He, Jun. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 136-156). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Chan Siu-chung Andrew. / "December 2000." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (p. 120-136). / 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 Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Wing Y. Chan. / "March 2000." / Thesis (M.D.)--Chinese University of Hong Kong, 2000. / 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 Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

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

Mak, Ko Fung. / Thesis submitted in: July 2003. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 113-138). / Abstracts in English and Chinese. / Chapter CHAPTER I: --- INTRODUCTION --- p.1 / Chapter I. --- AIM OF STUDY --- p.1 / Chapter II. --- LITERATURE REVIEW --- p.2 / Chapter 1. --- Epidemiology --- p.2 / Chapter 2. --- Etiology --- p.4 / Chapter A. --- Heredity --- p.4 / Chapter i. --- Family History --- p.4 / Chapter ii. --- Inherited Predisposition --- p.4 / Chapter B. --- Hormonal --- p.7 / Chapter C. --- Environmental --- p.9 / Chapter i. --- Diet --- p.9 / Chapter ii. --- Radiation --- p.10 / Chapter iii. --- Physical Activity --- p.11 / Chapter 3. --- Histopathology --- p.12 / Chapter 4. --- Clonality Nature of Cancer --- p.13 / Chapter 5. --- "Knudson ""two-hit"" Hypothesis on Cancer Development" --- p.14 / Chapter 6. --- Molecular Genetic Studies of Breast Cancer --- p.15 / Chapter A. --- Loss of Heterozygosity --- p.16 / Chapter B. --- Comparative Genomic Hybridization --- p.19 / Chapter C. --- Epigenetic Changes --- p.20 / Chapter 7. --- Genetic Changes in Breast Cancer --- p.21 / Chapter A. --- Chromosome 1 --- p.21 / 14-3-3σ Gene --- p.21 / Chapter B. --- Chromosome 3 --- p.22 / Chapter i. --- Retionoic Acid Receptor p2 Gene --- p.22 / Chapter ii. --- Fragile Histidine Traid Gene --- p.24 / Chapter iii. --- Ras Associated Domain Family 1A Gene --- p.25 / Chapter iv. --- Thyroid Hormone Receptor β1 Gene --- p.26 / Chapter C. --- Chromosome 5 --- p.27 / Adenomatous Polyposis Coli Gene --- p.27 / Chapter D. --- Chromosome 6 --- p.28 / Estrogen Receptor Gene --- p.28 / Chapter E. --- Chromosome 9 --- p.29 / p16 Gene --- p.29 / Chapter F. --- Chromosome 13 --- p.30 / Chapter i. --- BRCA2 Gene --- p.31 / Chapter ii. --- Retinoblastoma Gene --- p.32 / Chapter G. --- Chromosome 16 --- p.33 / E-cadherin Gene --- p.33 / Chapter H. --- Chromosome 17 --- p.34 / Chapter i. --- TP53 Gene --- p.34 / Chapter ii. --- BRCA1 Gene --- p.36 / Chapter CHAPTER II: --- MATERIALS AND METHODS --- p.38 / Chapter I. --- PATIENTS AND SPECIMENS --- p.38 / Chapter II. --- FROZEN SECTIONS AND MICRODISSECTION --- p.41 / Chapter III. --- DNA EXTRACTION --- p.43 / Chapter IV. --- ALLELOTYPING --- p.44 / Chapter 1. --- Polymerase Chain Reaction --- p.44 / Chapter 2. --- Electrophoresis --- p.45 / Chapter 3. --- Data Analysis --- p.46 / Chapter CHAPTER III: --- RESULTS --- p.49 / Chapter I. --- ALLELOTYPING ANALYSIS --- p.49 / Chapter II. --- FREQUENCY OF LOH --- p.68 / Chapter III. --- FRACTIONAL ALLELIC LOSS --- p.70 / Chapter IV. --- MINIMAL DELETION REGIONS --- p.72 / Chapter 1. --- Chromosome 6q --- p.83 / Chapter 2. --- Chromosome 8p --- p.83 / Chapter 3. --- Chromosome 8q --- p.84 / Chapter 4. --- Chromosome 10q --- p.84 / Chapter 5. --- Chromosome 13q --- p.85 / Chapter 6. --- Chromosome 14q --- p.85 / Chapter 7. --- Chromosome 16q --- p.86 / Chapter V. --- MICROSATELLITE INSTABILITY --- p.86 / Chapter CHATPER IV: --- DISCUSSION --- p.88 / Chapter I. --- COMPARISONS OF CHROMOSOMAL ALTERATIONS --- p.88 / Chapter II. --- MICROSATELLITE INSTABILITY --- p.92 / Chapter III. --- CHROMOSOMAL GAINS AND LOSSES --- p.93 / Chapter IV. --- CHROMOSOME 17 --- p.95 / Chapter V. --- MINIMAL DELETION REGIONS --- p.96 / Chapter 1. --- Chromosome 6q --- p.97 / Chapter 2. --- Chromosome 8p --- p.99 / Chapter A. --- 8p23.3.-22 --- p.99 / Chapter B. --- 8p21.3-12 --- p.101 / Chapter C. --- 8p22-21 --- p.103 / Chapter 3. --- Chromosome 8q --- p.104 / Chapter 4. --- Chromosome 10q --- p.106 / Haploinsufficiency and PTEN --- p.107 / Chapter 5. --- Chromosome 13q --- p.108 / Chapter 6. --- Chromosome 14q --- p.109 / Chapter 7. --- Chromosome 16q --- p.110 / Chapter CHAPTER V: --- CONCLUSION --- p.112 / REFERENCES --- p.113

