island hypermethylation, and their clinical implications, in the three cancers. Somatic mutations of the D-loop region of mtDNA were identified by direct sequencing in 48.9% of 45 endometrial cancers and 34.0% of 50 cervical cancers, and in 20.0% of 25 ovarian cancers reported previously. Mitochondrial microsatellite instability (mtMSI) was the commonest alteration with a frequency of 44.4% in endometrial cancer, higher than the other two cancers. The homopolymorphic C tracts located in the hypervariable segments 1 and 2 (HVS 1 & 2) were hotspots for instability. Abstract 1 nMSI for 7 microsatellite markers was investigated in 66 cervical cancers, 43 endometrial cancers and 44 ovarian cancers by PCR amplification. High-level MSI was identified in 20-30% of the three cancers. The occurrence of nMSI was not associated with mtMSI. HPV 16 DNA was identified by real-time quantitative PCR in 32.1% of 56 ovarian cancers and 15.2% of 46 endometrial cancers, significantly lower than in cervical cancers (64.3% of 129). HPV 18 DNA was detected in 20.9% of cervical cancers and one ovarian cancer (1.8%). The levels of HPV 16 DNA in cervical cancer were significantly higher than the other two cancers, and the incidence or levels of HPV DNA were associated with histological type, stage, grade, or response to radiotherapy. Plasma HPV DNA was identified in more than half of cervical cancers. Its level was associated with the viral load in the paired primary tumor, and changed with the treatment and progression of the disease. Using pooled DNA approach and methylation-specific PCR, the methylation status of the CpG islands of 34 genes was determined in the three cancers. Differential DNA methylation profiles were identified. Some gene changes were shared and others were specific. CpG island hypermethylation for 4 selected genes in each cancer type was identified in 50-70% of patients with the three cancers. The occurrence of DAPK hypermethylation was the highest, in 56.7% of cervical cancers, and independent on HPV infection. It was more common in squamous cell carcinoma than in adenocarcinoma and was an independent prognostic factor for shorter disease-free survival. Aberrant methylation for other genes (BRCA1, hMLH1, MGMT, p14, p16, and PTEN) was also associated with histological subtypes, stage, grade or treatment response. Furthermore, hypermethylated sequences were identified in 58.2% of Abstract 2 plasma samples of cervical cancers, significantly higher than in precancerous diseases and controls. In summary, molecular alterations are different in the three cancers. mtMSI at HVS 1 & 2 might be a potential molecular marker in endometrial cancer. Measurement of HPV DNA copy number in primary tumor and plasm