Nasopharyngeal cancer (NPC) is endemic in Southern China and Hong Kong. It has traditionally been treated by local radiotherapy with great success especially for early stage disease. However the treatment outcome in advanced stage disease is unsatisfactory. / Results from this series of combined clinical, translational and laboratory studies have redefined the role of hypoxia, angiogenesis and metastasis as new therapeutic targets in NPC. Novel biomarkers and new therapeutic approaches were developed based on these targets. / To develop new therapies in NPC, we demonstrated in a randomized controlled phase 2 clinical trial that sequential therapy of neoadjuvant chemotherapy followed by chemoradiotherapy was well tolerated with a manageable toxicity profile that allowed subsequent delivery of full dose chemoradiotherapy. This strategy reduced distant metastasis which translated into improved patient survival. In preclinical studies, the antiangiogenesis agent sunitinib demonstrated potent in vitro and in vivo growth inhibition in NPC. In a phase 2 clinical trial, sunitinib demonstrated modest clinical activity in heavily pretreated NPC patients. However, the unexpected high incidence of severe hemorrhage from upper aero-digestive tract in NPC patients who received prior high dose RT to the region is of concern. We propose to exclude NPC patients with disease recurrence within previous radiation field and/or with vascular invasion from future antiangiogenesis therapy. / To investigate potential new therapeutic targets and biomarkers in NPC, we first confirmed from the Hong Kong NPC study group of 2915 patients' database that distant metastasis was the leading cause of NPC failure after primary radiotherapy. We further showed that hypoxia induced broad changes of both up- and down-regulated gene expressions involved in diverse biological processes in NPC cells. Over-expression of biomarkers of hypoxia and angiogenesis (including HIF-1alpha, CA IX and VEGF) is common in NPC and is associated with poor prognosis. Elevated plasma osteopontin is a biomarker of distant metastasis, and pre-treatment plasma osteopontin level may be a useful biomarker of response to radiotherapy in NPC. / Hui, Pun. / "September 2010." / Adviser: Anthony Chan. / Source: Dissertation Abstracts International, Volume: 73-04, Section: B, page: . / Thesis (M.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 269-293). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.

Genes identified by SAGE may serve as potential prognostic marker or therapeutic target. 14-3-3 sigma is a putative tumor suppressor and can be induced in response to DNA damage following irradiation, leading to cell cycle arrest in G2/M in human cancer cells. Our SAGE results revealed that 14-3-3 sigma expression is significantly downregulated in C666-1 cells. The study of 72 primary NPCs showed that an increased expression of 14-3-3 sigma was associated with a poorer clinical outcome in terms of shorter overall survival (OS; p=0.0297) and shorter disease free survival (DFS; p=0.042) using univariate analysis. Hence, 14-3-3 sigma may be used as an independent prognostic marker for NPC patients. / In conclusion, a NPC transcription profile has been successfully generated and several candidate NPC-associated genes have been identified by Serial Analysis of Gene Expression (SAGE) and NPC transcriptome map. These novel findings lead to better understanding of the molecular basis of NPC development and provide a foundation for discovery of new therapeutic strategies. / Nasopharyngeal carcinoma (NPC) is one of the most prevalent cancers among Southern Chinese. To better understand the genetic basis of this disease, Serial Analysis of Gene Expression (SAGE) was performed to investigate the transcriptional profiles of an EBV-positive NPC cell line (C666-1) and a normal NP outgrowth (NP4). A total of 102,059 SAGE tags were extracted in both libraries and 250 genes with 10-fold or more differential expression were found in NPC cells compared to normal NP cells. Eleven differentially expressed genes identified by SAGE were selected for confirmation using real time RT-PCR. The transcripts for 5 of the 11 genes, CD 74, Transcriptional intermediary factor 1, Ferritin 1, Claudin 4, and fatty acid synthase were overexpressed in NPC cells. Conversely, the remaining transcripts including Keratin 17, Keratin 5, S100 calcium-binding A2, Cystatin A, 14-3-3 sigma and Caveolin 1 were underexpressed in NPC cells. The aberrant expression of CD74, Claudin 4, Fatty acid synthase, 14-3-3 sigma, Caveolin 1 were further validated by immunohistochemistry on 20 NPC patients. / On the other hand, fatty acid synthase (FASN), a key enzyme for de novo lipogenesis, is a putative therapeutic target in treating NPC. Immunohistochemical studies showed upregulation of FASN in 20.8% (15/72) of the NPC cases compared with the adjacent normal NP epithelium. In addition, FASN expression also had prognostic significance in predicting the outcome of patients after radiotherapy, as high levels of FASN expression were associated with worse overall survival (OS, p=0.032) and disease free survival (DFS, p=0.002) in NPC patients. FASN inhibitors, such as C75 which inhibit cell growth via cell cycle arrest in G2/M phase, are potential chemotherapeutic agents in treating NPC. / The genome-wide quantitative analysis of gene expression by SAGE with matched chromosomal positions enables the construction of a transcriptome map of NPC. A total of 8 and 29 overexpressed and underexpressed gene clusters were observed, respectively. Some novel regions that have never been illustrated in previous reports such as amplification regions at 2p11.2-p25.1, 2q33-q37, 9q22-q34, 17p11.2-p13.2 and deletion regions at 1p12-p31.2, 1q25-q42.12, 2q21.3-q33, 8p21.1-p22, 9q33-q34.3, 10q23.3-q26.3, 12p13, 16p13, 17q23.2-q25, 19p13, 19q12-q13.2, 20p11-p13, 22q13, Xp11.2-p11.4, and Xq26-q28 were also identified. A candidate tumor suppressor gene named MEG3 has been found within an underexpressed region at 14q32.2 in the NPC transcriptome map. Our FISH analysis revealed that chromosome loss at 14q32 is associated with hypermethylation of MEG3 promoter region in 9/13 (75%) of NPC patients. Loss of imprinting is the major mechanism that governs the MEG3 expression. Moreover, transient transfection of one of the MEG3 isoforms (accession no. AF119863) could obviously inhibit cell colony formation of NPC cells. Taken together, MEG3 gene on chromosome 14q32.2 might act as a tumor suppressor in NPC. / Chan, Yat Yee. / "March 2008." / Adviser: Lo Kwok Wai. / Source: Dissertation Abstracts International, Volume: 70-03, Section: B, page: 1605. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (p. 196-225). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Wong Yue Hang Albert. / Thesis submitted in: December 2005. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 108-119). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.v / List of Figures --- p.x / List of Tables --- p.xi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter Chapter 2 --- Literature Review --- p.4 / Chapter 2.1 --- Nasopharyngeal Carcinoma (NPC) --- p.4 / Chapter 2.1.1 --- Anatomy of Nasopharynx --- p.4 / Chapter 2.1.2 --- Histopathology of Nasopharyngeal Carcinoma --- p.5 / Chapter 2.1.3 --- Epidemiology and Etiology of Nasopharyngeal Carcinoma --- p.5 / Chapter 2.1.3.1 --- Endemic Regions of Nasopharyngeal Carcinoma --- p.5 / Chapter 2.1.3.2 --- Gender and Age Bias --- p.6 / Chapter 2.1.3.3 --- Nasopharyngeal Carcinoma in Hong Kong --- p.6 / Chapter 2.1.3.4 --- Environmental Factors and Diet --- p.7 / Chapter 2.1.3.5 --- HLA Haplotypes and Nasopharyngeal Carcinoma --- p.9 / Chapter 2.1.4 --- Epstein-Barr Virus (EBV) and Nasopharyngeal Carcinoma --- p.10 / Chapter 2.1.4.1 --- EBV and Human Cacners --- p.10 / Chapter 2.1.4.2 --- EBV Infection --- p.10 / Chapter 2.1.4.3 --- "Latent, Clonal EBV Infection" --- p.11 / Chapter 2.1.4.4 --- EBV Latency Form --- p.11 / Chapter 2.1.4.5 --- Reactivation of EBV --- p.12 / Chapter 2.1.5 --- Molecular Pathogenesis of Nasopharyngeal Carcinoma --- p.13 / Chapter 2.1.5.1 --- Genetic Changes --- p.13 / Chapter 2.1.5.2 --- Epigenetic Changes --- p.13 / Chapter 2.1.6 --- Therapy of Nasopharyngeal Carcinoma and its Deficiency --- p.14 / Chapter 2.1.6.1 --- Radiotherapy --- p.14 / Chapter 2.1.6.2 --- Concurrent Chemoradiotherapy --- p.16 / Chapter 2.1.6.3 --- Adjuvant and Neo-adjuvant Chemotherapy --- p.17 / Chapter 2.1.6.4 --- Chemotherapy in Metastatic Nasopharyngeal Carcinoma --- p.18 / Chapter 2.1.6.5 --- Novel Therapeutic Agents and Approach --- p.19 / Chapter 2.2 --- Histone Modification and Cancer --- p.20 / Chapter 2.2.1 --- Histone Modification and Transcription Regulation --- p.20 / Chapter 2.2.2 --- Carcinogenic Effect of Aberrant HAT and HDAC Activities --- p.21 / Chapter 2.2.3 --- Structural Classes of HDAC Inhibitors --- p.24 / Chapter 2.2.4 --- Anti-Cancer Mechanisms of HDAC Inhibitors --- p.25 / Chapter 2.3 --- Suberoylanilide Hydroxamic Acid (SAHA) --- p.27 / Chapter 2.3.1 --- Anti-tumor Effect of SAHA in Various Cancer Cell Lines --- p.27 / Chapter 2.3.2 --- SAHA Mediated Non-apoptotic Programmed Cell Death --- p.29 / Chapter 2.3.3 --- Anti-tumor and Preventive Effect of SAHA in Animal Model --- p.29 / Chapter 2.3.4 --- Clinical Trials of SAHA --- p.30 / Chapter 2.4 --- FK228 (Depsipeptide or FR901228) --- p.31 / Chapter 2.4.1 --- Anti-malignancy mechanism of FK228 --- p.31 / Chapter 2.4.2 --- Anti-angiogenesis --- p.32 / Chapter 2.4.3 --- Drug Resistance and FK228 --- p.33 / Chapter 2.4.4 --- Studies of FK228 on Animal Models --- p.33 / Chapter 2.4.5 --- Clinical Trials --- p.34 / Chapter 2.5 --- Histone Modification and Nasopharyngeal Carcinoma --- p.34 / Chapter Chapter 3 --- Materials and Methods --- p.36 / Chapter 3.1 --- Cell Lines --- p.36 / Chapter 3.2 --- EBER ish Hybridization (EBER ISH) --- p.37 / Chapter 3.3 --- HDAC Inhibitors --- p.38 / Chapter 3.4 --- Cellular Sensitivity of NPC Cell Lines to HDAC Inhibitors --- p.38 / Chapter 3.4.1 --- Drug Treatment --- p.38 / Chapter 3.4.2 --- Determining Relative Amount of Survival Cells (WST-1 Assay) --- p.39 / Chapter 3.5 --- Flow Cytometry Analysis --- p.40 / Chapter 3.5.1 --- Collecting Cells and Fixation --- p.40 / Chapter 3.5.2 --- Staining --- p.41 / Chapter 3.5.3 --- Flow Cytometry Analysis --- p.41 / Chapter 3.6 --- Protein Extraction --- p.41 / Chapter 3.6.1 --- Harvesting Samples --- p.41 / Chapter 3.6.2 --- Protein Extraction --- p.42 / Chapter 3.6.3 --- Protein Quantification --- p.42 / Chapter 3.7 --- Western Blotting --- p.43 / Chapter 3.7.1 --- SDS-Polyarcylamide Gel Electrophoresis (PAGE) (SDS-PAGE) --- p.43 / Chapter 3.7.2 --- Wet Transfer of Proteins --- p.43 / Chapter 3.7.3 --- Immunoblotting --- p.44 / Chapter 3.7.4 --- Signal Detection --- p.44 / Chapter 3.8 --- CodeLin´kёØ Oligonucleotide Microarray --- p.45 / Chapter 3.8.1 --- HDAC Inhibitor Treatment --- p.45 / Chapter 3.8.2 --- RNA Extraction --- p.45 / Chapter 3.8.3 --- Quality and Quantity Assessment of Total RNA Extracted --- p.46 / Chapter 3.8.4 --- CodeLinkIM Expression Bioarray System --- p.46 / Chapter 3.8.5 --- Data Analysis --- p.48 / Chapter 3.9 --- Real-time Reverse Transcription PCR (Real-time RT-PCR) --- p.48 / Chapter Chapter 4 --- Results --- p.50 / Chapter 4.1 --- Presence of EBV --- p.50 / Chapter 4.2 --- Anti-prolirative Effect of HDAC Inhibitors --- p.52 / Chapter 4.3 --- Histone Acetylation --- p.56 / Chapter 4.4 --- Induction of p21 Expression in NPC Cell Lines --- p.58 / Chapter 4.5 --- HDAC Inhibitors Induced Cell Cycle Arrest and Polyploidy Formation --- p.60 / Chapter 4.5.1 --- Trichostatin A Induced G2/M Arrest --- p.60 / Chapter 4.5.2 --- Suberoylanilide Hydroxamic Acid Induced G1 Arrest --- p.62 / Chapter 4.5.3 --- FK228 Mediated G2/M Arrest --- p.64 / Chapter 4.6 --- HDAC Inhibitors Altered the Expression of Cell Cycle Regulatory Proteins --- p.66 / Chapter 4.6.1 --- TSA Down-regulated Cyclin A and B --- p.66 / Chapter 