MicroRNA（miRNAs）是一類細小的非編碼RNA（ncRNA），能透過轉錄後機制調節靶標基因的表達。miRNA的發現，不僅提出一個嶄新的基因調節機制，更強調了小ncRNA於不同的生理和發展過程中的重要性。最近的研究更進一步展示了miRNA失調與癌症發展之間的因果關係。 / 我們此前曾利用陣列分析，發現miRNA在肝細胞癌（HCC）中的失調模式，揭示了miR-145在HCC的普遍下調。在本論文的第一部分，定量逆轉錄聚合酶鏈反應（qRT-PCR）進一步證實了miR-145在50的肝細胞癌患者（n=80）的腫瘤中出現表達下調，而且miR-145的表達下調更與較短的无病生存期相關。其中一個低內源性miR-145的肝癌腫瘤樣本被建立為細胞株─HKCI-C2。此體外模型保持低miR-145水平，並於恢復miR-145表達後，抑製細胞存活和增殖。多個計算機演算法均預測了miR-145可針對胰島素樣生長因子（IGF）信號通路中的多個基因，包括胰島素受體底物（IRS1）-1，IRS2和胰島素樣生長因子1受體。這些假定目標的蛋白表達亦被miR-145下調。熒光素酶檢測進一步驗證了miR-145和IRS1/IRS2 3'-非編碼區的直接目標關聯。隨後的分析也確定miR-145能下調 IGF信號通路下游的信號傳導，即活性β-catenin水平。 / 最近出現的深度測序技術，為研究miRNome提供了一個前所未有的平台，以識別已知和新的miRNA。此外，現代生物信息學技術可同時對不同類型的小ncRNA，如PIWI-interacting RNA（piRNAs）進行分析。在本論文的第二部分中，我們利用Illumina大規模並行測序對兩個肝癌細胞株（HKCI-4和HKCI-8）和正常肝細胞株（MIHA）的小RNA轉錄組進行研究。生物信息學和生物功能分析揭示一種新型piRNA（取名為piR-Hep1）在肝腫瘤發生中的重要角色。在73例肝癌中，qRT-PCR結果顯示piR-Hep1在47的肝癌組織出現上調。PiR-Hep1的沉默能抑制肝癌細胞存活、遷移和侵襲，同時亦減少了Akt的磷酸化。在miRNA的分析中，miR-1323被發現在肝癌組織中大量表達，並與肝硬化背景下產生的肝腫瘤相關。此外，miR-1323出現過表達的肝硬化肝癌患者的無病和整體存活率亦較差（P<0.009）。 / 總觀來說，本論文首次發現miR-145可同時抑制引致肝癌的IGF信號通路中的多個傳導因子，亦突出了piR-Hep1的功能重要性和miR-1323在肝癌患者中的預後意義。此外，本研究表明，傳統的陣列分析和新一代的測序技術均能發現重要的miRNA。新一代測序技術對轉錄組的全面分析，將對研究各種不同類型的ncRNAs在肝癌發生發展過程中的參與提供新的思路。 / MicroRNAs (miRNAs) are a class of small non-coding RNAs (ncRNA) that post-transcriptionally regulate gene expression. The discovery of miRNAs not only puts forth an alternate gene regulatory mechanism, but also underscores the importance of small ncRNAs as pivotal regulators of diverse physiological and developmental processes. Recent studies have emphasized a causal link between miRNA deregulation and cancer development. / Our group has previously reported on dysregulated miRNA pattern in hepatocellular carcinoma (HCC) by array-based profiling, which revealed common downregulation of miR-145. In the first part of this thesis, quantitative reverse transcription polymerase chain reaction (qRT-PCR) corroborated reduced miR-145 expression in 50% of tumors in a cohort of 80 HCC patients, which also correlated reduced miR-145 expression with shorter disease-free survival of patients. One HCC tumor analyzed with low endogenous miR-145 was propagated as cell line. This in vitro model HKCI-C2 maintained low miR-145 level and upon restoration of miR-145 expression, a consistent inhibitory effect on cell viability and proliferation was readily observed. Multiple in silico algorithms predicted that miR-145 could target a number of genes along the insulin-like growth factor (IGF) signaling, including insulin receptor substrate (IRS1)-1, IRS2 and insulin-like growth factor 1 receptor. Protein expression of these putative targets was concordantly downregulated in the presence of miR-145. Luciferase reporter assay further verified direct target association of miR-145 to specific sites of IRS1 and IRS2 3’-untranslated regions. Subsequent analysis also affirmed the modulation of IGF signaling cascade by miR-145 as evident by reduction of the downstream mediator, namely, the active β-catenin level. / The recent advent of deep sequencing has provided an unprecedented platform to study the miRNome, in which both known and novel miRNAs can be identified. Moreover, bioinformatics advances have enabled different types of small ncRNAs, e.g. piwi-interacting RNAs (piRNAs), to be analyzed simultaneously. In the second part of this thesis, small RNA transcriptomes of two HCC cell lines (HKCI-4 and HKCI-8) and an immortalized hepatocyte line (MIHA) were examined using Illumina massively parallel sequencing. Combined bioinformatic and biological analyses revealed the involvement of a novel piRNA, designated as piR-Hep1, in liver tumorigenesis. piR-Hep1 was found to be up-regulated in 47% of HCC in a cohort of 73 HCC patients by qRT-PCR. Silencing of piR-Hep1 inhibited cell viability, motility and invasiveness with a concomitant reduction of Akt phosphorylation. In the analysis of miRNA, miR-1323 was found to be abundantly expressed in HCC and distinctly associated with tumors arising from a cirrhotic background. Furthermore, miR-1323 