Colorectal cancer is a serious health problem that has concerned people for decades. In Hong Kong, it is the most common cancer and the second leading cause of death. Among colorectal cancer patients, around 40-50% of them will develop metastatic disease. Chemotherapy is playing an important role all the time in the treatment of advanced colorectal cancer. In the past decade, the application of targeted therapies in treatment has largely improved efficacy and prolonged survival. Bevacizumab and cetuximab are two commonly used targeted agents in daily clinical practice of Hong Kong. Since multiple clinical trials have studied bevacizumab and cetuximab in combination with other chemotherapies, limited data is available in Asian patients. Therefore, we conduct three 5-year retrospective analyses based on patients received treatment in Hong Kong Queen Mary Hospital, to investigate the clinical efficacy and toxicity of those two drugs. The first study examined the use of bevacizumab in treating KRAS mutated type patients. We found the efficacy and results were consistent with historical data. In the next analysis of cetuximab, comparable data were shown which suggested the consistency with previous studies. The last study is aim to compare bevacizumab and cetuximab in previously untreated wild-type KRAS patients. Identical response rates, progression-free survival and overall survival were finally reported. / published_or_final_version / Medicine / Master / Master of Philosophy

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Lectins (hemagglutinins) are a type of proteins that could recognize different sugar structures and specifically initiate reversible binding with them. Though they have been universally found in a variety of organisms, they are exceptionally abundant in legumes. From the initial finding of agglutinating red blood cells to the discovery of recognizing carbohydrates on cell membranes, multiple functions of lectins have been gradually unveiled by numerous researchers across a century. Based on its carbohydrate-binding property, lectins have found great value in the study of glycomics. Many lectin-based biological tools, like lectin affinity chromatography, lectin blotting, lectin histochemistry, lectin microarray and lectin-based biosensor have been developed and applied to the study of glycoproteins. Besides, lectins are also reported to be potential agents for anti-insect, anti-fungi, anti-HIV, anti-bacterial and anti-tumor applications. / The present study focuses on the isolation of a new hemagglutinin from an edible legume, exploration of its anti-colorectal cancer effect and mechanisms, its cytokine inducing function and anti-HIV activities. The protein was purified by liquid chromatography techniques which entailed affinity chromatography on Affi-Gel Blue Gel, ion exchange chromatography on Mono Q and gel filtration on Superdex 75 with an FPLC system. The hemagglutinating activity of this hemagglutinin was demonstrated to be ion-dependent and stable over a wide range of temperatures (20-60℃) and pH (2-11) values. Like most of the lectins or hemagglutinins, this novel hemagglutinin could also attenuate the activity of HIV-1 reverse transcriptase. / This hemagglutinin could potently suppress the proliferation of colorectal carcinoma HCT116 cells and colorectal adenocarcinoma HT29 