4.6.2 --- Suppressed Expression of Cyclin D1 and B by SAHA --- p.69 / Chapter 4.6.3 --- Effect of FK228 on Expression of Different Cyclins in NPC Cell Lines --- p.71 / Chapter 4.7 --- Effect of HDAC Inhibitors on EBV Proteins --- p.73 / Chapter 4.8 --- HDAC Inhibitors Modulated Gene Expression Profile --- p.76 / Chapter 4.8.1 --- SAHA and FK228-Induced Gene Expression Profile --- p.76 / Chapter 4.8.2 --- Validation of Expression Profile of Selected Genes by Real-time RT-PCR --- p.83 / Chapter Chapter 5 --- Discussion --- p.87 / Chapter 5.1 --- Anti-proliferative Effect of SAHA and FK228 on NPC Cell Lines --- p.88 / Chapter 5.2 --- Resistance of SAHA or FK228 in NPC --- p.93 / Chapter 5.3 --- Growth Inhibitory Mechanism of SAHA and FK228 in NPC Cells --- p.94 / Chapter 5.4 --- Induction of Polyploidy Cells in NPC Cell Lines --- p.98 / Chapter 5.5 --- Does EBV play a Role in HDAC Inhibiotrs Induced Growth Arrest in NPC Cell Lines? --- p.99 / Chapter 5.6 --- Transcriptional Signature of SAHA and FK228 in NPC Cell Lines --- p.100 / Chapter 5.7 --- Combining SAHA or FK228 with other Anti-tumor Agents --- p.104 / Chapter 5.8 --- Future Prospectus --- p.105 / Chapter Chapter 6 --- Summary --- p.106 / References --- p.108 / Appendix 1 --- p.120 / Appendix 2 --- p.121

鼻咽癌(nasopharyngeal carcinoma, NPC)與艾巴氏病毒(Epstein-Barr virus, EBV)和遺傳及表現遺傳變異有連串關係。儘管鼻咽癌腫瘤的發生機制仍然未知,最近研究顯示微核糖核酸(microRNA, miRNA)通過調節細胞增殖凋亡遷移和侵襲等方式對鼻咽癌的生成起著至關重要的作用。為了確定微核糖核酸與鼻咽癌發生的相關機制及其扮演的角色，我們集中研究微核糖核酸在鼻咽癌腫瘤中所發生的變異，探討這些異常表達的微核糖核酸的功能，並揭開與幹細胞相關的微核糖核酸在鼻咽癌幹細胞樣細胞(cancer stem-like cell, CSC)裏所扮演的角色。 / 通過使用微陣列技術(Agilent Microarray)， 我們運用了 866個人類與 89個病毒微核糖核酸探針,以識別出多個帶有艾巴氏病毒的鼻咽癌腫瘤細胞系裏的微核糖核酸表達圖譜。相比正常的鼻咽上皮細胞系NP69，113個微核糖核酸在鼻咽癌中的差異表達已被鑒定出來。其中58個在鼻咽癌裏下調的微核糖核酸表達，miR-31的轉錄下調現象在鼻咽癌腫瘤細胞系和原發腫瘤中被不斷地發現。在7個帶有艾巴氏病毒的腫瘤細胞系樣本裏, 其中6個(86%)樣本呈miR-31下調跡象。與此同時，以顯微切割技術所得的38個原發腫瘤樣本中全部(100%)都顯示有miR-31下調的跡象。相比之下，所有正常的鼻咽上皮細胞都顯出高表達的miR-31。 / miR-31位於染色體9p21.3上，距離CDKN2A (p16) 0.5Mb處。這是在鼻咽癌細胞裏通常缺失的位置。在X1915和X99186腫瘤細胞系中，已證實在miR-31和CDKN2A位點上都出現了純合性缺失。在四株不具備miR-31缺失的腫瘤細胞系裏，甲基化特異性聚合酶連鎖反應 (methylation-specific PCR, MSP) 和亞硫酸氫鈉測序法(bisulfite sequencing)發現了高甲基化的CpG島。使用5-aza-2’-deoxycytidine (5-Aza-dC) 治療後，鼻咽癌細胞株C666-1被證實恢復了miR-31轉錄。這些結果表明，純合性缺失和啟動子高甲基化是造成miR-31在鼻咽癌裏轉錄失效的主要發生機制。 / 微陣列技術和生物信息學分析找出了一些可能受miR-31影響的基因。其中FIH1和MCM2被確定為在鼻咽癌細胞裏受miR-31影響的基因。我們證實miR-31與FIH1和MCM2 信使核糖核酸的3’UTR處結合會抑制螢光素酶的活性。在鼻咽癌細胞裏miR-31的異位表達也會壓抑FIH1和MCM2蛋白的表達。更重要的是，恢復正常的miR-31表達或敲除FIH1表達能顯著地抑制C666-1細胞的增殖和移動能力。C666-1細胞的克隆形成能力和錨定依賴性生長都顯著地被miR-31的表達所抑制。穩定的miR-31表達亦能抑制鼻咽癌腫瘤在裸鼠體內的生長。此外，FIH1的敲除加強了p21和磷酸化p53 (Ser15) 的表達。這些結果暗示了miR-31是一個與鼻咽癌至關重要的微核糖核酸。 它通過了對FIH1的壓制，負面地調節細胞的增殖和移動。 / 使用微核糖核酸微陣列分析後，我們在鼻咽癌細胞中培養的懸浮細胞球裏篩選出差異表達的微核糖核酸。同樣地，實時螢光定量逆轉錄聚合酶鏈反應(qRT-PCR) 亦證實了miR-96和miR-183在C666-1懸浮細胞球裏是被抑制的。此外，miR-96和miR-183的異位表達顯著地降低了C666-1懸浮細胞球形成和克隆形成的能力。這項研究結果暗示, miR-96和miR-183的抑制對鼻咽癌幹細胞樣細胞的形成非常重要。 / 總的來說，某些微核糖核酸已被確定為潛在的鼻咽癌腫瘤抑制基因。 在帶艾巴氏病毒的鼻咽癌裏，miR-31的表達被證實是因純合性缺失和啟動子高甲基化而被下調的。miR-31抑制鼻咽癌細胞的增殖錨定依賴性生長細胞遷移和體內腫瘤的生長。同時，miR-96和miR-183也被發現對維持鼻咽癌的幹細胞樣特性起著一定作用。這些結果表明微核糖核酸對鼻咽癌腫瘤的生成扮演著抑制的角色。對微核糖核酸的機制作進一步全面了解將改進鼻咽癌的治療策略。 / Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) has been reported to be related to a number of genetic and epigenetic changes, however, the molecular mechanism leading to NPC tumorigenesis still remains unclear. Recently, microRNAs (miRNAs) have been demonstrated to play vital roles in NPC development via regulating cell proliferation, apoptosis, and cell migration and invasion. In this study, we aim to elucidate the role of miRNAs in NPC tumorigenesis in this study by identifying the miRNA aberration, investigating the possible functions of these aberrantly expressed miRNAs, and unraveling the role of stemness-related miRNAs in NPC cancer stem-like cells (CSCs). / By using Agilent Microarray with 866 human and 89 viral miRNA probes, miRNA expression profiles of multiple EBV-associated NPC tumor lines were generated. Compared to NP69, a nonmalignant nasopharyngeal epithelial cell line, 113 differentially expressed miRNAs were identified. Among the 58 down-regulated miRNAs in NPC, transcriptional silencing of miR-31 was consistently found in both NPC tumor lines and primary tumors. Down-regulation of miR-31 was detected in 6 of 7 (86%) EBV-positive tumor lines and 38 of 38 (100%) microdissected primary tumors, while all normal nasopharyngeal epithelia showed high expression of miR-31. / miR-31 is located at 0.5 Mb telomeric to CDKN2A (p16) on chromosome 9p21.3, which is commonly deleted in NPC. Homozygous deletion of both miR-31 and CDKN2A loci was confirmed in tumor lines X1915 and X99186. In the four tumor lines with intact miR-31, hypermethylation of 5’ CpG islands was detected by methylation-specific PCR (MSP) and bisulfite sequencing analysis. Restoration of miR-31 transcription was demonstrated in the EBV-positive NPC cell line C666-1 treated with 5-aza-2’-deoxycytidine. These findings suggested that homozygous deletion and promoter hypermethylation are the major mechanisms for transcriptional silencing of miR-31 in NPC. / By microarray and bioinformatic analysis, a number of putative targets of miR-31 were identified. Among these candidates, FIH1 and MCM2 were found to be the targets of miR-31 in NPC. We have shown that binding of miR-31 on FIH1 and MCM2 mRNA 3’UTR suppressed their luciferase activity. Ectopic expression of miR-31 in NPC cells resulted in repression of FIH1 and MCM2 protein expression. Importantly, the restoration of miR-31 or knockdown of FIH1 expression significantly suppressed proliferation as well as migration of C666-1 cells. Clone-forming ability and anchorage-independent growth of C666-1 were significantly inhibited by miR-31 expression. Stably expressed miR-31 was also demonstrated to inhibit NPC tumor growth in nude mice. Furthermore, expression of p21 and phospho-p53 (Ser15) was found to be increased by FIH1 knockdown. These results implied that miR-31 is a critical NPC-associated miRNA which negatively regulates cell proliferation and migration via FIH1 repression. / By miRNA microarray analysis, we have screened for