UNIVERSITE LOUIS PASTEUR DE STRASBOURG I

Anne Anstett

Description

Niveau: Supérieur, Doctorat, Bac+8
UNIVERSITE LOUIS PASTEUR DE STRASBOURG I ECOLE DOCTORALE SCIENCES DE LA VIE ET DE LA SANTE THESE pour obtenir le grade de DOCTEUR DE L'UNIVERSITE LOUIS PASTEUR DE STRASBOURG Discipline : Sciences du Vivant Spécialité : Aspects moléculaires et cellulaires de la biologie Soutenue le 12 septembre 2005 et présentée par Anne ANSTETT APPROACH OF COMBINED CANCER GENE THERAPY AND RADIATION: RESPONSE OF PROMOTERS TO IONIZING RADIATION devant le jury composé de : Pr. Kurt BALLMER-HOFER Rapporteur Dr. Pierre BISCHOFF Rapporteur Pr. Josef JIRICNY Examinateur Pr. Jean-Pierre CAZENAVE Directeur de thèse

• promoters cloned


• pathway


• profiling using


• radiation-inducible gene


• specific binding


• gene therapy


• gene expression

Sujets

• Cazenave
• Bischoff
• Path
• Gene therapy
• Gene expression

Informations

• Publié par: profil-zyak-2012
• Publié le: 01 septembre 2005
• Nombre de lectures: 64
• Poids de l'ouvrage: 4 Mo

Extrait

UNIVERSITE LOUIS PASTEUR DE STRASBOURG I
ECOLE DOCTORALE SCIENCES DE LA VIE ET DE LA SANTE



THESE

pour obtenir le grade de

DOCTEUR DE L’UNIVERSITE LOUIS PASTEUR DE
STRASBOURG

Discipline : Sciences du Vivant
Spécialité : Aspects moléculaires et cellulaires de la biologie




Soutenue le 12 septembre 2005 et présentée par


Anne ANSTETT



APPROACH OF COMBINED CANCER GENE THERAPY
AND RADIATION: RESPONSE OF PROMOTERS TO

IONIZING RADIATION




devant le jury composé de :


Pr. Kurt BALLMER-HOFER Rapporteur
Dr. Pierre BISCHOFF Rapporteur
Pr. Josef JIRICNY Examinateur
Pr. Jean-Pierre CAZENAVE Directeur de thèse

UNIVERSITE LOUIS PASTEUR DE STRASBOURG I
ECOLE DOCTORALE SCIENCES DE LA VIE ET DE LA SANTE



THESE

pour obtenir le grade de

DOCTEUR DE L’UNIVERSITE LOUIS PASTEUR DE
STRASBOURG

Discipline : Sciences du Vivant
Spécialité : Aspects moléculaires et cellulaires de la biologie




Soutenue le 12 septembre 2005 et présentée par


Anne ANSTETT



APPROCHE DE THERAPIE GENIQUE ANTI-
CANCEREUSE COMBINEE A L’IRRADIATION :

ETUDE DE LA REPONSE DE PROMOTEURS AUX
RADIATIONS IONISANTES



devant le jury composé de :


Pr. Kurt BALLMER-HOFER Rapporteur
Dr. Pierre BISCHOFF Rapporteur
Pr. Josef JIRICNY Examinateur
Pr. Jean-Pierre CAZENAVE Directeur de thèse

TABLE OF CONTENTS



SUMMARY..............................................................................................................................iii

RÉSUMÉ................................................................................................................................... v

SYNTHÈSE DE LA THÈSE ................................................................................................. vii