cells. It induced cell cycle arrest in G0/G1 phase, downregulated the expression of Cyclin D1 and upregulated P21expression. The protein initially bound on the cell membranes most probably through glycoproteins and subsequently entered the cytoplasm, which was achieved as early as 3h post treatment. The hemagglutinin was found to be preferentially localized in Golgi apparatus and initiated aggregation of the Golgi apparatus, which may possibly attenuate its protein processing capacity by reducing total superficial area or even partially blocking the transportation of proteins from the endoplasmic reticulum (ER). The impaired protein reception ability of Golgi apparatus may lead to the protein accumulation in the ER and induce cell apoptosis. Accordingly, two ER stress sensors (IRE1α and ATF6) and one late product of ER stress (CHOP) were found to up-regulated. Apoptosis-inducing effect of this hemagglutinin on HT29 and HCT116 cells were further confirmed using methods based on different principles. Cells treated with the hemagglutinin were observed to undergo obvious chromatin condensation, mitochondrial membrane depolarization and phosphatidylserine exposure. An apoptosis initiator (Apaf-1) and one important indicator (cleaved PARP) of cell apoptosis were accordingly detected. Besides, intraperitoneal administration of this hemagglutinin to colorectal tumor bearing nude mice could slow down the growth of tumors. / At last, this hemagglutinin exerted an immunomodulatory function on splenocytes by stimulating the mRNA expression level of interleukin-2 (IL-2), interleukin-6 (IL-6), interleukin-1 beta (IL-1β), interferon- gamma (IFN-γ), and tumor necrosis factor alpha (TNF-α). Secretion of IL-1β and IL-2 from splenocytes also increased with the concentration of this hemagglutinin. / In a short conclusion, we have isolated a new hemagglutinin with anti-HIV RT, anti-colorectal cancer and immunomodulatory activities. / 凝集素（血凝素）是一类能够识别不同糖结构并能和它们发生可逆性结合的蛋白。虽然他们在许多生物体内均有发现，但这类蛋白在豆科植物中的含量尤其丰富。经过一个多世纪来众多研究者的努力，从最初认识到其具有红血细胞凝集功能到糖类识别作用，凝集素的诸多功能已被逐步挖掘。基于其独特的糖结构识别特性，凝集素在糖组学的研究中具有重大意义。许多基于凝集素的生物方法，如凝集素亲和层析法，凝集素印迹法，凝集素组织化学，凝集素生物芯片以及基于凝集素的生物传感器已被研究出来, 并用于研究糖蛋白。除此之外，研究表明，凝集素还具有抗虫，抗真菌，抗HIV，抗细菌和抗癌等活性。 / 该凝集素可以极大抑制结肠直肠癌HCT116细胞和结直肠腺癌HT29细胞增殖，引发细胞周期停滞，分别下调和上调Cyclin D1和P21的表达。该蛋白极有可能首先通过和细胞表面的糖蛋白结合而附在细胞膜上，然后进入细胞内。该过程可在往细胞培养液内加入该蛋白后的3小时内完成。该凝集素优先与细胞内的高尔基体结合，随后引发高尔基体聚集。该聚集作用可能会通过减少高尔基体总表面积甚至阻塞内质网和高尔基体间的蛋白运输，进而减弱高尔基体处理蛋白质的能力。当高尔基体接受蛋白的能力降低时，蛋白可能会堆积在内质网上并进一步引发细胞程序性死亡。相应地，两个内质网应激感受蛋白IRE1α和 ATF6以及内质网应激后期产物CHOP均被发现上调。该凝集素对HT29细胞和HCT116细胞的凋亡诱导作用采用不同的方法进行了进一步的确认，这些方法都是基于不同检测原理进行的。结果表明，该凝集素可导致细胞产生明显的染色质凝缩，线粒体膜电位去极化和磷脂酰丝氨酸外翻。与此相应地，凋亡启动蛋白Apaf-1和凋亡后期蛋白(被剪切的PARP)可在处理后的细胞中检测到。通过腹腔注射的方法给接种大肠癌细胞的裸鼠给药可降低肿瘤的生长速度。 / 本研究的工作包括：从一种可食用豆类中提取一种新的凝集素；检测其抗大肠癌的作用和机制；研究其细胞素诱导作用以及抗HIV活性。该蛋白采用液相色谱法分离提纯，其中包括亲和层析柱Affi-Gel Blue Gel, 离子交换层析柱Mono Q 和凝胶层析柱Superdex 75，后两种层析法在FPLC系统上操作。该蛋白的红血细胞凝集作用具有金属阳离子依赖性，并在20-60℃和pH2-11范围内保持活性稳定。像许多其它的凝集素一样，该蛋白也可以削弱HIV逆转录酶活性。 / 最后，该蛋白还具有免疫调节作用，它可促进白细胞介素-2，白细胞介素-6，白细胞介素-1β，干扰素-γ和肿瘤坏死因数-α在mRNA水平上的表达并刺激白细胞介素-2和细胞介素-1β的分泌。 / 综上所诉，本研究分离提纯了一种新凝集素，它具有抗HIV，抗大肠癌和免疫调节作用。 / Dan, Xiuli. / Thesis Ph.D. Chinese University of Hong Kong 2015. / Includes bibliographical references (leaves 153-170). / Abstracts also in Chinese. / Title from PDF title page (viewed on 05, October, 2016). / 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.