differentially expressed miRNAs in sphere-forming cells of EBV-associated NPC. In concordance with microarray findings, suppression of miR-96 and miR-183 in C666-1 spheroids was confirmed by qRT-PCR. Ectopic expression of miR-96 and miR-183 significantly reduced the sphere-forming and clone-forming ability of C666-1 cells. The findings implied that miR-96 and miR-183 repression is important in the formation of NPC CSCs. / In summary, several miRNAs were identified as potential tumor suppressor genes in NPC. miR-31 was found down-regulated by homozygous deletion or promoter hypermethylation in EBV-associated NPC. It plays roles in NPC pathogenesis by suppressing NPC cell proliferation, clone-forming ability, cell anchorage-independent growth, migration and in vivo tumor growth. Moreover, miR-96 and miR-183 were found to have a role in the maintenance of NPC stem-like properties. These findings suggested important tumor suppressive roles of miRNAs in regulating NPC tumorigenesis, and a better understanding on the miRNA mechanisms may potentiate better therapeutic strategies for NPC. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Cheung, Ching Mei. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 177-209). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts also in Chinese. / Abstract --- p.i / 摘要 --- p.iv / Thesis / Assessment Committee --- p.vii / Acknowledgements --- p.viii / Table of contents --- p.ix / List of figures --- p.xv / List of tables --- p.xviii / List of publications --- p.xix / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Nasopharyngeal carcinoma (NPC) --- p.1 / Chapter 1.1.1 --- Histopathology and epidemiology --- p.1 / Chapter 1.1.2 --- Etiology --- p.2 / Chapter 1.1.2.1 --- Environmental factors --- p.2 / Chapter 1.1.2.2 --- Genetic factors --- p.3 / Chapter 1.1.2.3 --- Epstein-Barr virus (EBV) infection --- p.3 / Chapter 1.2 --- Molecular pathogenesis of NPC --- p.5 / Chapter 1.2.1 --- Cytogenetic changes --- p.5 / Chapter 1.2.2 --- NPC-associated tumor suppressor genes (TSGs) --- p.6 / Chapter 1.2.3 --- NPC-associated oncogenes --- p.8 / Chapter 1.3 --- MicroRNAs --- p.10 / Chapter 1.3.1 --- Biogenesis of microRNAs --- p.10 / Chapter 1.3.2 --- MicroRNAs and cancers --- p.15 / Chapter 1.3.2.1 --- MicroRNAs - tumor suppressors --- p.15 / Chapter 1.3.2.2 --- MicroRNAs - oncogenes --- p.16 / Chapter 1.4 --- MicroRNAs in nasopharyngeal carcinoma --- p.18 / Chapter 1.4.1 --- MicroRNA profiling in NPC --- p.18 / Chapter 1.4.2 --- OncomiRs in NPC --- p.20 / Chapter 1.4.3 --- Tumor suppressor miRNAs in NPC --- p.22 / Chapter 1.4.4 --- miRNAs and cancer stem-like cells (CSCs) --- p.27 / Chapter 1.4.5 --- Clinical implication of miRNAs in NPC --- p.29 / Chapter 1.5 --- Aims of study --- p.32 / Chapter Chapter 2 --- Materials and methods --- p.34 / Chapter 2.1 --- Patient biopsies --- p.34 / Chapter 2.2 --- NPC cell lines and xenografts --- p.34 / Chapter 2.2.1 --- Cell lines --- p.34 / Chapter 2.2.2 --- Xenografts --- p.36 / Chapter 2.3 --- Total RNA Isolation --- p.39 / Chapter 2.4 --- DNA extraction --- p.39 / Chapter 2.5 --- Protein Extraction --- p.40 / Chapter 2.6 --- Western Blotting --- p.40 / Chapter 2.7 --- Microarray analysis --- p.43 / Chapter 2.7.1 --- MicroRNA microarray --- p.43 / Chapter 2.7.2 --- Gene expression microarray --- p.44 / Chapter 2.8 --- Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR) --- p.45 / Chapter 2.8.1 --- Conventional qRT-PCR --- p.45 / Chapter 2.8.2 --- Stem-looped qRT-PCR --- p.46 / Chapter 2.9 --- Preparation of stable clone of miR-31 --- p.51 / Chapter 2.9.1 --- Cloning and plasmid DNA preparation --- p.51 / Chapter 2.9.1.1 --- Bacterial Transformation --- p.51 / Chapter 2.9.1.2 --- Plasmid DNA Extraction --- p.51 / Chapter 2.9.2 --- DNA Sequencing --- p.52 / Chapter 2.9.3 --- Stable transfection --- p.52 / Chapter 2.9.4 --- Clone selection --- p.53 / Chapter 2.10 --- Transient transfection --- p.55 / Chapter 2.11 --- Flow cytometry --- p.55 / Chapter 2.11.1 --- Apoptosis analysis by Annexin V --- p.55 / Chapter 2.11.2 --- Cell cycle analysis by propidium iodide (PI) --- p.56 / Chapter 2.11.3 --- Detection of stem-like cell markers --- p.56 / Chapter 2.12 --- Cell proliferation analysis --- p.56 / Chapter 2.12.1 --- WST-1 assay --- p.56 / Chapter 2.12.2 --- BrdU assay --- p.57 / Chapter 2.13 --- Anchorage-independent growth assay --- p.58 / Chapter 2.14 --- Clone formation assay --- p.58 / Chapter 2.15 --- Cell migration assay --- p.54 / Chapter 2.16 --- In vivo tumorigenicity --- p.59 / Chapter 2.17 --- Dual luciferase reporter assay --- p.60 / Chapter 2.17.1 --- Luciferase reporter vectors --- p.60 / Chapter 2.17.2 --- Luciferase reporter assay --- p.60 / Chapter 2.18 --- Mapping homozygous deletion and genes in chromosome 9p21.3 --- p.64 / Chapter 2.19 --- 5-aza-2’-deoxycytidine (5-Aza-dC) and Trichostatin A (TSA) treatments --- p.64 / Chapter 2.20 --- Methylation specific-PCR (MSP) and bisulfite sequencing analysis --- p.68 / Chapter 2.20.1 --- Bisulfite modification --- p.68 / Chapter 2.20.2 --- Methylation specific-PCR (MSP) --- p.69 / Chapter 2.20.3 --- Bisulfite sequencing analysis --- p.69 / Chapter 2.21 --- Statistical analysis --- p.70 / Chapter 2.22 --- In situ hybridization (ISH) analysis --- p.73 / Chapter Chapter 3 --- Identification of novel deregulated microRNAs in nasopharyngeal carcinoma --- p.74 / Chapter 3.1 --- Introduction --- p.74 / Chapter 3.2 --- Results --- p.80 / Chapter 3.2.1 --- Aberrant expression of microRNAs in NPC --- p.80 / Chapter 3.2.2 --- Homozygous deletion of miR-31 in NPC --- p.90 / Chapter 3.2.3 --- Hypermethylation of 5’ CpG islands of miR-31 in NPC --- p.92 / Chapter 3.2.4 --- Detection of miR-31 expression in normal epithelia and NPC by in situ hybridization --- p.99 / Chapter 3.3 --- Discussion --- p.101 / Chapter Chapter 4 --- Characteristics of miR-31 and its targets in NPC --- p.105 / Chapter 4.1 --- Introduction --- p.105 / Chapter 4.2 --- Results --- p.107 / Chapter 4.2.1 --- Effects of exogenous miR-31 on NPC