overexpression in cirrhotic-HCC correlated with poorer disease-free and overall survivals of patients (P<0.009). / Taken together, results from this thesis showed for the first time the pleiotropic effect of miR-145 on targeting multiple components of the oncogenic IGF signaling pathway in HCC. In addition, the functional importance of piR-Hep1 and the prognostic significance of miR-1323 in HCC were highlighted. Studies conducted demonstrated that important miRNAs can be discovered by both traditional array-based profiling and next-generation sequencing. Moreover, comprehensive definition of transcriptome by next-generation sequencing unveils virtually all types of ncRNAs and provides new insight into the liver carcinogenic events. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Law, Tak Yin. / "December 2012." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 180-200). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstracts also in Chinese. / Acknowledgements --- p.i / Publications --- p.ii / Abstract --- p.iii / 摘要 --- p.vi / Contents --- p.viii / List of Figures --- p.xiii / List of Tables --- p.xv / Abbreviations --- p.xvi / Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Hepatocellular Carcinoma - One of the world’s most deadly killers --- p.2 / Chapter 1.2 --- MicroRNAs - a tiny molecule with enormous impacts --- p.10 / Chapter 1.2.1 --- Discovery of miRNAs --- p.11 / Chapter 1.2.2 --- Biogenesis and actions of miRNA --- p.13 / Chapter 1.3 --- MiRNAs and cancer --- p.16 / Chapter 1.4 --- Involvements of miRNAs in HCC etiological factors --- p.18 / Chapter 1.4.1 --- Viral hepatitis infection --- p.19 / Chapter 1.4.2 --- Chronic heavy alcohol consumption --- p.26 / Chapter 1.4.3 --- Dietary aflatoxin exposure --- p.28 / Chapter 1.4.4 --- Male gender --- p.31 / Chapter 1.4.5 --- Obesity --- p.33 / Chapter 1.5 --- Regulation of cancer-associated signaling network by microRNAs --- p.34 / Chapter 1.5.1 --- Apoptotic pathway --- p.37 / Chapter 1.5.1.1 --- Intrinsic pathway --- p.38 / Chapter 1.5.1.2 --- Extrinsic pathway --- p.39 / Chapter 1.5.2 --- Cell cycle regulators --- p.41 / Chapter 1.5.2.1 --- G₁/S transition --- p.42 / Chapter 1.5.2.2 --- G₂/M transition --- p.43 / Chapter 1.5.3 --- Receptor tyrosine kinase-mediated pathways --- p.45 / Chapter 1.5.3.1 --- c-MET-activated signaling --- p.45 / Chapter 1.5.3.2 --- PI3K-Akt --- p.47 / Chapter 1.5.3.3 --- RAS-RAF-MEK-ERK cascade --- p.48 / Chapter 1.5.4 --- TGF-ß signaling pathways --- p.50 / Chapter 1.5.5 --- Metastatic pathways --- p.52 / Chapter 1.5.5.1 --- MiRNAs with metastatic suppressing effects --- p.52 / Chapter 1.5.5.2 --- MiRNAs with metastatic promoting effects --- p.53 / Chapter 1.6 --- Clinical potentials of microRNAs - a killer or a cure? --- p.56 / Chapter 1.6.1 --- MiRNAs involvements in HCC risk prediction --- p.57 / Chapter 1.6.2 --- MiRNAs as diagnostic biomarkers --- p.59 / Chapter 1.6.3 --- MiRNAs as prognostic biomarkers --- p.60 / Chapter 1.6.4 --- Effects of miRNAs on responses to therapy --- p.61 / Chapter 1.7 --- Non-coding RNAs --- p.62 / Chapter 1.8 --- Aims of study --- p.63 / Chapter 2 --- Materials and Methods --- p.65 / Chapter 2.1 --- Quantitative reverse transcription polymerase chain reaction (qRT-PCR) --- p.66 / Chapter 2.1.1 --- Materials --- p.66 / Chapter 2.1.1.1 --- Total RNA extraction --- p.66 / Chapter 2.1.1.2 --- DNase treatment --- p.66 / Chapter 2.1.1.3 --- Reverse transcription --- p.66 / Chapter 2.1.1.4 --- Quantitative polymerase chain reaction --- p.66 / Chapter 2.1.2 --- Methods --- p.67 / Chapter 2.1.2.1 --- Total RNA extraction --- p.67 / Chapter 2.1.2.2 --- DNase treatment --- p.68 / Chapter 2.1.2.3 --- Reverse transcription --- p.69 / Chapter 2.1.2.4 --- Quantitative polymerase chain reaction --- p.69 / Chapter 2.2 --- Transfection --- p.70 / Chapter 2.2.1 --- Materials --- p.70 / Chapter 2.2.2 --- Methods --- p.70 / Chapter 2.2.2.1 --- Evaluation of HCC cells transfection efficiency --- p.70 / Chapter 2.2.2.2 --- Transfection --- p.71 / Chapter 2.3 --- In vitro functional assay --- p.72 / Chapter 2.3.1 --- Materials --- p.72 / Chapter 2.3.1.1 --- Cell viability assay --- p.72 / Chapter 