TCP-1是一种新型的定向于肿瘤血管的多肽，通过小鼠体内的噬菌体展示技术筛选得到。在之前的研究中，我们已证明TCP-1具有定向于肿瘤血管并有效靶向运输抗肿瘤药物和显像剂的特性。本研究的目的是进一步研究在原位结肠癌模型中定向运输抗肿瘤药物肿瘤坏死因子(TNFα)，以及在结肠癌临床样本中运输显像剂异硫氰酸荧光素(FITC)的能力。并对TCP-1与肿瘤坏死因子的融合蛋白TCP-1/TNFα的抗肿瘤机制进行阐述。 / 本研究中，我们首先尝试用TCP-1作为载体，将增强绿色荧光蛋白靶向运输至肿瘤血管。结果证明TCP-1可以成功将蛋白运输到在肿瘤血管而非其它正常的组织器官上。TCP-1还可以靶向运输肿瘤坏死因子并增强其抗肿瘤作用。和肿瘤坏死因子比较，融合蛋白TCP-1/TNFα处理组的凋亡细胞数量增多，肿瘤微血管数目减少，并且无明显毒副作用。与结肠癌的一线化疗药物5-氟尿嘧啶(5-FU)联合给药后，与TNFα与5-FU联合给药相比较，融合蛋白TCP-1/TNFα联合5-FU在以下方面具有更明显的作用：抑制肿瘤生长，增加肿瘤细胞凋亡和抑制肿瘤细胞增殖，促进肿瘤血管正常化，升高瘤内免疫细胞以及减轻骨髓和脾内的免疫抑制反应。经检测TCP-1的靶向运输增加了瘤内的TNFα以及5-FU的浓度。这些都表明TCP-1不但可以靶向运输TCP-1/TNFα至肿瘤血管，还可以增加CD8+细胞的浸润增加瘤内免疫反应以及增加血管对抗肿瘤药物的通透性。以上都对抗肿瘤起到重要作用。 / 在临床的结肠癌样本中，TCP-1对肿瘤血管的结合能力也得到了证实。48.98%的结肠癌样本对TCP-1的结合为阳性。统计学分析显示TCP-1的结合与结肠癌的分期和肿瘤位置有关，对于N2期，位于乙状结肠的肿瘤的结合尤为明显。本研究的主要目的是将分离鉴定出的TCP-1发展成为结肠癌的生物标记，并且作为运输抗肿瘤药物和显像剂的载体应用于结肠癌的诊断和治疗中。鉴于TCP-1的靶向运输特点，将会有机会研发更多的抗肿瘤药物，同时增强传统化疗药的抗肿瘤作用。这些都可以优化肿瘤治疗的方案。综上所述，TCP-1是一种在结肠癌治疗诊断中具有广阔前景的多肽。 / TCP-1 is a novel vasculature-targeting peptide. It was discovered through the in vivo phage library selection in mice. It was demonstrated that TCP-1 peptide exhibited a homing ability to the neovasculature of colon tumors and was capable of efficiently delivering imaging agents and chemotherapeutic drugs to this target site. The current study is to further investigate the targeting ability of TCP-1 to deliver a known immunomodulator, tumor necrosis factor α (TNFα) as an example of anti-cancer drug in an orthotopic colorectal cancer (CRC) model and fluorescein isothiocyanate (FITC) as imaging agent for testing the binding capacity for tumors in colorectal cancer patients. The mechanisms for the action of this novel biologic TCP-1/TNFα in the treatment of colon cancer in mice were also defined. / In this study, we observed that TCP-1 peptide delivered enhanced green fluorescent protein (EGFP) only to tumor blood vessel other than normal organs after TCP- 1/EGFP injection. This was not observed after EGFP injection. This finding showed that TCP-1 can deliver biologic protein to the tumor blood vessels. Furthermore, results from TNFα or TCP-1/TNFα targeted delivery experiments showed that TCP- 1/TNFα displayed stronger anti-cancer effects than TNFα alone on the induction of apoptosis and reduction in number of microvessels in the tumors, without significant effect in systemic toxicity. In the combined therapy with 5-fluorouracil (5-FU), a standard drug for colon cancer treatment, pretreatment with low dose (1 ng TNFα /mouse) of TNFα or TCP-1/TNFα potentiated the anti-cancer action of 5-FU. In this regard, TCP-1/TNFα could significantly reduce tumor size and weight, increase number of apoptotic cells, inhibit tumor cell proliferation, normalize tumor blood vessels, facilitate infiltration of immune cells to tumor mass and attenuate immunosuppression in bone marrow and spleen. Moreover, TCP-1 could significantly increase intratumoral levels of TNFα and 5-FU. It was also suggested that TCP-1 could selectively deliver TNFα to the tumor blood vessels and modulate the immune response by increasing CD8+ cells infiltration to tumors and increase vascular permeability to 5-FU. These observations may be the key actions to reduce tumor growth. / The binding ability of TCP-1 was also detected in clinical samples from colorectal cancer patients in which 24/49 (48.98%) tumor tissues were positive with TCP-1 binding signal. Statistical analysis showed that TCP-1 had a strong and significant binding with colorectal cancer at the N2 stage among the different colorectal cancer stages (P=0.028) and location in the colon at the sigmoid (P<0.001). / Our study also focused on the isolation and identification of the binding molecule of TCP-1 in order to develop it into a biomarker for CRC and using TCP-1 as a carrier in delivering anti-cancer drugs and imaging agents to colon tumors for cancer therapy and diagnosis. With the homing property of TCP-1 on colon tumor blood vessels, new types of anti-cancer drugs will be developed and their combinations with conventional chemotherapy drugs will optimize the therapeutic outcome and improve regimen of treatment for CRC. Taken together, TCP-1 peptide appears to be a promising agent in molecular imaging and drug delivery for CRC diagnosis and therapy. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Lu, Lan. / Thesis (Ph.D.) Chinese University of Hong Kong, 2014. / Includes bibliographical references (leaves 157-177). / Abstracts also in Chinese. / Lu, Lan.