cells --- p.107 / Chapter 4.2.1.1 --- miR-31 effect on C666-1 cell proliferation and cell cycle progression --- p.107 / Chapter 4.2.1.2 --- Clone-forming ability and anchorage-independent growth of C666-1 --- p.113 / Chapter 4.2.1.3 --- Migration ability of C666-1 --- p.113 / Chapter 4.2.2 --- Effects of stably expressed miR-31 on NPC cells --- p.117 / Chapter 4.2.2.1 --- Stable clones selection by restoring precursor of miR-31 into C666-1 --- p.117 / Chapter 4.2.2.2 --- Cell proliferation and cell cycle progression in stable clones of miR-31 --- p.117 / Chapter 4.2.2.3 --- Anchorage-independent growth of C666-1 stable clones --- p.117 / Chapter 4.2.2.4 --- Tumorigenicity of C666-1 stable clones expressing miR-31 in vivo --- p.118 / Chapter 4.2.3 --- Identification of miR-31 targets in NPC cells --- p.125 / Chapter 4.2.3.1 --- miR-31 targets FIH1 and MCM2 --- p.125 / Chapter 4.2.3.2 --- Other reported targets of miR-31 in NPC --- p.131 / Chapter 4.2.4 --- Functional analysis of FIH1 in NPC cells --- p.133 / Chapter 4.2.4.1 --- Repression of FIH1 by siRNAs --- p.133 / Chapter 4.2.4.2 --- Proliferation of C666-1 with FIH1 knockdown --- p.133 / Chapter 4.2.4.3 --- Clone-forming and migration ability of C666-1 transfected with siFIH1 --- p.133 / Chapter 4.2.4.4 --- Putative downstream targets of FIH1 --- p.139 / Chapter 4.2.5 --- Identification of novel miR-31 targets by gene expression microarray --- p.139 / Chapter 4.3 --- Discussion --- p.145 / Chapter Chapter 5 --- MicroRNAs regulation on NPC stem-like properties --- p.154 / Chapter 5.1 --- Introduction --- p.154 / Chapter 5.2 --- Results --- p.156 / Chapter 5.2.1 --- MicroRNA expression profiles in NPC sphere-forming cells --- p.155 / Chapter 5.2.2 --- Ectopic expression of miR-183 family and miR-203 in NPC --- p.161 / Chapter 5.2.2.1 --- Sphere-forming ability of NPC cells --- p.161 / Chapter 5.2.2.2 --- Clone-forming ability of C666-1 --- p.161 / Chapter 5.2.3 --- Sphere-forming ability of NPC cells transfected with anti-miR-96 and anti-miR-183 --- p.164 / Chapter 5.2.4 --- Expression of cacner stem cell markers in NPC cells transfected with miR-96 and miR-183 --- p.164 / Chapter 5.3 --- Discussion --- p.167 / Chapter Chapter 6 --- General discussion --- p.170 / Reference --- p.177

Ng Kenway. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 169-189). / Abstracts in English and Chinese ; questionnaire also in Chinese. / ABSTRACT --- p.I / 摘要 --- p.IV / ACKNOWLEDGEMENT --- p.V / TABLE OF CONTENTS --- p.VII / ABBREVIATION --- p.XI / LIST OF TABLES --- p.XIII / LIST OF FIGURES --- p.XIV / Chapter CHAPTER 1 --- INTRODUCTION --- p.1 / Chapter CHAPTER 2 --- LITERATURE REVIEW --- p.4 / Chapter 2.1 --- SIDE EFFECTS OF RADIATION THERAPY IN HEAD & NECK CACNER PATIENTS --- p.6 / Chapter 2.2 --- NUTRITIONAL ALTERATIONS IN CANCER PATIENTS --- p.9 / Chapter 2.3 --- FACTORS INFLUENCING ALTERATION IN CALORIE INTAKE IN CANCER PATIENTS --- p.12 / Chapter 2.3.1 --- Evidence for impaired calorie intake in cancer patients --- p.12 / Chapter 2.3.2 --- Anorexia --- p.13 / Chapter 2.3.2.1 --- Mucositis of upper food passage --- p.13 / Chapter 2.3.2.2 --- "Change in saliva and taste, food aversions" --- p.14 / Chapter 2.3.2.3 --- Psychological and emotional factors --- p.14 / Chapter 2.3.2.4 --- Cytokines --- p.15 / Chapter 2.4 --- FACTORS INFLUENCING ENERGY EXPENDITURE IN CANCER PATIETNS --- p.17 / Chapter 2.4.1 --- Introduction --- p.17 / Chapter 2.4.2 --- Components of total energy expenditure --- p.22 / Chapter 2.4.2.1 --- Measurement of Basal metabolic rate --- p.22 / Chapter 2.4.2.2 --- Energy cost of physical activity --- p.25 / Chapter 2.4.2.3 --- Thermic effect of food (TEF) --- p.26 / Chapter 2.5 --- METHODS FOR NUTRITIONAL ASSESSMENT --- p.27 / Chapter 2.5.1 --- Body weight and body composition --- p.27 / Chapter 2.5.2 --- Dietary intake --- p.30 / Chapter 2.6 --- METHODS FOR ENERGY EXPENDITURE MEASUREMENT --- p.35 / Chapter 2.7 --- CYTOKINES AND LEPTIN CHANGES IN CANCER PATIENTS --- p.40 / Chapter 2.7.1 --- Cytokines --- p.40 / Chapter 2.7.1.1 --- Tumor necrosis factor --- p.40 / Chapter 2.7.1.2 --- Interleukin 1 and interleukin 6 --- p.44 / Chapter 2.7.2 --- Leptin --- p.45 / Chapter 2.8 --- THE IMPACT OF MALNUTRITION ON CANCER SURVIVAL --- p.49 / Chapter CHAPTER 3 --- OBJECTIVES OF STUDY --- p.53 / Chapter CHAPTER 4 --- METHODS --- p.55 / Chapter 4.1 --- RETROSPECTIVE STUDY --- p.56 / Chapter 4.1.1 --- Patients --- p.56 / Chapter 4.1.2 --- Cancer staging --- p.56 / Chapter 4.1.3 --- Cancer treatment --- p.57 / Chapter 4.1.4 --- Outcome endpoints --- p.57 / Chapter 4.1.5 --- Determinants --- p.58 / Chapter 4.1.6 --- Statistical analysis --- p.58 / Chapter 4.2 --- PROSPECTIVE STUDY --- p.59 / Chapter 4.2.1 --- "Patients, oncological treatment, and assessment time points" --- p.59 / Chapter 4.2.2 --- Assessment of nutritional intake by food record --- p.60 / Chapter 4.2.3 --- Assessment of radiotherapy-induced symptoms --- p.61 / Chapter 4.2.4 --- Assessment of Basal metabolic rate --- p.62 / Chapter 4.2.5 --- Assessment of total energy expenditure and energy balance --- p.63 / Chapter 4.2.6 --- Assessment of body composition --- p.65 / Chapter 4.2.7 --- Measurement of cytokines --- p.68 / Chapter 4.2.7.1 --- Serum TNF-α --- p.69 / Chapter 4.2.7.2 --- Serum Human Leptin --- p.72 / Chapter CHAPTER 5 --- RESULTS --- p.80 / Chapter 5.1 --- RETROSPECTIVE STUDY --- p.81 / Chapter 5.1.1 --- The 5-year profile of bodyweight change during and after the end of radiotherapy --- p.81 / Chapter 5.1.2 --- Analysis on relation between weight loss and survival --- p.82 / Chapter 5.1.2.1 --- Patient and cancer stage --- p.82 / Chapter 5.1.2.2 --- Percentage of patients with weight loss at end of radiotherapy --- p.82 / Chapter 5.1.2.3 --- Cancer treatment outcome --- p.82 / Chapter 5.1.2.4 --- Univariate analysis --- p.82 / Chapter 5.1.2.5 --- Multivariate analysis --- p.83 / Chapter 5.2 --- PROSPECTIVE STUDY --- p.84 / Chapter 5.2.1 --- The profile of nutritional measurements during radiotherapy --- p.84 / Chapter 5.2.1.1 --- Bodyweight and body composition before and during RT --- p.84 / Chapter 5.2.1.2 --- Calorie intake before and during RT --- p.85 / Chapter 5.2.1.3 --- Energy expenditure before and during RT --- p.86 / Chapter 5.2.1.4 --- Energy balance before and during RT --- p.88 / Chapter 5.2.2 --- The profile of nutritional measurements during the 6-month period after radiotherapy --- p.88 / Chapter 5.2.2.1 --- Body weight and body composition during the 6 months after radiotherpay --- p.88 / Chapter 5.2.2.2 --- Calorie intake during the 6 months after radiotherapy --- p.90 / Chapter 5.2.2.3 --- Energy expenditure during the 6 months after radiotherapy --- p.91 / Chapter 5.2.2.4 --- Energy balance during the 6 months after radiotherapy --- p.92 / Chapter 5.2.3 --- Radiotherapy-induced Symptoms --- p.92 / Chapter 5.2.3.1 --- Profile of symptoms after RT --- p.92 / Chapter 5.2.3.2 --- Detailed profile of symptoms during RT --- p.95 / Chapter 5.2.4 --- The profile of cytokines during and after completion of RT --- p.95 / Chapter 5.2.4.1 --- SerumTNF-α --- p.96 / Chapter 5.2.5.2 --- Serum leptin --- p.96 / Chapter CHAPTER 6 --- DISCUSSION --- p.144 / Chapter 6.1 --- RETROSPECTIVE STUDY --- p.145 / Chapter 6.2 --- PROSPECTIVE STUDY --- p.147 / Chapter 6.2.1 --- The magnitude of the problem --- p.147 / Chapter 6.2.2 --- The potential determinants of weight loss --- p.147 / Chapter 6.2 3 --- Is weight loss due to the cancer or due to its treatment? --- p.148 / Chapter 6.2.4 --- "Is the weight loss influenced by pre-treatment factors, i. e. anthropometrical data?" --- p.148 / Chapter 6.2.5 --- "Is the pattern weight loss compatible with the model of calorie-protein malnutrition, similar to a starvation state?" --- p.149 / Chapter 6.2.6 --- Is the weight loss due to increased energy expenditure? --- p.150 / Chapter 6.2.7 --- Is the weight loss due to reduced calorie intake? --- p.151 / Chapter 6.2.8 --- Is weight loss during radiotherapy due to negative energy balance? --- p.151 / Chapter 6.2.9 --- What causes reduced calorie intake: Are radiotherapy-induced symptoms contributive? --- p.152 / Chapter 6.2.10 --- What are the observations on and implications of cytokine changes? --- p.155 / Chapter 6.2.11 --- What determines the recovery of body weight during 6 months after end of radiotherapy? --- p.158 / Chapter 6.2.12 --- Is the weight loss in the post RT recovery period due to negative energy balance? --- p.159 / Chapter 6.2.13 --- What are the implications on nutritional intervention? --- p.159 / Chapter 6.2.14 --- Limitations and future studies --- p.164 / Chapter CHAPTER 7 --- CONCLUSIONS --- p.166 / REFERENCES --- p.169 / APPENDIX 1 CONSENT FORM I (IN ENGLISH) --- p.190 / APPENDIX 2 CONSENT FORM I (IN CHINESE) --- p.193 / APPENDIX 3 CONSENT FORM II (IN ENGLISH) --- p.196 / APPENDIX 4 CONSENT FORM II (IN CHINESE) --- p.199 / APPENDIX 5 3-DAY DIET RECORD --- p.202 / APPENDIX 6 24-HOUR DIETARY RECALL --- p.206 / APPENDIX 7 SUBJECTIVE NUTRITIONAL ASSESSMENT --- p.208 / APPENDIX 8 PHYSICAL ACTIVITY QUESTIONNAIRE --- p.210 / APPENDIX 9 BONE SCAN REPORT --- p.215

Hui Wai Ying. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 158-189). / Abstracts in English and Chinese. / Abstract / Acknowledgements / List of Tables / List of Figures / Table of Contents / Table of Contents / Chapter Chapter I: --- Introduction --- p.1 / Chapter I. --- Aim of Study --- p.1 / Chapter II. --- Literature Review --- p.3 / Chapter 1. --- Background --- p.3 / Chapter A. --- Epidemiology --- p.3 / Chapter B. --- Histopathology --- p.3 / Chapter C. --- Etiology --- p.4 / Chapter i. --- Environmental Factors --- p.5 / Chapter ii. --- Epstein-Barr Virus (EBV) Infection --- p.6 / Chapter iii. --- Genetic Factors --- p.9 / Chapter 2. --- Molecular Genetics of NPC --- p.11 / Chapter A. --- Genome-Wide Studies --- p.11 / Chapter i. --- Comparative Genomic Hybridization (CGH) --- p.11 / Chapter ii. --- Loss of Heterozygosity (LOH) Studies --- p.12 / Chapter iii. --- Homozygous Deletion Study --- p.12 / Chapter B. --- NPC-related Oncogenes and Tumor Suppressor Genes --- p.13 / Chapter i. --- Oncogenes --- p.13 / Chapter ii. --- Tumor Suppressor Genes --- p.14 / Chapter 3. --- Chromosome 14q and NPC --- p.19 / Chapter A. --- Tumor Suppressor Loci and Cancer-Related Genes on Chromosome14q --- p.20 / Chapter i. --- Tumor Suppressor Loci on Chromosome 14q --- p.20 / Chapter ii. --- Cancer-Related Genes on Chromosome 14q --- p.26 / Chapter 4. --- Chromosome 16q and NPC --- p.28 / Chapter A. --- Tumor Suppressor Loci and Candidate Tumor Suppressor genes on Chromosome16q --- p.28 / Chapter i. --- Tumor Suppressor Loci on Chromosome 16q --- p.28 / Chapter ii. --- Metastasis Suppressor Loci on Chromosome 16q --- p.34 / Chapter iii. --- Candidate Tumor Suppressor Genes on Chromosome 16q --- p.34 / Chapter Chapter II: --- Materials and Methods --- p.40 / Chapter I. --- Cell Lines and Xenografts --- p.40 / Chapter 1. --- Cell Lines --- p.40 / Chapter 2. --- Xenografts --- p.41 / Chapter 3. --- DNA Extraction --- p.42 / Chapter II. --- Patients and Biopsy Specimens --- p.44 / Chapter 1. --- Manual Microdissection --- p.44 / Chapter 2. --- Laser Captured Microdissection (LCM) --- p.46 / Chapter 3. --- DNA Extraction --- p.46 / Chapter III. --- Comprehensive Screening for Homozygous Deletion Regions on Chromosomes 14q32.12-32.33 and 16q23.1-24.3 in Human Cancers --- p.48 / Chapter 1. --- DNA of Human Cancer Cell Lines --- p.48 / Chapter 2. --- Sequence-Tagged Sites (STS) Markers --- p.48 / Chapter 3. --- Polymerase Chain Reaction (PCR) --- p.49 / Chapter IV --- . Investigation of Inactivation of Potential Tumor Suppressor Genes on Chromosome 14q32.12-32.33 and 16q23.1-24.3 --- p.58 / Chapter 1. --- Detection of Homozygous Deletion --- p.58 / Chapter 2. --- Expression Analysis --- p.58 / Chapter A. --- RNA Extraction --- p.58 / Chapter B. --- Reverse-Transcription (RT) PCR --- p.61 / Chapter i. --- DNase I Digestion --- p.62 / Chapter ii. --- First-strand cDNA Synthesis