2.3.1.2 --- Colony formation assay --- p.72 / Chapter 2.3.1.3 --- Cell cycle analysis --- p.72 / Chapter 2.3.1.4 --- Apoptosis assay --- p.72 / Chapter 2.3.1.5 --- Cell motility and invasion assay --- p.73 / Chapter 2.3.2 --- Methods --- p.73 / Chapter 2.3.2.1 --- Cell viability assay --- p.73 / Chapter 2.3.2.2 --- Colony formation assay --- p.74 / Chapter 2.3.2.3 --- Cell cycle analysis --- p.75 / Chapter 2.3.2.4 --- Apoptosis assay --- p.75 / Chapter 2.3.2.5 --- Cell motility and invasion assay --- p.76 / Chapter 2.4 --- Luciferase reporter assay --- p.78 / Chapter 2.4.1 --- Materials --- p.78 / Chapter 2.4.1.1 --- Cloning --- p.78 / Chapter 2.4.1.2 --- Cycle sequencing --- p.78 / Chapter 2.4.1.3 --- Luciferase reporter assay --- p.79 / Chapter 2.4.2 --- Methods --- p.79 / Chapter 2.4.1.1 --- Cloning --- p.79 / Chapter 2.4.2.2 --- Cycle sequencing --- p.81 / Chapter 2.4.2.3 --- Luciferase reporter assay --- p.82 / Chapter 2.5 --- Western blot --- p.84 / Chapter 2.5.1 --- Materials --- p.84 / Chapter 2.5.2 --- Methods --- p.85 / Chapter 2.5.2.1 --- Cell harvesting and protein quantitation --- p.86 / Chapter 2.5.2.2 --- Western blotting --- p.86 / Chapter 2.6 --- Small RNA Sequencing --- p.88 / Chapter 2.6.1 --- Materials --- p.88 / Chapter 2.6.2 --- Methods --- p.88 / Chapter 2.6.2.1 --- Sample preparation --- p.88 / Chapter 2.6.2.2 --- Cluster generation by bridge amplification --- p.88 / Chapter 2.6.2.3 --- Sequencing by synthesis --- p.89 / Chapter 2.7 --- Northern blot analysis --- p.94 / Chapter 2.7.1 --- Materials --- p.94 / Chapter 2.7.2 --- Methods --- p.94 / Chapter 2.7.2.1 --- Polyacrylamide gel electrophoresis (PAGE) --- p.94 / Chapter 2.7.2.2 --- Probe preparation --- p.95 / Chapter 2.7.2.3 --- Hybridization, stringency washes and signal detection --- p.95 / Chapter 3 --- Conventional miRNA profiling reveals miR-145 as a tumor suppressor in HCC --- p.97 / Chapter 3.1 --- Introduction --- p.98 / Chapter 3.2 --- Materials and Methods --- p.102 / Chapter 3.2.1 --- Patients --- p.102 / Chapter 3.2.2 --- qRT-PCR --- p.104 / Chapter 3.2.3 --- Cell line --- p.105 / Chapter 3.2.4 --- Transfection --- p.106 / Chapter 3.2.5 --- In vitro functional assay --- p.107 / Chapter 3.2.5.1 --- Cell viability assay --- p.107 / Chapter 3.2.5.2 --- Colony formation assay --- p.107 / Chapter 3.2.5.3 --- Flow cytometry assay --- p.107 / Chapter 3.2.6 --- miRNA target prediction --- p.109 / Chapter 3.2.7 --- Luciferase reporter assay --- p.110 / Chapter 3.2.8 --- Western blot --- p.112 / Chapter 3.2.9 --- Immunohistochemistry --- p.113 / Chapter 3.2.10 --- Statistical analysis --- p.114 / Chapter 3.3 --- Results --- p.115 / Chapter 3.3.1 --- Down-regulation of miR-145 in primary HCC --- p.115 / Chapter 3.3.2 --- Re-expression of miR-145 induced G₂-M arrest and apoptosis --- p.119 / Chapter 3.3.3 --- IRS1, IRS2 and IGF1R expressions --- p.124 / Chapter 3.3.4 --- miR-145 targeted both IRS1 and IRS2 and elicited IGF signaling --- p.126 / Chapter 3.4 --- Discussion --- p.131 / Chapter 4 --- Small RNA Deep sequencing reveals novel non-coding RNAs in HCC --- p.134 / Chapter 4.1 --- Introduction --- p.135 / Chapter 4.2 --- Materials and Methods --- p.136 / Chapter 4.2.1 --- Cell lines --- p.136 / Chapter 4.2.2 --- Patients --- p.137 / Chapter 4.2.3 --- Small RNA Sequencing --- p.139 / Chapter 4.2.4 --- Bioinformatics analysis --- p.140 / Chapter 4.2.4.1 --- Sequence mapping and ncRNA identification --- p.140 / Chapter 4.2.4.2 --- Putative miRNA prediction --- p.140 / Chapter 4.2.4.3 --- Putative piRNA identification --- p.140 / Chapter 4.2.4.4 --- Differentially-expressed ncRNAs identification --- p.141 / Chapter 4.2.5 --- qRT-PCR --- p.142 / Chapter 4.2.6 --- Northern blot analysis --- p.143 / Chapter 4.2.7 --- Transfection --- p.144 / Chapter 4.2.8 --- In vitro functional assays --- p.145 / Chapter 4.2.8.1 --- Cell viability assay --- p.145 / Chapter 4.2.8.2 --- Cell motility and invasion assay --- p.145 / Chapter 4.2.9 --- Western blot analysis --- p.147 / Chapter 4.2.10 --- Statistical analysis --- p.148 / Chapter 4.3 --- Results --- p.149 / Chapter 4.3.1 --- Small RNA Sequencing --- p.149 / Chapter 4.3.2 --- Up-regulation of putative piR-Hep1 in HCC --- p.155 / Chapter 4.3.3 --- piR-Hep1 silencing reduced cell viability and invasiveness --- p.159 / Chapter 4.3.4 --- Novel miR-1323 overexpression in HCC --- p.162 / Chapter 4.4 --- Discussion --- p.171 / Chapter 5 --- Concluding remarks and future perspectives --- p.175 / Chapter 6 --- References --- p.179