Colorectal cancer (CRC) is one of the most common malignancies worldwide. However, adjuvant chemotherapeutic agents exhibit poor accumulation in the tumor mass and frequently result in serious side effects due to nonspecific damage to normal organs. Therefore, the development of more selective anticancer drugs with targeted delivery to tumor sites is the current trend in cancer therapies. Among these sites, tumor neovasculature is an attractive target for anticancer agents. It is because tumor growth is largely limited by blood supply which is dependent on the extent of angiogenesis in the tumor. / Experimental analysis suggested that TCP-1 phage and synthetic TCP-1 peptide specifically homed to colorectal cancer tissues and co-localized with the tumor vasculature. Moreover, TCP-1 peptide also recognized the vasculature of human colorectal cancer specimens. Subsequently, the homing abilities of TCP-1 phage were extensively tested in other cancer models. Results showed that TCP-1 peptide could also target the vasculature of orthotopic gastric cancer induced by human colon cancer cell line (MKN45) in BALB/c nude mice. Meanwhile, TCP-1 phage exhibited binding activity to colorectal cancer cells such as colon 26 and SW1116. TCP-1 peptide could carry a pro-apoptotic peptide into these cells and markedly enhanced its pro-apoptotic action. / In summary, we have used the phage display technology to isolate two unique peptides TCP-1 and TCP-2, which targeted the vasculature of orthotopic colorectal cancer and also recognized the vasculature of human colorectal cancer. Moreover, they could deliver fluorescein or pro-apoptotic peptide only to the tumor vasculature but not to other normal tissues, for imaging detection and targeted therapy. In conclusion, both TCP-1 and TCP-2 may have significant clinical applications as carriers in diagnostic imaging and ligand-mediated targeted therapy for human colorectal cancer. / Similarly, TCP-2 phage or its peptide also targeted specifically the orthotopic colorectal cancer, and co-localized with the tumor vasculature in mice. Meanwhile, TCP-2 peptide recognized the vasculature of human colorectal cancer specimens. FITC-labeled TCP-2 peptide could also be used to detect cancer tissues in tumor-bearing mice. / To identify specific ligands targeting the tumor neovasculature, in vivo phage display technology has been extensively used. Several dozens of peptides homing to normal or diseased vasculature have been identified through this technology. However, these peptides target mainly the tumors growing at distant sites but not at the primary organ, thus limiting their clinical application. To obtain specific peptides targeting the neovasculature of colorectal cancer growing in situ, we established an orthotopic colorectal cancer model in normal BALB/c mice by using syngeneic colon cancer cells (colon 26). Subsequently, in vivo phage display technology was utilized to isolate peptides which specifically recognized the vasculature of the cancer. Four peptides (termed TCP-1, 2, 3, 4) were enriched more than once after four-round selections. Further investigation disclosed that TCP-1 and TCP-2 phages had relatively stronger binding abilities to cancer tissues among the four phage clones. They were chosen for further study. / We further demonstrated that TCP-1 could serve as a carrier for image detection and drug delivery. FITC-labeled TCP-1 could specifically produce a strong fluorescence signal in the tumors after intravenous injection into the orthotopic tumor-bearing mice. Moreover TCP-1, when conjugated with a pro-apoptotic peptide, could also specifically induce apoptosis of tumor vasculature in vivo. / Li, Zhijie. / Adviser: Cho Chiltin. / Source: Dissertation Abstracts International, Volume: 72-04, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 194-221). / 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. / Abstract also in Chinese.

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.