and RNase Digestion --- p.62 / Chapter iii. --- Reverse-Transcription (RT)-PCR --- p.63 / Chapter C. --- Real-Time RT PCR --- p.63 / Chapter 3. --- Methylation Analysis --- p.68 / Chapter A. --- Sodium Bisulfite Modification --- p.68 / Chapter B. --- Methylation-Specific PCR (MSP) --- p.69 / Chapter C. --- Bisulfite Sequencing --- p.70 / Chapter D. --- Combined Bisulfite Restriction Analysis (COBRA) --- p.75 / Chapter E. --- 5 -aza-2' -deoxycytidine Treatment --- p.76 / Chapter ChapterIII： --- Results --- p.78 / Chapter I. --- Comprehensive Screening for Homozygous Deletion Regions in Human Cancers --- p.78 / Chapter 1. --- Chromosome 14q32.12-3233 --- p.78 / Chapter 2. --- Chromosome 16q23.1-243 --- p.79 / Chapter II. --- Investigation of Inactivation of Potential Tumor Suppressor Genes in NPC --- p.86 / Chapter 1. --- Chromosome 14q --- p.86 / Chapter A. --- "WW Domain-Containing Protein, 45-kD （WW45)" --- p.86 / Chapter B. --- Apoptosis Stimulating Protein of p53（ASPP1) --- p.88 / Chapter 2. --- Chromosome 16q --- p.92 / Chapter A. --- WW Domain-Containing Oxidoreductase （WWOX) --- p.92 / Chapter i. --- Homozygous Deletion Screening of WWOX --- p.92 / Chapter ii. --- Expression of Aberrant Splicing Transcripts of WWOX in NPC --- p.94 / Chapter iii. --- Sequencing of WWOX Aberrant Transcripts --- p.95 / Chapter iv. --- Quantitative Analysis of WWOX Transcripts in NPC --- p.95 / Chapter v. --- Methylation Analysis --- p.99 / Chapter B. --- H-Cadherin (CDH13) --- p.102 / Chapter i. --- Analysis of H-cadherin Deletion on Cancer Cell Lines and Xenografts --- p.102 / Chapter ii. --- Expression Analysis of H-Cadherin by RT-PCR and Real-Time RT-PCR --- p.102 / Chapter iii. --- Analysis of Promoter Hypermethylation by Methylation-Specific PCR (MSP) and Bisulfite Sequencing in NPC Cell Lines and Xenografts --- p.104 / Chapter iv. --- Demethylation Study of H-Cadherin in C666-1 Cell Line --- p.105 / Chapter v. --- Methylation Analysis of H-Cadherin in Primary Tumors --- p.105 / Chapter vi. --- Methylation Analysis of H-Cadherin in Human Cancer Cell Lines --- p.106 / Chapter C. --- Myeloid Translocation Gene on Chromosome 16 (MTG16) --- p.113 / Chapter i. --- Deletion Analysis of MTG16 in Cancer Cell Lines and Xenografts --- p.113 / Chapter ii. --- Differential Expression of MTG16a and MTG16b Transcripts in NPC Cell Lines and Xenografts --- p.113 / Chapter iii. --- Methylation Analysis of MTG16b in NPC Cell Lines and Xenografts --- p.118 / Chapter iv. --- Sequencing of MTGl 6b RT-PCR Products --- p.119 / Chapter v. --- Demethylation Study of MTG16b in HK-1 Cell Line --- p.119 / Chapter vi. --- Promoter Methylation Analysis of MTG16b by MSP in Primary NPC and Cancer Cell Lines --- p.120 / Chapter Chapter IV: --- Discussion --- p.124 / Chapter I. --- Comprehensive Homozygous Deletion Screening of Chromosomes 14q32.12-32.33 and 16q23.1-24.3 in Human Cancer Cell Lines and Xenografts --- p.124 / Chapter II. --- Investigation of Candidate Tumor Suppressor Genes on Chromosome 14q in NPC --- p.128 / Chapter III. --- Alterations of Candidate Tumor Suppressor Genes on Chromosome 16q in NPC --- p.133 / Chapter 1. --- Expression of Aberrant Transcripts of WWOX in NPC --- p.133 / Chapter 2. --- Methylation-Associated Silencing of H-Cadherin and MTG16b in NPC --- p.140 / Chapter Chapter V: --- Conclusion --- p.154 / Chapter Chapter VI: --- References --- p.158

1. Study on improving the diagnostic accuracy of treatment-naive nasopharyngeal carcinoma. / 2. Study on diagnostic accuracy of EBV-DNA on recurrent nasopharyngeal carcinoma. / 3. Studies on EBV-DNA as a screening tool for nasopharyngeal carcinoma. Part 1. To define the detection rate of NPC and the false-positive rate of IgA-VCA in an IgA-VCA-based screening problem, and to define the specificity of IgA-EA in IgA-VCA-positive screenees. Part 2. To define the specificity of EBV-DNA in IgA-VCA-positive screenees. Part 3. To define the sensitivity of IgA-EA, and EBV-DNA in IgA-positive NPC patients. / 4. Studies on pre-therapy prognostication of nasopharyngeal carcinoma Study Part 1. Objective. To assess the role of EBV-DNA in pre-therapy prognostication of early-stage NPC. / Background. The specific association between nasopharyngeal carcinoma (NPC) and the Epstein-Barr virus (EBV) had been exploited to develop a spectrum of EBV-antibodies-based blood markers. Among these markers, the Immunoglobulin A antibody against the viral capsid antigen (IgA-VCA) of the EBV has been the most popularly employed marker to assist diagnosis of NPC. There is however a relative paucity of data on the application of blood markers for screening, for detection of relapse, and for prognostification of patient cohorts managed in present-day therapy oncology protocols. Peripheral blood EBV-DNA, measured by quantitative polymerase chain reaction assay, is a newly-developed marker, and represents a prototype model of a nuclei acid-based, as opposed to antibody-based, EBV tumor marker for NPC. The present thesis describes the translation of this basic scientific advance into clinical applications, through several prospective and retrospective studies that address the diagnosis of treatment-naive NPC, the detection of recurrent NPC, the screening of individuals at risk of NPC, the pre-therapy prognostication for NPC to guide for choice of therapy. The role of integration of conventional markers and EBV-DNA in clinical applications was also examined. / Study Part 2. Objectives. To assess whether incorporation of EBV-DNA data to TNM staging improves prognostic discrimination of patients subsets within individual cancer stage, to assess if EBV-DNA is an independent prognostic factor for survival after ontological therapy. (Abstract shortened by UMI.) / Leung Sing-fai. / "February 2005." / Source: Dissertation Abstracts International, Volume: 67-07, Section: B, page: 3695. / Thesis (M.D.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references. / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / School code: 1307.