大腸癌是全世界最常見的癌症之一，亦是一個癌症死亡率的首要原因。大腸癌患者約50%在病程中會出現轉移病灶。近十年來，雖然多種被批准用於臨床治療的新化療藥顯著提高了大腸癌的治療效果，但是轉移性大腸癌病人的預後仍然很差。隨著各種分子生物技術的進步，新的治療標靶可能在大腸癌細胞株中被發現，並得以在病人標本中驗證。 / 在本研究中，我們採用即時定量多聚酶鏈反應(qPCR)陣列分析，比較大腸癌細胞株和正常大腸細胞株基因表達譜，試圖識別潛在的新的治療標靶。結果提示，與正常大腸細胞株 CCD-18Co 比較，Jagged 2 (JAG2) 和 Frizzled-3 (FZD3)基因 在大腸癌細胞株 SW480 和 SW620 中表達升高。病人大腸癌組織的免疫組織化學染色 (IS) 檢查進一步證實了上述結果，大腸癌組織較其癌旁正常組織表達3.1倍JAG2和6.6倍FZD3蛋白。因此， 我們假設JAG2和FZD3在大腸癌的發生中起重要作用。 / 為了檢驗該假設的真偽，我們運用RNA 干擾的方法進行功能缺失研究。通過該方法，大腸癌細胞株中JAG2 信使RNA和蛋白均能夠被下調，但是FZD3蛋白卻沒有顯示降低。為了弄清JAG2基因的功能，我們進行了單層細胞劃痕傷口癒合試驗和Matrigel 侵襲試驗。結果提示，JAG2 基因下調顯著抑制大腸癌細胞遷移和侵襲的能力。 / 為了調查參與上述功能的機制，我們利用腫瘤轉移相關基因的qPCR陣列分析，試圖檢測出JAG2基因敲除後上調或下調表達的轉移相關基因。結果顯示組織蛋白酶K (CTSK)，一種溶酶體半胱氨酸蛋白酶，在JAG2基因沉默的大腸癌細胞株中表達下調。為了闡明CTSK 活性在大腸癌細胞株侵襲能力中起到的作用，我們採用CTSK抑制劑處理大腸癌細胞株HCT116和DLD-1，發現這兩種細胞株的侵襲能力分別下降了36%和59%。總之, 這些發現表明CTSK可能是JAG2的下游靶基因，活性CTSK可能參與了JAG2介導的大腸癌細胞株侵襲能力。 / 以前的研究表明p38 MAPK通路參與癌細胞遷和侵襲能力的調控。通過Western blot方法，磷酸化的p38和磷酸化的STAT3被發現在JAG2基因沉默的大腸癌細胞中表達降低。p38抑制劑處理的 HCT116和DLD-1細胞降低了侵襲能力下降，同時遷移能力也由於p38抑制劑的處理而降低，支持p38可調控癌細胞遷移和侵襲能力的事實。 / 總之，我們的結果顯示JAG2高表達通過啟動CTSK和p38 MAPK通路，可能促進大腸癌轉移。因此，JAG2可能成為轉移性大腸癌治療的潛在標靶。 / Colorectal cancer (CRC) is one of the most frequent cancers worldwide and is a leading cause of cancer mortality. Around 50% of patients with CRC will experience metastases. Although significant progress has been made in CRC treatment within the last decade with the approval of multiple new chemotherapeutic agents, the prognosis for patients with metastatic CRC remains poor. With the advancement of molecular techniques, novel therapeutic targets are able to be discovered in CRC cell lines and validated in patient samples. / Therefore in this project, I aim to identify potential novel therapeutic targets by comparing the gene expression profile of colon cancer cell lines and a normal colon cell line using quantitative polymerase chain reaction (qPCR) arrays. Results showed that Jagged 2 (JAG2) and Frizzled-3 (FZD3) were up-regulated in the CRC cell lines SW480 and SW620 as compared to the normal colon cell line CCD-18Co. Those results were further validated by immunohistochemical staining (IS), which detected up-regulated JAG2 (3.1-fold) and FZD3 (6.6-fold) proteins expression in CRC tissues as compared to adjacent normal tissues. Thus I hypothesized that JAG2 and FZD3 may play an important role in CRC carcinogenesis. / In order to study the roles of FZD3 and JAG2 in CRC, loss-of-function studies by RNA interference (RNAi) were carried out. While the expression of FZD3 protein failed to be down-regulated by RNAi, JAG2 expression was successfully knocked down in CRC cell lines at both the mRNA and protein levels. Functional analyses using the monolayer scratch wound-healing assay and Matrigel invasion assay showed that JAG2 knockdown significantly inhibited migration and invasion in CRC cell lines. / To investigate the mechanisms involved, a tumour metastasis qPCR array was used to examine the changes in the expression level of metastasis-related genes after JAG2 gene knockdown. Results showed that the expression of Cathepsin K (CTSK), a lysosomal cystein protease, was found to be down-regulated in CRC cell lines following JAG2 silencing. To demonstrate the importance of CTSK activity in CRC cell invasion, HCT116 and DLD-1 CRC cell lines were treated with a CTSK inhibitor and its effect were assessed by the Matrigel invasion assay. Results showed that CTSK inhibition led to a 36% and 59% reduction in number of invaded cells in HCT116 and DLD-1 cell lines, respectively. Taken together, these findings show that CTSK may be a downstream target of JAG2 and that active CTSK may involve in JAG2 mediated invasion in CRC cell lines. / Previous works by others have shown that the p38 MAPK pathway is involved in the regulation of migration and invasive activity of cancer cell lines. Using Western blot analysis, the expression