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Identification of Novel Therapeutic Targets and Tumor Suppressor Genes in Colon Cancer Using Genome‐Wide High‐ Throughput Approaches

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Identification of Novel Therapeutic Targets and Tumor Suppressor Genes in Colon Cancer Using Genome‐Wide High‐ Throughput Approaches Identification of novel thera peutic targets and tu m or sup press or gene s in colon cancer using genome - wide high - throughput approac hes Sarah Bazzocco ADVERTIMENT . La consulta d’aquesta tesi queda condicionada a l’acceptació de les següents condicions d'ús: La difusió d’aquesta tesi per mitjà del servei TDX ( www.tdx.cat ) i a través del Dipòsit Digital de la UB ( diposit.ub.edu ) ha estat autoritzada pels titulars dels drets de propietat intel·lectual únicament per a usos privats emmarcats en activitats d’investigació i docència. No s’autoritza la seva reproducció amb finalitats de lucre ni la seva difusió i posada a disposici ó d es d’un lloc aliè al servei TDX ni al Dipòsit Digital de la UB . No s’autoritza la presentació del seu contingut en una finestra o marc aliè a TDX o al Dipòsit Digital de la UB (framing). Aquesta reserva de drets afecta tant al resum de presentació de la te si com als seus continguts. En la utilització o cita de parts de la tesi és obligat indicar el nom de la persona autora. ADVERTENCIA . La consulta de esta tesis queda condicionada a la aceptación de las siguientes condiciones de uso: La difusión de esta t esis por medio del servicio TDR ( www.tdx.cat ) y a través del Repositorio Digital de la UB ( diposit.ub.edu ) ha sido autorizada por los titulares de los derechos de propiedad intelectual únicamente para usos privados enmarcados en actividades de investigació n y docencia. No se autoriza su reproducción con finalidades de lucro ni su difusión y puesta a disposición desde un sitio ajeno al servicio TDR o al Repositorio Digital de la UB . No se autoriza la presentación de su contenido en una ventana o marco ajeno a TDR o al Repositorio Digital de la UB (framing). Esta reserva de derechos afecta tanto al resumen de presentación de la tesis como a sus contenidos. En la utilización o cita de partes de la tesis es obligado indicar el nombre de la persona autora. WARNING . On having consulted this thesis you’re accepting the following use conditions: Spreading this thesis by the TDX ( www.tdx.cat ) service and by the UB Digital Repository ( dipo sit.ub.edu ) has been authorized by the titular of the intellectual property rights only for private uses placed in investigation and teaching activities. Reproduction with lucrati ve aims is not authorized n or its spreading and availability from a site fore ign to the TDX service or to the UB Digital Repository . Introducing its content in a window or frame foreign to the TDX service or to the UB Digital Repository is not authorized (framing). Tho s e rights affect to the presentation summary of the thesis as well as to its contents. In the using or citation of parts of the thesis it’s obliged to indicate the name of the author. Identification of novel therapeutic targets and tumor suppressor genes in colon cancer using genome‐wide high‐ throughput approaches by Sarah Bazzocco A thesis submittedd in fulfillment of the requirements for the title of PhD by the University of Barcelona PhD program in Biomedicine The thesis was carried out at Molecular oncology Group Centre for Molecular Biology and Biochemistry Investigation in Nanomedicine (CIBBIM‐Nanomedicine) Vall d'Hebroon Hospital Research Institute Director: PhD candidate: Diego Arango del Corro Sarah Bazzocco Table of Contents Table of Content TABLE OF CONTENT ......................................................................................................................... I INDEX OF FIGURES ......................................................................................................................... IV INDEX OF TABLES ............................................................................................................................ V ABSTRACT ..................................................................................................................................... VI 1 INTRODUCTION ....................................................................................................................... 1 1.1 THE DIGESTIVE TRACT ................................................................................................................... 1 1.2 THE HUMAN INTESTINE ................................................................................................................. 2 1.2.1 Anatomy of the small intestine ....................................................................................... 2 1.2.2 Histology of the epithelium of the small intestine........................................................... 4 1.2.3 Anatomy and histology of the large intestine ................................................................. 6 1.2.4 Homeostasis of the human intestinal epithelium ............................................................ 6 1.2.4.1 Proliferation and differentiation of intestinal stem cells ............................................ 7 1.2.4.2 Signaling in the intestinal stem cells ........................................................................... 8 1.3 GENERAL CHARACTERISTICS OF CANCER ......................................................................................... 10 1.3.1 Types of cancer .............................................................................................................. 10 1.3.2 World and European cancer statistic ............................................................................ 11 1.3.3 Hallmarks of cancer ....................................................................................................... 13 1.3.4 Tumor suppressor genes and oncogenes ...................................................................... 16 1.3.5 Epigenetic alterations in cancer .................................................................................... 18 1.4 COLORECTAL CANCER ................................................................................................................. 19 1.4.1 Origin and clinical classification of colorectal cancer .................................................... 19 1.4.2 Incidence and statistics of colorectal cancer ................................................................. 22 1.4.3 Molecular pathways in colorectal cancer ...................................................................... 24 1.4.3.1 The multistep nature from normal epithelium to colorectal cancer ........................ 24 1.4.3.2 The distinct paths to colorectal cancer ..................................................................... 25 1.4.3.3 Microsatellite instability (MSI) ‐ DNA mismatch repair defects in colorectal cancer 26 1.4.3.4 – Chromosomal instability (CIN)/microsatellite stability (MSS) ‐ Genetic Instability in colorectal cancer ....................................................................................................................... 28 1.4.3.5 CpG island methylator phenotype (CIMP) ‐ Epigenetic changes in colorectal cancer 28 1.4.3.6 Somatic oncogene and tumor suppressor gene mutations in colorectal cancer ...... 29 1.4.4 Treatment of colorectal cancer ..................................................................................... 33 1.4.5 Treatment of colon cancer by stage .............................................................................. 33 1.4.5.1 Chemotherapeutic agents available for colorectal cancer treatment ...................... 34 1.4.5.2 Strategies to improve the objective response rate in chemotherapy for colorectal cancer patients .......................................................................................................................... 36 2 AIMS OF THE STUDY .............................................................................................................. 40 I Results 3 MATERIALS AND METHODS .................................................................................................. 41 3.1 MATERIALS USED IN THIS STUDY ................................................................................................... 41 3.1.1 Human colorectal cancer cell lines ................................................................................ 41 3.1.2 Primary colorectal tumor samples ................................................................................. 43 3.1.3 Antibodies ...................................................................................................................... 43 3.1.4 Primers ........................................................................................................................... 44 3.2 METHODS USED IN THIS STUDY ..................................................................................................... 45 3.2.1 Microarray data ............................................................................................................. 45 3.2.1.1 mRNA expression microarray analysis ...................................................................... 45 3.2.1.2 DNA methylation microarray analysis ....................................................................... 45 3.2.2 Identification of genes ................................................................................................... 46 3.2.2.1 Associations between mRNA expression and the doubling time of cell lines ........... 46 3.2.2.2 Associations between mRNA expression and promoter methylation levels ............ 46 3.2.3 Functional group enrichment analysis ........................................................................... 46 3.2.4 In vitro experiments ......................................................................................................
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