The Best PhD Thesis in the Faculties of Dentistry, Engineering, Medicine and Science (University of Hong Kong), Li Ka Shing Prize,2001-2003 / published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

Tang Angie. / Thesis submitted in: November 2005. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 151-172). / Abstracts in English and Chinese. / Acknowledgements --- p.i / Abstract --- p.iii / Abstract in Chinese --- p.vi / Table of Contents --- p.vii / List of Publications --- p.xi / Awards --- p.xii / List of Abbreviations --- p.xiii / List of Tables --- p.xv / List of Figures --- p.xvi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter Chapter 2 --- Literature Review --- p.3 / Chapter 2.1 --- Gastric cancer-overview --- p.3 / Chapter 2.1.1 --- Epidemology --- p.3 / Chapter 2.1.2 --- Pathology --- p.3 / Chapter 2.1.3 --- Etiologies and Risk Factors --- p.4 / Chapter I. --- Environmental factors --- p.4 / Chapter a. --- Helicobacter pylori infections --- p.4 / Chapter b. --- Epstein-Barr virus (EBV) --- p.6 / Chapter c. --- Dietary factors --- p.6 / Chapter d. --- Smoking --- p.6 / Chapter II. --- Genetic Factors --- p.7 / Chapter a. --- Hereditary Gastric Cancer --- p.7 / Chapter b. --- Genetic polymorphism --- p.8 / Chapter III. --- Cyclooxygenases (COX) enzymes --- p.10 / Chapter IV. --- Molecular carcinogenesis --- p.11 / Chapter a. --- Activation of proto-oncogenes --- p.11 / Chapter b. --- Candidate tumor suppressor genes --- p.12 / Chapter 1. --- Gene mutation and deletion --- p.12 / Chapter 2. --- Epigenetic Silencing --- p.13 / Chapter 2.2 --- Epigenetics --- p.14 / Chapter 2.2.1 --- DNA methylation --- p.15 / Chapter 2.2.2 --- Histone modification --- p.28 / Chapter I. --- Histone acetylation and deacetylation --- p.32 / Chapter II. --- Histone methylation --- p.32 / Chapter III. --- Histone phosphorylation --- p.34 / Chapter IV. --- Histone ubiquitylation --- p.34 / Chapter 2.3 --- "HAT, HDAC and HDAC inhibitors" --- p.36 / Chapter 2.3.1 --- HAT --- p.38 / Chapter 2.3.2 --- HDAC --- p.39 / Chapter (a) --- Class I --- p.40 / Chapter (b) --- Class II --- p.41 / Chapter (c) --- Class III --- p.42 / Chapter (d) --- Mammalian HDAC and their mechanism of deacetylation --- p.44 / Chapter 2.3.3 --- HDAC inhibitors --- p.45 / Chapter I. --- Class I/II natural inhibitors --- p.47 / Chapter II. --- Class I/II synthetic inhibitors --- p.48 / Chapter III. --- Sirtuins inhibitors --- p.49 / Chapter IV. --- Activity of HDAC inhibitors in vitro --- p.50 / Chapter a. --- Effect in the gene expression --- p.50 / Chapter b. --- Non-transcriptional effects --- p.55 / Chapter c. --- Activity of HDAC inhibitors with other agents --- p.57 / Chapter d. --- Effects in xenograft tumor models --- p.57 / Chapter V. --- Clinical trials of HDAC inhibitors --- p.59 / Chapter Chapter 3 --- Aims of the study --- p.63 / Chapter Chapter 4 --- Materials and Methods --- p.64 / Chapter 4.1 --- Cell culture --- p.64 / Chapter 4.2 --- Drug treatment --- p.64 / Chapter 4.2.1 --- Suberoylanilide Hydroxamic Acid treatment --- p.64 / Chapter 4.2.2 --- Trichostatin A treatment --- p.65 / Chapter 4.3 --- Cell proliferation assay --- p.66 / Chapter 4.4 --- Apoptotic assay --- p.67 / Chapter 4.5 --- Flow cytometry --- p.67 / Chapter 4.5.1 --- Cell preparation --- p.67 / Chapter 4.5.2 --- Propidium Iodide staining --- p.68 / Chapter 4.5.3 --- Annexin V-FITC staining --- p.68 / Chapter 4.5.4 --- Flow cytometer analysis --- p.69 / Chapter 4.6 --- Total RNA extraction --- p.70 / Chapter 4.7 --- DNA extraction --- p.71 / Chapter 4.8 --- Protein extraction --- p.72 / Chapter 4.9 --- Western blottng --- p.72 / Chapter 4.10 --- Microarray analysis --- p.74 / Chapter 4.10.1 --- Sample preparation for microarray --- p.74 / Chapter 4.10.2 --- Hybridization --- p.75 / Chapter 4.10.3 --- Scanning and data processing --- p.75 / Chapter 4.10.4 --- Data analysis --- p.76 / Chapter 4.11 --- Primer design --- p.77 / Chapter 4.12 --- RT-PCR --- p.77 / Chapter 4.12.1 --- Reverse transcription --- p.77 / Chapter 4.12.2 --- Quantitative RT-PCR --- p.78 / Chapter 4.13 --- Methlyation study --- p.79 / Chapter 4.13.1 --- Demethylation by 5-aza-2'deoxycytidine --- p.79 / Chapter 4.13.2 --- Bisulfite modification --- p.79 / Chapter 4.13.3 --- Methylation-specific PCR (MSP) --- p.79 / Chapter Chapter 5 --- Results --- p.81 / Chapter 5.1 --- Morphological changes in AGS cells --- p.81 / Chapter 5.2 --- Anti-cancer effects of HDAC inhibitors --- p.81 / Chapter 5.2.1 --- Effect of HDAC inhibitors on cell growth --- p.81 / Chapter a. --- SAHA inhibits cell proliferation --- p.82 / Chapter b. --- TSA inhibits cell proliferation --- p.82 / Chapter 5.2.2 --- Cell cycle analysis --- p.87 / Chapter a. --- Effect of SAHA on cell cycle --- p.87 / Chapter b. --- Effect of TSA on cell cycle --- p.88 / Chapter 5.2.3 --- Induction of apoptosis on AGS cells --- p.92 / Chapter a. --- SAHA induces apoptotic cell death --- p.92 / Chapter b. --- TSA induces apoptotic cell death --- p.94 / Chapter 5.3 --- Induction of histone expression on AGS cells --- p.102 / Chapter 5.3.1 --- HDAC inhibitors induced acetylation of histone H3 --- p.102 / Chapter 5.3.2 --- HDAC inhibitors induced acetylation of histone H4 --- p.103 / Chapter 5.4 --- SAHA- and TSA-induced gene expression profiles --- p.106 / Chapter 5.5 --- Verification of gene expression by quantitative RT-PCR --- p.108 / Chapter 5.6 --- Methylation study --- p.113 / Chapter Chapter 6 --- Discussion --- p.116 / Chapter 6.1 --- Improved treatment strategy is needed for gastric cancer. --- p.116 / Chapter 6.2 --- HDAC inhibitors as potential anti-cancer agents --- p.117 / Chapter 6.3 --- Potential anti-cancer effect of TSA and SAHA on AGS cells --- p.120 / Chapter I. --- Morphological changes of AGS gastric cancer cells --- p.120 / Chapter II. --- Inhibition of cell proliferation --- p.120 / Chapter III. --- Induction of cell cycle arrest --- p.121 / Chapter IV. --- Induction of apoptosis --- p.122 / Chapter 6.4 --- Expression of acetylated histones upon treatment with TSA and SAHA --- p.124 / Chapter 6.5 --- Identify potential target genes upon treatment with TSA and SAHA --- p.125 / Chapter 6.5.1 --- Candidate genes involved in cell cycle --- p.126 / Chapter a. --- P21WAF1 --- p.126 / Chapter b. --- p27kip1. --- p.128 / Chapter c. --- Cyclin E & Cyclin A --- p.128 / Chapter d. --- Signal-induced proliferation-associated gene 1 (SIPA1) .… --- p.129 / Chapter 6.5.2 --- Candidate genes involved in apoptosis and anti-proliferation --- p.130 / Chapter a. --- BCL2-interacting killer (apoptosis-inducing) (BIK) (Pro-apoptotic gene) --- p.131 / Chapter b. --- Thioredoxin interacting protein (TXNIP) (Proapoptotic gene) / Chapter c. --- Cell death-inducing DFFA-like effector b (CIDEB) (apoptosis induction) --- p.132 / Chapter d. --- B-cell translocation gene 1 (BTG1) - (anti-proliferation) --- p.133 / Chapter e. --- Quiescin 6 (QSCN6) (anti-proliferation) --- p.133 / Chapter f. --- "Cysteine-rich, angiogenic inducer, 61 (CYR61) (anti-proliferative)" --- p.134 / Chapter g. --- Metallothionein 2A (MT2A) (apoptosis induction and anti-proliferative) --- p.134 / Chapter 6.5.3 --- Other genes reported to be up-regulated with HDAC inhibitors treatment --- p.135 / Chapter a. --- Glia maturation