of phosphorylated p38 MAPK and phosphorylated STAT3 were found to be down-regulated following JAG2 depletion in CRC cell lines. In support of a role for p38 MAPK in the regulation of cancer cell migration and invasive capability, treatment with a p38 MAPK inhibitor was found to reduce the percentage of invasive cells and distance moved by migratory cells in HCT116 and DLD-1 cell lines. / In conclusion, my results show that JAG2 over-expression in CRC may promote cancer cell migration and invasion through activation of CTSK and the p38 MAPK pathway. Therefore, JAG2 may be a potential therapeutic target for treatment of metastatic CRC. / 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. / He, Wan. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 164-207). / 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 in English --- p.i / Abstract in Chinese --- p.iv / Acknowledgements --- p.vi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Colorectal Cancer (CRC) --- p.1 / Chapter 1.1.1 --- Epidemiology and Incidence --- p.1 / Chapter 1.1.2 --- Histology --- p.2 / Chapter 1.1.3 --- Gender and Age --- p.4 / Chapter 1.1.4 --- Etiology of CRC --- p.4 / Chapter 1.1.4.1 --- Environment --- p.4 / Chapter 1.1.4.2 --- Hereditary Factors --- p.5 / Chapter 1.1.4.3 --- Dietary Factors --- p.6 / Chapter 1.1.4.4 --- Obesity --- p.6 / Chapter 1.1.4.5 --- Tobacco and alcoho --- p.7 / Chapter 1.1.4.6 --- Inflammatory bowel disease (IBC) --- p.7 / Chapter 1.1.5 --- Genetic Changes in CRC --- p.8 / Chapter 1.1.5.1 --- Chromosomal Aberration --- p.8 / Chapter 1.1.5.2 --- Tumor Suppressor Genes --- p.10 / Chapter 1.1.5.2.1 --- APC gene --- p.10 / Chapter 1.1.5.2.2 --- P53 gene --- p.11 / Chapter 1.1.5.2.3 --- SMAD4 gene --- p.11 / Chapter 1.1.5.3 --- Oncogenes --- p.12 / Chapter 1.1.5.3.1 --- Epidermal Growth Factor Receptor (EGFR) gene --- p.12 / Chapter 1.1.5.3.2 --- RAS gene and BRAF gene --- p.13 / Chapter 1.1.5.4 --- Proposed Two-hit Model for the Multistep Pathogenesis of CRC --- p.15 / Chapter 1.1.6 --- Clinical Presentation and Diagnosis --- p.16 / Chapter 1.1.7 --- Theatment --- p.16 / Chapter 1.1.7.1 --- Surgery --- p.16 / Chapter 1.1.7.2 --- Radiotherapy (RT) --- p.17 / Chapter 1.1.7.3 --- Concurrent Chemotherapy --- p.17 / Chapter 1.1.7.4 --- Target Therapy --- p.18 / Chapter 1.1.7.5 --- Colorectal Cancer Treatment by Stage --- p.19 / Chapter 1.1.7.6 --- Novel Strategies --- p.20 / Chapter 1.1.7.6.1 --- Epigenetic therapy --- p.20 / Chapter 1.1.7.6.2 --- Immunotherapy --- p.21 / Chapter 1.2 --- Pathways Involved in CRC Carcinogenesisand Progression --- p.22 / Chapter 1.2.1 --- Wnt Signaling Pathway --- p.22 / Chapter 1.2.2 --- Notch Signaling --- p.23 / Chapter 1.2.3 --- Nuclear Factor-kappa B (NF-κB) Signaling Pathway --- p.23 / Chapter 1.2.4 --- Phosphatidylinositol 3-kinase (PI3K) Signaling Pathway --- p.24 / Chapter 1.2.5 --- Crosstalk Among WNT, NOTCH, NF-κB and PI3K Signaling Pathway in CRC --- p.24 / Chapter 1.3 --- Hypothesis and Objectives of this Study --- p.28 / Chapter Chapter 2 --- Identification of Differentially Expressed Genes between Colorectal Cancer Cell Lines and A Normal Colon Cell Line --- p.29 / Chapter 2.1 --- Background --- p.29 / Chapter 2.2 --- Materials and Methods --- p.33 / Chapter 2.2.1 --- Cell Lines --- p.33 / Chapter 2.2.2 --- Identification of Differetially Expressed Genes by qPCR Arrays --- p.33 / Chapter 2.2.2.1 --- Total RNA Extraction --- p.33 / Chapter 2.2.2.2 --- RNA Quality Contol --- p.34 / Chapter 2.2.2.3 --- Reverse Transcription (RT) --- p.34 / Chapter 2.2.2.4 --- PCR Arrays --- p.34 / Chapter 2.3 --- Results --- p.36 / Chapter 2.3.1 --- Differentially Expressed Genes in WNT Signaling Pathway --- p.36 / Chapter 2.3.2 --- Differentially Expressed Genes in Notch Signaling Pathway --- p.40 / Chapter 2.3.3 --- Differentially Expressed Genes in NF-κB Signaling Pathway --- p.43 / Chapter 2.3.4 --- Differentially Expressed Genes in PI3K-AKT Signaling Pathway --- p.46 / Chapter 2.3.5 --- Choice of over-expressed genes for further validation and characterization --- p.49 / Chapter 2.4 --- Discussions --- p.53 / Chapter 2.4.1 --- WNT Signaling Pathway --- p.53 / Chapter 2.4.2 --- NOTCH