Celecoxib is a COX-2 selective non-steroidal anti-inflammatory drug which has been shown to inhibit growth and induce apoptosis in various cancer cell lines. Using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and an apoptosis detection kit, we demonstrated that celecoxib was able to induce growth inhibition and apoptosis in a dose-dependent manner in 3 NPC cell lines: HK-1, Hone-1, and C666-1. Afterwards, a proteomic approach was used to study the underlying mechanisms involved in celecoxib-mediated effects on two COX-2 positive NPC cell lines (HK-1 and C666-1). Results showed that a total of 18 protein spots were differentially expressed in the HK-1 and C666-1 cells. On the other hand, we also compared the proteomic expression profile between an NPC cell line (C666-1) and a normal nasopharynx cell line (NP69) in order to study whether those differentially expressed proteins after celecoxib treatment were also involved in NPC carcinogenesis. Proteomics results with confirmation using Western blotting discovered that HSP27 phosphorylated of serine 82 (HSP27-pSer82) protein was up-regulated in C666-1 cells when compared with that in NP69 cells. After treatment with celecoxib, expression of HSP27-pSer82 protein was down-regulated in both HK-1 and C666-1 cells. These findings suggest that down-regulation of HSP27-pSer82 protein expression may have mediated the growth-inhibitory effects of celecoxib in HK-1 and C666-1 cells. Finally, other differential expressed proteins identified from proteomics with confirmation by immunocytochemical staining in the 2 NPC cell lines and 40 NPC patient specimens showed that down-regulation of annexin 2 and beta2-tubulin may be important in NPC formation. / COX-2 over-expression has been found in various cancers such as colorectal cancer, liver cancer and lung cancer. In vivo studies have shown that mice overexpressing COX-2 developed breast cancer whereas COX-2 knockout mice had reduced rates of cancer formation in the intestines and skin. In the present study, COX-2 expression in NPC patient biopsies was examined and correlated with the clinicopathological data of the patients. Immunocytochemical staining showed that COX-2 protein was over-expressed in 84.6% (66/78) of non-metastatic NPC patients and was associated with an advanced nodal stage (P<0.05). All these data support an important role for COX-2 in NPC pathogenesis. / In summary, this study is the first to identify HSP27-pSer82 protein as a potential target of celecoxib in NPC cells. Detailed investigations of the functional role of molecular targets identified in this study would improve our understanding of the chemotherapeutic effects of celecoxib and, in the long run, may lead to a more effective chemotherapeutic treatment to this common cancer. / Nasopharyngeal carcinoma (NPC) is prevalent in southern China. Although early stage patients have a high rate of cure with radiotherapy alone, the prognosis for those with stage III or IV disease remains poor due to subsequent development of distant metastases. Therefore there is an urgent need to develop novel biologic agents to improve treatment outcomes. / Chan, Ming Lok. / Adviser: Anthony T.C. Chan. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3418. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 141-171). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

(1) MRI is shown to be an accurate test for detecting NPC and one which has the potential to be used in screening to (a) screen out normal patients who do not require endoscopie biopsy; and (b) identify small tumours that would be missed on endoscopy. / (2) At diagnosis staging NPC by MRI reveals that oropharyngeal and maxillary sinus invasion are markers of more advanced disease than reflected in current staging system. Tumour involvement of the preclival/prevertebral region, skull base and retropharyngeal nodes are more common than previously recognised by computed tomography, while parapharyngeal tumour invasion is less common. The latter resuits from the superior ability of MRI to differentiate primary tumours with true parapharyngeal invasion from those contained within the nasopharynx which are causing bowing of the wall or lie adjacent to a retropharyngeal node. / (3) Post treatment complications were detected by MRI in over 50% of patients. Neural damage, especially to the temporal lobes and 12th cranial nerves, was the most frequent complication (48%), followed by osteoradionecrosis (20%) involving the mandible, upper cervical spine and skull base, the latter including destruction of the roof of the nasopharynx. Malignant tumours and unusual benign masses (6%) showed radiological features useful in the differential diagnosis from NPC recurrence. Malignant tumours were mainly squamous cell carcinomas or sarcomas showing a predilection for the maxillary region, tongue and external auditory canal. The unusual benign masses were found in the nasopharynx/sphenoid sinus. / (4) Finally functional MRI using DWI and 1H-MRS are feasible in the technically challenging region of the head and neck. Choline ratios and ADC values of NPC are established. The successful demonstration of differences between the biomarkers of NPC and those of lymphoma and squamous cell carcinoma, show that functional MRI is a new tool for the evaluation of NPC, opening up the possibility that these biomarkers can be used for monitoring NPC treatment response in the future. / Magnetic resonance imaging (MRI) provides excellent anatomical detail and functional information about cancer. This thesis explores the role of MRI in the assessment of nasopharyngeal carcinoma (NPC) from detection through to the long term complications of radiotherapy treatment. / Ann D King. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3468. / Thesis (M.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 180-191). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English only. / School code: 1307.

Advanced-stage nasopharyngeal carcinoma (NPC) presents very difficult scenarios for radiation therapy (RT) planning. The infiltration of tumor to the skull base and beyond means that the tumor is very close to critical normal organs (organs at risk, OARs). Despite the advent of intensity-modulated radiotherapy (IMRT) treatment technique---the state-of-art RT technique, conflicting requirements between organ protection and target dose conformity is still problematic. The objectives of the present research are (1) to investigate the dosimetry properties of IMRT treatment in advanced-stage NPC in respect of its dosimetric limitations and planning problems, (2) to develop new methods and tools to resolve such problems, in particular to improve the quality of treatment plans and efficiency of the dose planning and optimization process. A series of four inter-linked studies were conducted to address these issues. / In conclusion, the solutions to several major problems in IMRT planning for advanced-stage NPC were investigated and established. It has been demonstrated in this research that, by applying these methods and tools, significant improvement in the dosimetry and efficiency of IMRT treatment planning can be accomplished as compared with conventional IMRT planning techniques. It is expected that such would translate into an improvement in treatment throughput, better tumor control and reduction in normal tissues complications. The methods developed have potential to be applied to all stages of NPC and to other tumor sites. / The first study was to improve the efficacy in target coverage and organs sparing using an "organ-splitting" approach. The OARs which overlapped with targets were split into target-overlapping and non-overlapping segments and each segment was assigned with different constraints parameters to increase the degree of flexibility during optimization. As a result, a steep gradient in the dose distribution at the regions of interface between the targets and normal critical organs could be achieved and treatment quality was improved. In the second study, a thorough dosimetric comparison between conventional 2-dimensional (2D) RT and IMRT plans was conducted to determine, with reference to outcome of 2D treatments, the extended tolerance dose limits for the critical organs, especially that of the brainstem and spinal cord, and their planning organ at risk volume. Such data could then serve as reference in IMRT planning when the dose of critical organs need be exceeded in order to allow adequate dose to a very close by target. In the third study, the feasibility of using interpolated contours for segmentation of targets and OARs in IMRT planning was investigated. The result indicated that the use of interpolated contours in IMRT planning could significantly reduce the contouring time by about 50% without degrading the target coverage and OARS sparing. In the final study, an array of dose constraint templates that could accommodate different degrees of overlap between the targets and OARs, together with a template selection program, were developed to improve the efficiency of IMRT planning. By applying the methods and tools developed, IMRT treatment planning of advanced NPC could become more efficient and less dependent on planner's experience. / Chau, Ming Chun. / Adviser: Anthony Chan Tak Cheung. / Source: Dissertation Abstracts International, Volume: 69-02, Section: B, page: 0948. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 118-128). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / School code: 1307.