Wong, Wai Ki Vicky. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 105-131). / Abstracts in English and Chinese. / Abstract --- p.ii / 摘要 --- p.iv / Acknowledgements --- p.vi / Table of contents --- p.vii / List of tables --- p.xii / List of figures --- p.xiii / List of abbreviations --- p.xiv / Chapter Chapter 1: --- Introduction --- p.1 / Chapter Chapter 2: --- Colorectal cancer / Chapter 2.1 --- Literature Review / Chapter 2.1.1 --- Colorectal cancer --- p.8 / Chapter 2.1.2 --- Incident rate of colorectal cancer --- p.8 / Chapter 2.1.3 --- Hereditary colorectal cancer --- p.9 / Chapter 2.1.4 --- Sporadic colorectal cancer and Wnt signaling pathway --- p.10 / Chapter 2.1.5 --- Chemotherapy treatment of colorectal cancer --- p.11 / Chapter 2.1.5.1 --- 5-Fluorouracil --- p.12 / Chapter 2.1.5.2 --- Oxaliplatin --- p.14 / Chapter 2.1.5.3 --- Irinotecan --- p.14 / Chapter 2.1.6 --- Biomarkers for colorectal cancer --- p.15 / Chapter 2.1.6.1 --- Thymidylate synthase --- p.15 / Chapter 2.1.6.2 --- Dihydropyrimidine dehydrogenase --- p.16 / Chapter 2.1.6.3 --- Thymidine phosphorylase --- p.16 / Chapter 2.1.6.4 --- Microsatellite-instability status --- p.16 / Chapter 2.1.6.5 --- Clinical uses of biomarkers for colorectal cancer --- p.17 / Chapter 2.1.7 --- Choice of cell line as colorectal cancer model --- p.17 / Chapter 2.1.8 --- Aims of study --- p.17 / Chapter 2.2 --- Materials and Methods / Chapter 2.2.1 --- Verification of SW480 as a nuclear β-catenin positive cell line / Chapter 2.2.1.1 --- Maintenance of cell lines --- p.21 / Chapter 2.2.1.2 --- Antibody --- p.21 / Chapter 2.2.1.3 --- Agar block preparation for SW480 and CCD-18C0 cells --- p.22 / Chapter 2.2.1.4 --- Immunocytochemical staining --- p.22 / Chapter 2.2.2 --- Effect of anti-cancer drugs on cell viability / Chapter 2.2.2.1 --- Maintenance of cell lines --- p.22 / Chapter 2.2.2.2 --- MTT cell viability assay --- p.23 / Chapter 2.3 --- Results / Chapter 2.3.1 --- SW480 is a β-catenin positive cell line --- p.24 / Chapter 2.3.2 --- Antiproliferative effects of cytotoxic drugs in SW480 cells / Chapter 2.3.2.1 --- 5-Fluorouracil --- p.26 / Chapter 2.3.2.2 --- Oxaliplatin --- p.29 / Chapter 2.3.2.3 --- Irinotecan --- p.31 / Chapter 2.4 --- Discussion / Chapter 2.4.1 --- SW480 as a nuclear β-catenin positive cell line --- p.33 / Chapter 2.4.2 --- Antiproliferative effects of 5-fluorouracil in SW480 cells --- p.33 / Chapter 2.4.3 --- Summary --- p.34 / Chapter Chapter 3: --- Effect of 5-fluorouracil on mRNA expression in SW480 cells / Chapter 3.1 --- Literature Review / Chapter 3.1.1 --- Application of quantitative real-time polymerase chain reaction in cancer research / Chapter 3.1.1.1 --- Principles of quantitative real-time polymerase chain reaction --- p.36 / Chapter 3.1.1.2 --- Advantages of quantitative real-time polymerase chain reaction over conventional polymerase chain reaction --- p.39 / Chapter 3.1.1.3 --- Determination of colorectal cancer biomarkers by quantitative real-time polymerase chain reaction --- p.39 / Chapter 3.2 --- Materials and Methods / Chapter 3.2.1 --- Determination of the effect of 5-fluorouracil on mRNA expression in SW480 cells / Chapter 3.2.1.1 --- Treatment of cells --- p.40 / Chapter 3.2.1.2 --- Extraction of total RNA from SW480 cells --- p.40 / Chapter 3.2.1.3 --- Removal of genomic DNA --- p.41 / Chapter 3.2.1.4 --- Determination of the efficiency of genomic DNA removal --- p.42 / Chapter 3.2.1.5 --- Determination of the purity and concentration of RNA --- p.42 / Chapter 3.2.1.6 --- Determination of the integrity of RNA --- p.43 / Chapter 3.2.1.7 --- First strand cDNA synthesis --- p.44 / Chapter 3.2.1.8 --- Real-time polymerase chain reaction using human Wnt signaling pathway RT2 ProfileŕёØ PCR array --- p.44 / Chapter 3.2.1.9 --- Calculation of the fold-change in genes expression