Signaling Pathway --- p.54 / Chapter 2.4.3 --- NF-κB Signaling Pathway --- p.55 / Chapter 2.4.4 --- PI3K-AKT Signaling Pathway --- p.56 / Chapter 2.4.5 --- Choice of over-expressed genes for further validation and characterization --- p.56 / Chapter Chapter 3 --- JAG2, FZD3 and NOTCH4 Expression in Colorectal Cancer Cell Lines and Colorectal Cancer Tissues --- p.59 / Chapter 3.1 --- Background --- p.59 / Chapter 3.1.1 --- JAG2 Ligand --- p.59 / Chapter 3.1.2 --- FZD3 Receptor --- p.61 / Chapter 3.1.3 --- NOTCH4 Receptor --- p.62 / Chapter 3.2 --- Materials and Methods --- p.64 / Chapter 3.2.1 --- CRC Cell Lines --- p.65 / Chapter 3.2.2 --- CRC Tissues --- p.65 / Chapter 3.2.3 --- Quantitative RT-PCR --- p.66 / Chapter 3.2.4 --- Detection of JAG2, FZD3 and NOTCH4 Protein Expression in CRC Tissues by Immunohistochemical Staining (IS) --- p.67 / Chapter 3.2.5 --- Western Blot Assays --- p.68 / Chapter 3.2.5.1 --- Protein extraction --- p.68 / Chapter 3.2.5.2 --- SDS-PAGE gel electrophroresis --- p.68 / Chapter 3.2.5.3 --- Protein blotting --- p.68 / Chapter 3.2.6 --- Detection of JAG2 and FZD3 Protein Expression in CRC and Normal Colon Cell Lines by Western Blotting --- p.69 / Chapter 3.2.7 --- Statistical Analysis --- p.70 / Chapter 3.3 --- Results --- p.71 / Chapter 3.3.1 --- JAG2 and FZD3 but not NOTCH4 mRNA were Over -expressed in CRC Cell Lines --- p.71 / Chapter 3.3.2 --- Over-expression of JAG2 and FZD3 Proteins in CRC Tissues --- p.72 / Chapter 3.3.3 --- FZD3 Over-expression Correlated with Tumour-Node Metastasis (TNM) stages --- p.76 / Chapter 3.3.4 --- JAG2 and FZD3 Protein Expression in Colorectal Cancer and Normal Cell Lines --- p.77 / Chapter 3.4 --- Discussions --- p.78 / Chapter Chapter 4 --- Functional Analyses of JAG2 and FZD3 in CRC Cell Lines by RNA Interference --- p.81 / Chapter 4.1 --- Background --- p.81 / Chapter 4.2 --- Materials and Methods --- p.84 / Chapter 4.2.1 --- Transfection of siRNA into CRC Cell Lines --- p.84 / Chapter 4.2.2 --- Cell Proliferation Assay --- p.85 / Chapter 4.2.3 --- Monolayer Scratch Wound Healing Assay --- p.85 / Chapter 4.2.4 --- Matrigel Invasion Assay --- p.86 / Chapter 4.2.5 --- Statistical Analysis --- p.87 / Chapter 4.3 --- Results --- p.88 / Chapter 4.3.1 --- Knockdown of JAG2 and FZD3 Expression by RNA Interference --- p.88 / Chapter 4.3.2 --- Effect of JAG2 Knockdown on Migration of CRC Cell Lines --- p.91 / Chapter 4.3.3 --- JAG2 Knockdown by siRNA 2 Transfection Reduced Migratory Capability of HCT116, DLD-1and HT29 cell lines --- p.94 / Chapter 4.3.4 --- JAG2 Knockdown Impaired the Invasiveness of HCT116 and DLD-1 Cell Lines --- p.97 / Chapter 4.3.5 --- Decreased Migratory and Invasive Capabilities Induced by JAG2 Knockdown was not Due to Reduced Cell Proliferation --- p.100 / Chapter 4.4 --- Discussions --- p.102 / Chapter Chapter 5 --- NOTCH Pathway Inactivation by JAG2 Silencing Reduces Oncogenic Properties of HT29 but not HCT116 andDLD-1 CRC Cell Lines --- p.106 / Chapter 5.1 --- Background --- p.106 / Chapter 5.2 --- Materials and Methods --- p.109 / Chapter 5.2.1 --- CRC Cell lines --- p.109 / Chapter 5.2.2 --- Pharmacological Inhibition of NOTCH signaling by DAPT --- p.109 / Chapter 5.2.3 --- Combination of DAPT Treatment and JAG2 Silencing by siRNA --- p.109 / Chapter 5.2.4 --- Western Blotting --- p.109 / Chapter 5.2.5 --- Cell Proliferation Assay (MTS Assay) --- p.110 / Chapter 5.2.6 --- Monolayer Scratch Wound Healing Assay --- p.110 / Chapter 5.2.7 --- Matrigel Invasion Assay --- p.111 / Chapter 5.2.8 --- Statistical Analysis --- p.111 / Chapter 5.3 --- Results --- p.112 / Chapter 5.3.1 --- JAG2 Silencing Down-regulates Notch Pathway Signaling in CRC Cell Lines --- p.112 / Chapter 5.3.2 --- Inhibition of NOTCH Signaling by DAPT Treatment in CRC Cell Lines --- p.112 / Chapter 5.3.3 --- NOTCH Inhibition Does not Significantly Affect Cell Proliferation in CRC Cell Lines --- p.114 / Chapter 5.3.4 --- Suppression of NOTCH Signaling by DAPT Inhibits Migration in HT29 but not in HCT116 and DLD-1 CRC Cell Lines --- p.115 / Chapter 5.3.5 --- Suppression of NOTCH Signaling by DAPT does not Significantly Affect Invasiveness of HCT116 and DLD-1 CRC