LIST OF ABBREVIATIONS............................................................................................... xvi

1 INTRODUCTION ................................................................................................................ 1
1.1 Cancer and cancer treatments............................................................................................ 1
1.2 Ionizing radiation as a cancer treatment modality ............................................................ 2
1.3 The cellular and molecular response to ionizing radiation ............................................... 4
1.3.1 Plasma membrane damage signaling to ionizing radiation......................................... 4
1.3.1.1 The EGF receptor signaling .................................................................................. 5
1.3.1.2 The death receptors signaling................................................................................ 6
1.3.1.3 The ceramide signaling pathway........................................................................... 6
1.3.2 DNA damage signaling to ionizing radiation.............................................................. 8
1.3.2.1 DNA damage signal transduction ......................................................................... 8
1.3.2.2 The cell cycle arrest in response to DNA double strand breaks............................ 9
1.3.2.3 The protein p53 ................................................................................................... 12
1.3.3 Apoptosis following DNA damage 18
1.3.3.1 The death receptor apoptotic pathway ................................................................ 18
1.3.3.2 The mitochondrial apoptotic pathway................................................................. 18
1.3.4 Examples of transcription factors activated by ionizing radiation............................ 20
1.3.4.1 Activating protein-1 (AP-1) ................................................................................ 20
1.3.4.2 Nuclear factor kappa B (NF- κB)......................................................................... 23
1.3.4.3 Early growth response gene-1 (EGR-1).............................................................. 25
1.4 Ionizing radiation-combined cancer therapies: inhibition of angiogenesis and
ionizing radiation............................................................................................................. 29
1.4.1 Angiogenesis and angiogenesis inhibition ................................................................ 29
1.4.2 Angiogenesis inhibition and ionizing radiation ........................................................ 32

2 AIM OF THE STUDY ........................................................................................................ 35

3 TRANSCRIPTIONAL RESPONSES OF ENDOTHELIAL CELLS TO IONIZING
RADIATION........................................................................................................................... 37
3.1 Introduction ..................................................................................................................... 37
3.2 Materials and Methods.................................................................................................... 38
3.3 Results ............................................................................................................................. 44
3.3.1 Biological responses of endothelial cells to ionizing radiation................................. 44
3.3.1.1 Cell cycle arrest................................................................................................... 44
3.3.1.2 Signal transduction.............................................................................................. 45
3.3.1.3 Anproliferation activity....................................................................................... 46
3.3.1.4 Apoptosis activity................................................................................................ 47
3.3.2 Gene expression profiling of endothelial cells exposed to ionizing radiation .......... 49
3.3.2.1 Gene expression profiling of irradiated EA.hy 926 ............................................ 49
i
3.3.2.2 Comparative gene expression profiling induced by ionizing radiation............... 72
3.4 Discussion ....................................................................................................................... 76

4 PROMOTER STUDY IN RESPONSE TO IONIZING RADIATION.......................... 81
4.1 Introduction ..................................................................................................................... 81
4.2 Materials and Methods.................................................................................................... 82
4.3 Results ............................................................................................................................. 91
4.3.1 Native radiation-inducible promoter screening......................................................... 91
4.3.1.1 BTG2 gene and promoter.................................................................................... 91
4.3.1.2 4-1BB gene and promoter ................................................................................... 97
4.3.1.3 NK4 gene and 5’-upstream region .................................................................... 100
4.3.2 Synthetic NF- κB binding sites to ionizing radiation exposure ............................... 108
4.4 Discussion ..................................................................................................................... 112

5 CONCLUSIONS AND PERSPECTIVES....................................................................... 117

6 REFERENCES.................................................................................................................. 119

ACKNOWLEDGEMENTS................................................................................................. 131





iiSUMMARY

Gene therapy is an emerging cancer treatment modality. We are interested in developing a
radiation-inducible gene therapy system to sensitize the tumor vasculature to the effects of
ionizing radiation (IR) treatment. An expression system based on irradiation-inducible
promoters will drive the expression of anti-tumor genes in the tumor vasculature. Solid
tumors are dependent on angiogenesis, a process in which new blood vessels are formed from
the pre-existing vasculature. Vascular endothelial cells are untransformed and genetically
stable, thus avoiding the problem of resistance to the treatments. Vascular endothelial cells
may therefore represent a suitable target for this therapeutic gene therapy strategy.
The identification of IR-inducible promoters native to endothelial cells was performed by
gene expression profiling using cDNA microarray technology. We describe the genes
modified by clinically relevant doses of IR. The extension to high doses aimed at studying the
effects of total radiation delivery to the tumor. The radio-inducibility of the genes selected for
pr