between the 5-FU treated and control SW480 cells --- p.45 / Chapter 3.3 --- Results / Chapter 3.3.1 --- The quality and quantity of RNA --- p.46 / Chapter 3.3.2 --- Effects of 5-fluorouracil on genes expression in SW480 cells --- p.48 / Chapter 3.4 --- Discussion / Chapter 3.4.1 --- Alterations in mRNA expression in 5-fluorouracil treated SW480 cells --- p.55 / Chapter 3.4.1.1 --- Extracellular signaling molecules --- p.55 / Chapter 3.4.1.2 --- Canonical Wnt signaling pathway --- p.56 / Chapter 3.4.1.3 --- Regulators of cell cycle --- p.57 / Chapter 3.4.1.4 --- Regulators of growth and proliferation --- p.58 / Chapter 3.4.1.5 --- Regulators of transcription --- p.58 / Chapter 3.4.1.6 --- Regulators of Wnt receptor signaling pathway --- p.60 / Chapter 3.4.1.7 --- Other genes involved in Wnt signaling --- p.61 / Chapter 3.4.2 --- Limitations of Q-RT-PCR --- p.61 / Chapter 3.4.3 --- Summary --- p.62 / Chapter Chapter 4: --- Effect of 5-fluorouracil on proteins expression in SW480 cells / Chapter 4.1 --- Literature Review / Chapter 4.1.1 --- From mRNA to proteins --- p.63 / Chapter 4.1.2 --- Application of proteomics in cancer research --- p.63 / Chapter 4.1.3 --- Two-dimensional gel electrophoresis --- p.64 / Chapter 4.1.4 --- Principles of MALDI TOF mass spectrometry --- p.64 / Chapter 4.1.5 --- Peptide mass fingerprinting --- p.65 / Chapter 4.1.6 --- Drug response proteins detected by proteomics in colorectal cancer cell lines --- p.65 / Chapter 4.1.7 --- Detection of biomarker in colorectal cancer formation using proteomics --- p.66 / Chapter 4.2 --- Materials and Methods / Chapter 4.2.1 --- Determination of the effect of 5-fluorouracil on proteins expression in SW480 cells / Chapter 4.2.1.1 --- Treatment of cells --- p.67 / Chapter 4.2.1.2 --- Cell lysis --- p.67 / Chapter 4.2.1.3 --- Protein quantitation of cell lysate --- p.67 / Chapter 4.2.1.4 --- Sample preparation for two-dimensional electrophoresis --- p.68 / Chapter 4.2.1.5 --- Two-dimensional electrophoresis --- p.69 / Chapter 4.2.1.6 --- Silver staining --- p.69 / Chapter 4.2.1.7 --- Image analysis --- p.70 / Chapter 4.2.1.8 --- In-gel protein digestion --- p.70 / Chapter 4.2.1.9 --- Peptide mass fingerprinting using mass spectrometry --- p.71 / Chapter 4.3 --- Results / Chapter 4.3.1 --- Protein expression patterns of 5-fluorouracil treated and untreated SW480 cells by 2-dimensional electrophoresis --- p.72 / Chapter 4.3.2 --- Identification of the differentially expressed proteins after 5-fluorouracil treatment in SW480 cells --- p.75 / Chapter 4.4 --- Discussion / Chapter 4.4.1 --- Effects of 5-fluorouracil on protein expression in SW480 cells --- p.82 / Chapter 4.4.1.1 --- Identified upregulated proteins after 5-fluorouracil treatment in SW480 cells / Chapter 4.4.1.1.1 --- Cyclophilin A --- p.83 / Chapter 4.4.1.1.2 --- Cytokeratin 19 --- p.83 / Chapter 4.4.1.1.3 --- Cytokeratin 8 --- p.84 / Chapter 4.4.1.1.4 --- RAN --- p.84 / Chapter 4.4.1.1.5 --- Heat shock protein 27 --- p.84 / Chapter 4.4.1.1.6 --- Peroxiredoxin 6 --- p.85 / Chapter 4.4.1.2 --- Identified dowiiregulated proteins after 5-fluorouracil treatment in SW480 cells / Chapter 4.4.1.2.1 --- Heat shock protein 60 --- p.86 / Chapter 4.4.1.2.2 --- Cytokeratin 18 --- p.86 / Chapter 4.4.1.2.3 --- Cytokeratin 9 --- p.86 / Chapter 4.4.1.2.4 --- Carbamoylphosphate synthetase I --- p.87 / Chapter 4.4.1.2.5 --- a-Enolase --- p.87 / Chapter 4.4.1.2.6 --- Heat shock protein 70 --- p.87 / Chapter 4.4.1.2.7 --- nm23 --- p.88 / Chapter 4.4.1.2.8 --- β-actin --- p.88 / Chapter 4.4.2 --- Limitations of proteomics profiling --- p.89 / Chapter 4.4.3 --- Summary --- p.90 / Chapter Chapter 5: --- Verification of proteinśة identities by immunocytochemical staining / Chapter 5.1 --- Materials and Methods / Chapter 5.1.1 --- Antibodies --- p.91 / Chapter 5.1.2 --- Treatment