factor-gamma (GMFG) --- p.135 / Chapter b. --- v-fos FBJ murine osteosarcoma viral oncogene homolog (FOS) / Chapter c. --- Interleukin 8 (IL-8) --- p.136 / Chapter d. --- Insulin-like growth factor binding protein- 2 (IGFBP2) --- p.137 / Chapter e. --- Integrin alpha chain 7 (ITGA7) --- p.138 / Chapter 6.5.4 --- Selected highly up-regulated genes with HDAC inhibitors treatment --- p.139 / Chapter a. --- Aldo-keto reductase family 1，member C3 (AKR1C3) --- p.139 / Chapter b. --- GPI-anchored metastasis-associated protein homolog (C4.4A) --- p.139 / Chapter c. --- "Serine (or cysteine) proteinase inhibitor，clade I (neuroserpin), member 1 (SERPINI1)" --- p.140 / Chapter d. --- "Serine (or cysteine) proteinase inhibitor，clade E (nexin, plasminogen activator inhibitor type 1), member 1 (SERPINE1)" --- p.140 / Chapter e. --- Adrenomedullin (ADM) --- p.141 / Chapter f. --- Dehydrogenase/reductase (SDR family) member 2 (HEP27) --- p.142 / Chapter g. --- Cholecystokinin (CCK) --- p.142 / Chapter h. --- Silver homolog (mouse) (SILV) --- p.143 / Chapter 6.6 --- Genes regulated by gene promoter hypermethylation in AGS cells --- p.143 / Chapter Chapter 7 --- Conclusion --- p.147 / Chapter Chapter 8 --- Further Studies --- p.150 / References --- p.151 / Appendix I --- p.151 / Appendix II --- p.III / Appendix III --- p.IV / Appendix IV --- p.VI

In the past, patients with peritoneal surface malignancy were considered incurable and were only offered palliative treatments. However, in a substantial number of patients, disease progression that is isolated to peritoneum may occur. It has been realised that elimination of peritoneal surface tumours may have an impact on the survival of these cancer patients, in whom a prominent cause of death is peritoneal carcinomatosis. The focus of this PhD. thesis is on the combined treatment of cytoreductive surgery and perioperative intrapersonal chemotherapy for diffuse malignant peritoneal mesothelioma, pseudomyxoma peritonei, colorectal peritoneal carcinomatosis and resectable gastric cancer. Section one describes the major principles of management for peritoneal surface malignancy, covering the historical perspectives, the treatment rationales and the learning curve associated with the combined procedure. Section two is devoted to peritoneal mesothelioma, in trying to examine this disease from its clinical, radiologic and histopathologic aspects. A radiologic classification and a histopathologic staging system for this disease are proposed. In section three, the results of the combined treatment for pseudomyxoma peritonei are presented, including a systematic review of the literature, a case series of 50 patients from our Australian centre and a treatment failure analysis of 402 patients from the Washington Cancer Institute. These studies suggest that a disease-free state is important for long-term survival for patients with pseudomyxoma peritonei. In section four, the current evidence on the combined treatment for colorectaI peritoneal carcinomatosis is demonstrated by conducting a systematic review of the literature and survival and perioperative outcome analyses of two separate patient cohorts. These results suggest that the combined treatment is associated with an improved survival, as compared with historical controls. In the last section, a metaanalysis of the randomised controlled trials on adjuvant intraperitoneal chemotherapy for resectable gastric cancer shows that a significant improvement in survival is associated with hyperthermic intraoperative intraperitoneal chemotherapy alone or in combination with early postoperative intraperitoneal chemotherapy.