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Alteration of drug sensitivity in human squamous carcinoma A431 cells by chronic exposure to epidermal growth factor.

Cheung Tsz Man. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 187-203). / Abstracts in English and Chinese. / Acknowledgements --- p.is / Abbreviations --- p.II / Abstracts --- p.V / List of Figures --- p.IX / List of Tables --- p.XIII / Contents / Chapter Chapter 1. --- General Introduction / Chapter 1.1 --- Cancer --- p.1 / Chapter 1.2 --- Growth Factor --- p.2 / Chapter 1.3 --- Growth Factor and Growth Factor Receptor --- p.4 / Chapter Chapter 2. --- Alteration of EGF Responses and EGFR Signaling in EGF-conditioned A431 cells / Chapter 2.1 --- Background Information / Chapter 2.1.1 --- Epidermal Growth Factor (EGF) --- p.6 / Chapter 2.1.2 --- Epidermal Growth Factor Receptor (EGFR) --- p.10 / Chapter 2.1.2.1 --- The Structure of EGFR --- p.10 / Chapter 2.1.2.2 --- The EGFR Family --- p.11 / Chapter 2.1.2.3 --- EGFR Activation --- p.13 / Chapter 2.1.3 --- The Intracellular Signal Transduction Pathways in EGFR Signaling --- p.18 / Chapter 2.1.3.1 --- The Ras/Raf/MAPK Pathway (MAPK pathway) --- p.19 / Chapter 2.1.3.2 --- The Jak/Stat Pathway --- p.23 / Chapter 2.1.3.3 --- The PI3K/Akt Pathway --- p.28 / Chapter 2.1.4 --- EGFR and Cancer --- p.31 / Chapter 2.1.5 --- EGFR-targeted Cancer Therapy --- p.35 / Chapter 2.1.5.1 --- Monoclonal Antibody (MAb) --- p.36 / Chapter 2.1.5.2 --- Immunotoxin Conjugates --- p.37 / Chapter 2.1.5.3 --- Bispecific Antibody --- p.37 / Chapter 2.1.5.4 --- Small-molecule EGFR Tyrosine Kinase Inhibitor (EGFR-TKI) --- p.38 / Chapter 2.1.5.5 --- Antisense Oligonucleotide --- p.39 / Chapter 2.2 --- Objectives --- p.41 / Chapter 2.3 --- Materials and Methods / Chapter 2.3.1 --- Materials --- p.42 / Chapter 2.3.2 --- Methods / Chapter 2.3.2.1 --- Cell Lines --- p.44 / Chapter 2.3.2.1.1 --- Establishment of Epidermal Growth Factor (EGF)-conditioned A431 Cells (EGF-conditioned Cells) ´ؤ AC Cells --- p.44 / Chapter 2.3.2.2 --- Growth Curve between A431 Parent Cells and EGF-conditioned Cells --- p.45 / Chapter 2.3.2.3 --- Epidermal Growth Factor (EGF) Sensitivity Assay --- p.45 / Chapter 2.3.2.4 --- Western Blot Analysis --- p.47 / Chapter 2.3.2.4.1 --- Protein Samples Preparation --- p.47 / Chapter 2.3.2.4.2 --- Protein Assay (by BCA Protein Assay Reagent) --- p.48 / Chapter 2.3.2.4.3 --- Protein Electrophoresis --- p.49 / Chapter 2.3.2.4.4 --- Electroblot (Protein Transfer) --- p.50 / Chapter 2.3.2.4.5 --- Antibody Probing (Immunoblotting) --- p.51 / Chapter 2.4 --- Results / Chapter 2.4.1 --- Growth Curve --- p.53 / Chapter 2.4.2 --- EGF Responses of A431 Parent Cells and EGF-conditioned Cells by MTT Assay --- p.55 / Chapter 2.4.3 --- The EGFR Expression Levels in A431 Parent Cells and EGF-conditioned Cells by Western Blot Analysis --- p.57 / Chapter 2.4.4 --- EGF-induced Protein Tyrosine Phosphorylation Pattern in A431 Parent Cells and EGF-conditioned Cells by Western Blot Analysis --- p.59 / Chapter 2.4.5 --- The Expression Profiles of EGFR Signaling Molecules in A431 Parent Cells and EGF-conditioned Cells by Western Blot Analysis --- p.61 / Chapter 2.4.5.1 --- The Ras/Raf/MAPK Pathway --- p.62 / Chapter 2.4.5.2 --- The Jak/Stat Pathway --- p.63 / Chapter 2.4.5.3 --- The PI3K/Akt Pathway --- p.64 / Chapter 2.4.6 --- The Cellular Responses to the Modifiers that Targeting the EGFR Signaling --- p.68 / Chapter 2.4.6.1 --- The Sensitivity of A431 Parent Cells and EGF-conditioned Cells to Various Signaling Modifiers --- p.69 / Chapter 2.4.6.2 --- The Influence of EGFR Signaling Modifiers on EGF --- p.76 / Chapter 2.5 --- Discussion --- p.85 / Chapter Chapter 3. --- The Inter-relationship between the Differential Anti-cancer Drugs Sensitivity and Alteration of EGFR Signaling in EGF-conditioned A431 Cells / Chapter 3.1 --- Background Information / Chapter 3.1.1 --- Drug Resistance and its Mechanisms in Tumor Cells --- p.90 / Chapter 3.1.2 --- Anti-cancer Drugs ´ؤ Introduction / Chapter 3.1.2.1 --- Camptothecin (CPT) --- p.93 / Chapter 3.1.2.2 --- Methotrexate (MTX) --- p.95 / Chapter 3.1.2.3 --- 5-fluorouracil (5-Fu) --- p.98 / Chapter 3.1.2.4 --- Vincristine (VCR) and Taxol --- p.104 / Chapter 3.1.2.5 --- Cisplatin (cis-DDP) --- p.108 / Chapter 3.2 --- Objectives --- p.110 / Chapter 3.3. --- Materials and Methods / Chapter 3.3.1 --- Materials --- p.112 / Chapter 3.3.2 --- Methods / Chapter 3.3.2.1 --- Cell Lines --- p.115 / Chapter 3.3.2.2 --- Determination of Drug Sensitivity by MTT Assay --- p.115 / Chapter 3.3.2.2.1 --- Determination the Influence of EGFR Signaling Modifiers on the Differential Anticancer Drugs Sensitivity by MTT Assay --- p.115 / Chapter 3.3.2.3 --- Semi-quantitative RT-PCR --- p.116 / Chapter 3.3.2.3.1 --- Preparation of RNA Samples --- p.116 / Chapter 3.3.2.3.2 --- RT-PCR --- p.117 / Chapter 3.3.2.4 --- DNA Fragmentation Assay --- p.118 / Chapter 3.3.2.5 --- Western Blot Analysis --- p.120 / Chapter 3.3.2.6 --- Northern Blot Analysis --- p.120 / Chapter 3.4 --- Results / Chapter 3.4.1 --- The Responses to Various Anti-cancer Drugs / Agents in A431 Parent Cells and EGF-conditioned Cells --- p.122 / Chapter 3.4.2 --- The Expressions of Classical Cellular Drug Resistant Factors in EGF-conditioning-associated Differential Anti-cancer Drugs Sensitivity --- p.126 / Chapter 3.4.2.1 --- Camptothecin Sensitivity --- p.126 / Chapter 3.4.2.2 --- Methotrexate Sensitivity --- p.130 / Chapter 3.4.2.3 --- 5-fluorouracil Sensitivity --- p.135 / Chapter 3.4.2.4 --- Vincristine and Taxol Sensitivity --- p.141 / Chapter 3.4.3 --- EGFR Signaling Modifiers and Differential Anti-cancer Drugs Sensitivity by MTT Assay --- p.143 / Chapter 3.4.3.1 --- Methotrexate --- p.143 / Chapter 3.4.3.2 --- Vincristine --- p.147 / Chapter 3.4.3.3 --- Taxol --- p.149 / Chapter 3.5 --- Discussion --- p.153 / Chapter Chapter 4. --- Identification of Differentially Expressed Genes in A431 Parent Cells and EGF-conditioned Cells by Differential Display (DD) / Chapter 4.1 --- Introduction 一 Differential Display (DD) --- p.156 / Chapter 4.2 --- Objectives --- p.160 / Chapter 4.3 --- Materials and Methods / Chapter 4.3.1 --- Materials --- p.161 / Chapter 4.3.2 --- Methods / Chapter 4.3.2.1 --- Cell Lines --- p.163 / Chapter 4.3.2.2 --- RT-PCR-based mRNA Differential Display --- p.163 / Chapter 4.3.2.2.1 --- Preparation of RNA Samples --- p.163 / Chapter 4.3.2.2.2 --- Identification of Differentially Expressed Genes by RT-PCR --- p.164 / Chapter 4.3.2.2.3 --- Reamplification of cDNA Probes --- p.164 / Chapter 4.3.2.2.4 --- Subcloning of the Differentially Expressed cDNAs --- p.165 / Chapter 4.3.2.2.4.1 --- Preparation of the Ultra-competent E.coli Cells for Transformation --- p.165 / Chapter 4.3.2.2.4.2 --- Preparation of Cloning Vector --- p.166 / Chapter 4.3.2.2.4.3 --- Transformation --- p.166 / Chapter 4.3.2.2.5 --- Verification of cDNA Differentially Expression by Colony-PCR and Northern Blot Analysis --- p.167 / Chapter 4.3.2.2.5.1 --- Colony-PCR --- p.167 / Chapter 4.3.2.2.5.2 --- Preparation of Cloned Plasmid cDNA and Bacterial Glycerol Stocks --- p.167 / Chapter 4.3.2.2.5.3 --- Preparation of cDNA Probes for Northern Blot Analysis --- p.168 / Chapter 4.3.2.2.5.4 --- Northern Blot Analysis --- p.168 / Chapter 4.3.2.2.6 --- Sequencing of the Desired Cloned cDNA Inserts --- p.170 / Chapter 4.3 --- Results --- p.171 / Chapter 4.4 --- Discussion --- p.180 / Chapter Chapter 5. --- General Conclusion and Future Perspectives / Chapter 5.1 --- General Conclusion --- p.182 / Chapter 5.2 --- Future Perspectives --- p.185 / References --- p.187

Identiferoai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_324974
Date January 2004
ContributorsCheung, Tsz Man., Chinese University of Hong Kong Graduate School. Division of Biochemistry.
Source SetsThe Chinese University of Hong Kong
LanguageEnglish, Chinese
Detected LanguageEnglish
TypeText, bibliography
Formatprint, xviii, 203 leaves : ill. (some col.) ; 30 cm.
RightsUse of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/)

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