of cells --- p.91 / Chapter 5.1.3 --- Agar block preparation of SW480 cells --- p.92 / Chapter 5.1.4 --- Immunocytochemical staining and evaluation --- p.92 / Chapter 5.1.5 --- Polymer-based immunohistochemical detection system --- p.93 / Chapter 5.1.6 --- Statistical analyses --- p.93 / Chapter 5.2 --- Results / Chapter 5.2.1 --- Confirmation of proteomic findings using immunocytochemical stainings in paraffin-embedded sections of 5-fluorouracil treated and untreated SW480 cells --- p.94 / Chapter 5.3 --- Discussion / Chapter 5.3.1 --- Immunocytochemical staining to verify proteomics findings of 5-fluorouracil treated and untreated SW480 cells --- p.99 / Chapter 5.3.2 --- Limitations of ICC staining --- p.100 / Chapter 5.3.3 --- Summary --- p.100 / Chapter Chapter 6: --- Conclusions and future perspectives / Chapter 6.1 --- Significance of study --- p.101 / Chapter 6.2 --- Future perspectives --- p.102 / References --- p.105

Ng, Po Yan. / Thesis submitted in: November 2006. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 109-120). / Abstracts in English and Chinese. / Acknowledgement --- p.i / Abstract --- p.ii / 摘要 --- p.iii / Table of Contents --- p.iv / List of Figures --- p.viii / List of Tables --- p.xi / List of Abbreviation --- p.xii / Chapter Chapter 1 --- Introduction / Chapter 1.1 --- A General Introduction to the Development and Clinical Activities of Platinum Drugs --- p.1 / Chapter 1.1.1 --- Platinum Drugs used in a Clinical Setting --- p.4 / Chapter 1.1.2 --- Platinum Drugs under Clinical Trials --- p.5 / Chapter 1.1.3 --- Platinum Compounds with Dual Mechanisms --- p.7 / Chapter 1.2 --- Platinum Drug Antitumor Mechanism --- p.9 / Chapter 1.3 --- Limitations of Platinum Drugs --- p.12 / Chapter 1.3.1 --- Toxicity --- p.12 / Chapter 1.3.2 --- Drug Resistance or Cross Resistance --- p.15 / Chapter 1.3.2.1 --- Reduced Drug Accumulation or Increased Drug Efflux --- p.16 / Chapter 1.3.2.2 --- Drug Inactivation --- p.18 / Chapter 1.3.2.3 --- Enhanced DNA Repair --- p.19 / Chapter 1.4 --- Why Combinational Therapy? --- p.21 / Chapter 1.4.1 --- Antimetabolites --- p.20 / Chapter 1.4.2 --- Topoisomerase Inhibitors --- p.22 / Chapter 1.4.3 --- Tubulin-Active Antimitotic Agents --- p.24 / Chapter 1.4.4 --- Demethylcantharidin as a potential candidate for drug combination --- p.28 / Chapter 1.5 --- Study Objectives --- p.31 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1 --- Cell Lines --- p.33 / Chapter 2.2 --- Cancer Cell Preparation / Chapter 2.2.1 --- Chemicals and Reagents --- p.33 / Chapter 2.2.2 --- Cell Culture Practice --- p.34 / Chapter 2.2.2.1 --- Subcultures --- p.35 / Chapter 2.2.2.2 --- Cryopreservation --- p.37 / Chapter 2.2.2.3 --- Thawing Cryopreservated Cells --- p.38 / Chapter 2.2.3 --- Development of Drug-Resistant Cell Lines --- p.39 / Chapter 2.3 --- Growth Inhibition Assay / Chapter 2.3.1 --- Evaluation of Cytotoxicity in vitro --- p.40 / Chapter 2.3.2 --- Drug Pretreatment --- p.43 / Chapter 2.3.3 --- Drug Pre-sensitization with Concurrent Treatment --- p.44 / Chapter 2.4 --- Calculations for Drug Combinations --- p.46 / Chapter 2.5 --- Statistical Analysis --- p.49 / Chapter Chapter 3 --- Results and Discussions / Chapter 3.1 --- In vitro Cytotoxicity and Evaluation of Drug Resistance --- p.50 / Chapter 3.2 --- Role of Leaving Ligand in a Platinum Complex --- p.58 / Chapter 3.3 --- Priority in Selecting the Most Effective Drug Combination --- p.66 / Chapter 3.4 --- Drug Combination Studies / Chapter 3.4.1 --- Drug Combination Prescreening --- p.68 / Chapter 3.4.1.1 --- Comparison of the effectiveness of the three Drug Combinations --- p.72 / Chapter 3.4.1.2 --- Rationale for Drug Combination Studies presented in Section 3.4.2 & 3.4.3 --- p.73 / Chapter 3.4.2 --- Drug Pre-sensitization Studies in Colorectal Cancer Cell Lines --- p.74 / Chapter 3.4.2.1 --- Comparison of Drug Pre-sensitization Treatment in Sensitive Colorectal Cancer Cell Lines --- p.84 / Chapter 3.4.2.2 --- Comparison of Drug Pre-sensitization Treatment in Sensitive and Oxaliplatin Resistant HCT116 Colorectal Cancer Cell Lines --- p.87 / Chapter 3.4.3 --- Drug Pre-sensitization Studies in Liver Cancer Cell Lines --- p.89 / Chapter 3.4.3.1 --- Comparison of Drug Pre-sensitization Treatment in Sensitive Liver Cancer Cell Lines --- p.99 / Chapter 3.4.3.2 --- Comparison of Drug Pre-sensitization Treatment in Sensitive and Cisplatin Resistant SK-Hepl Liver Cancer Cell Line --- p.101 / Chapter 3.5 --- Possible Explanation to the Observed Drug Combination Effect --- p.103 / Chapter 3.6 --- General Protocols for Drug Combinations --- p.105 / Chapter Chapter 4 --- Conclusions / Reference --- p.109 / Appendices --- p.121 / Chapter I a. --- "Raw Data of Pre-screening for HCT116 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.122 / Chapter I b. --- "Raw Data of Pre-screening for HCT116 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.123 / Chapter II a. --- "Raw Data of Pre-screening for SK-Hepl (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.124 / Chapter II b. --- "Raw Data of Pre-screening for SK-Hepl ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.125 / Chapter III a. i) --- "Isobolograms for HCT116 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.126 / Chapter III a. ii) --- "Raw Data for HCT116 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.127 / Chapter III b. i) --- "Isobolograms for HCT116 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.128 / Chapter III b. ii) --- "Raw Data for HCT116 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.129 / Chapter IV a. i) --- "Isobolograms for HCT1160xaR (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.130 / Chapter IV a. ii) --- "Raw Data for HCT1160xaR (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.131 / Chapter IV b. i) --- "Isobolograms for HCT1160xaR ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.132 / Chapter IV b. ii) --- "Raw Data for HCT1160xaR ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.133 / Chapter V a. i) --- "Isobolograms for HT29 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.134 / Chapter V a. ii) --- "Raw Data for HT29 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.135 / Chapter V b. i) --- "Isobolograms for HT29 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.136 / Chapter V b. ii) --- "Raw Data for HT29 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.137 / Chapter VI a. i) --- Isobolograms for Hep G2 (Cisplatin and [Pt(DMC)(NH3)2]) --- p.138 / Chapter VI a. ii) --- Raw Data for Hep G2 (Cisplatin and [Pt(DMC)(NH3)2]) --- p.139 / Chapter VI b. i) --- "Isobolograms for Hep G2 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.140 / Chapter VI b. ii) --- "Raw Data for Hep G2 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.141 / Chapter VII a. i) --- "isobolograms for SK Hep 1 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.142 / Chapter VII a. ii) --- "Raw Data for SK Hep 1 (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.143 / Chapter VII b.i) --- "Isobolograms for SK Hep 1 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.144 / Chapter VII b. ii) --- "Raw Data for SK Hep 1 ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.145 / Chapter VIII a. i) --- "Isobolograms for SK Hep ICisR (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.146 / Chapter VIII a. ii) --- "Raw Data for SK Hep ICisR (Cisplatin, [Pt(DMC)(NH3)2] and Pt(DMC)(NH2CH3)2])" --- p.147 / Chapter VIII b. i) --- "Isobolograms for SK Hep ICisR ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.148 / Chapter VIII b. ii) --- "Raw Data for SK Hep ICisR ([Pt(DMC)(R,R-DACH)] and Oxaliplatin)" --- p.149