Cell Lines --- p.117 / Chapter 5.4 --- Discussions --- p.118 / Chapter Chapter 6 --- JAG2 Knockdown Inhibits Invasion in CRC Cell Lines through Inactivation of Cathepsin K --- p.121 / Chapter 6.1 --- Background --- p.121 / Chapter 6.2 --- Materials and Methods --- p.123 / Chapter 6.2.1 --- Human Tumour Metastasis RT2 Profiler[superscript TM] PCR Array --- p.123 / Chapter 6.2.2 --- Measurement of CTSK Gene expression level by Quantitative Real-Time PCR --- p.123 / Chapter 6.2.3 --- Immunohistochemical Staining (IS) of CTSK in CRC Tissues --- p.124 / Chapter 6.2.4 --- Pharmacological Inhibitior of CTSK in CRC Cell Lines --- p.124 / Chapter 6.2.5 --- Inhibition of CTSK in CRC Cell Lines for Migration Study --- p.124 / Chapter 6.2.6 --- Inhibition of CTSK in CRC Cell Lines for Invasion Study --- p.125 / Chapter 6.2.7 --- Western Blotting --- p.125 / Chapter 6.2.8 --- Statistical Analysis --- p.125 / Chapter 6.3 --- Results --- p.126 / Chapter 6.3.1 --- Identification of Metastasis Related Genes Which were Down-regulated by JAG2 Knockdown in HCT116 Cells --- p.126 / Chapter 6.3.2 --- Validation of Down-regulation of CTSK Gene by JAG2 Knockdown in HCT116 Cell Line by qRT-PCR --- p.126 / Chapter 6.3.3 --- JAG2 Knockdown Reduced Expression of Active CTSK Protein in CRC Cell Lines --- p.128 / Chapter 6.3.4 --- CTSK Protein Expression in CRC Tissue Samples --- p.130 / Chapter 6.3.5 --- Pharmacological Inhibition of CTSK Suppressed Invasiveness of CRC Cell Lines --- p.131 / Chapter 6.3.6 --- Pharmacological Inhibition of CTSK did not Affect Migration of CRC Cell Lines --- p.132 / Chapter 6.4 --- Discussions --- p.133 / Chapter Chapter 7 --- Depletion of JAG2 Inhibits Migration and Invasion in CRC Cell Lines through Inactivation of p38 MAPK/HSP27 Pathway --- p.137 / Chapter 7.1 --- Background --- p.137 / Chapter 7.2 --- Materials and Methods --- p.140 / Chapter 7.2.1 --- Pharmocological Inhibition of p38 MAPK Phosphorylation CRC Cell Lines --- p.140 / Chapter 7.2.2 --- Inhibition of p38 MAPK Phosphorylation for Migration Study in CRC Cell Lines --- p.140 / Chapter 7.2.3 --- Inhibition of p38 MAPK Phosphorylation for Invasion Study in CRC Cell Lines --- p.140 / Chapter 7.2.4 --- Knockdown of STAT3 by RNA interference --- p.141 / Chapter 7.2.5 --- Knockdown of STAT3 for Migration Study in CRC Cell Lines --- p.141 / Chapter 7.2.6 --- Knockdown of STAT3 for Invasion Study in CRC Cell Lines --- p.141 / Chapter 7.2.7 --- Western Blotting --- p.141 / Chapter 7.2.8 --- Statistical Analysis --- p.142 / Chapter 7.3 --- Results --- p.143 / Chapter 7.3.1 --- JAG2 Knockdown Inhibits p38 MAPK / HSP27 Pathway in CRC Cell Lines --- p.143 / Chapter 7.3.2 --- Inhibition of p38 MAPK / HSP27 Signaling Pathway Down-regulated Invasive Capability of CRC Cell Line --- p.145 / Chapter 7.3.3 --- Inhibition of p38 MAPK / HSP27 Signaling Pathway Down-regulated Migration of CRC Cell lines --- p.147 / Chapter 7.3.4 --- JAG2 Knockdown Inactivated p38 MAPK / HSP27 Pathway Independently of NOTCH Pathway in CRC Cell Lines --- p.149 / Chapter 7.3.5 --- JAG2 Knockdown Inhibits STAT3 Activation in CRC Cell Lines --- p.151 / Chapter 7.3.6 --- STAT3 Silencing Reduced Invasive Capability in CRC Cell Lines --- p.152 / Chapter 7.3.7 --- STAT3 Silencing Reduced Migratory Capability in CRC Cell Lines --- p.154 / Chapter 7.3.8 --- Inhibition of p38 MAPK Activity Suppressed STAT3 Activation in HCT116 Cells --- p.156 / Chapter 7.4 --- Discussions --- p.157 / Chapter Chapter 8 --- Conclusions and Future Works --- p.161 / Chapter 8.1 --- Conclusions --- p.161 / Chapter 8.2 --- Future work --- p.163 / References --- p.164 / Chapter Appendix 1 --- List of Figures and Tables --- p.208 / Chapter Appendix 2 --- Abbrevations used in this thesis --- p.212

Dong Shumin. / "February 2002." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (p. 187-220). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Yun Jing-ping. / "March 2002." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (p. 185-226). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Wang Ya-Ping. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (p. 152-161). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.