Polyethylenimine and its derivatives [Elektronische Ressource] : investigation of in vivo fate, subcellular trafficking and development of novel vector systems / vorgelegt von Thomas Merdan

Sujets

• Gesundheit

Informations

• Publié par: philipps-universitat_marburg
• Publié le: 01 janvier 2003
• Nombre de lectures: 32
• Langue: English
• Poids de l'ouvrage: 3 Mo

Extrait

POLYETHYLENIMINE AND ITS DERIVATIVES:
INVESTIGATION OF IN VIVO FATE, SUBCELLULAR
TRAFFICKING AND DEVELOPMENT OF NOVEL
VECTOR SYSTEMS
Dissertation
zur
Erlangung des Doktorgrades
der Naturwissenschaften
(Dr. rer. nat.)
dem Fachbereich Pharmazie der
Philipps-Universität Marburg
vorgelegt von
Thomas Merdan
aus Bitburg
Marburg/Lahn 2003

























Vom Fachbereich Pharmazie der Philipps-Universität Marburg als Dissertation
am 06.10.2003 angenommen.
Erstgutachter: Prof. Dr. T. Kissel
Zweitgutachter: Prof. Dr. U. Bakowsky
Drittgutachter: Prof Dr. Jindrich Kopecek
Tag der mündlichen Prüfung: 11.11.2003


3
Die vorliegende Arbeit
entstand auf Anregung und unter der Leitung von
Herrn Prof. Dr. Thomas Kissel
am Institut für Pharmazeutische Technologie und Biopharmazie
der Philipps-Universität Marburg.









5
GEWIDMET MEINEN ELTERN
IN LIEBE UND DANKBARKEIT


ACKNOWLEDGEMENTS
My greatest thanks belong to my supervisors, Prof. Thomas Kissel in Marburg
and Prof. Jindrich Kopecek in Salt Lake City, USA, who always provided me
with motivation and guidance in my research. Their broad knowledge and
creative ideas were crucial for the success of this dissertation.
Next I would like to thank Prof. Pavla Kopecková for her great help and advice
in the laboratory, as well as for organizing everything necessary to settle down
in Utah.
I would like to thank my colleagues from Philipps University in Marburg.
Especially the members of the PEI research group Dr. Dagmar Fischer, Holger
Petersen, Klaus Kunath, Carola Brus and Michael Neu for valuable discussions
about our research projects. Prof. Xintao Shuai thank you so much for a very
fruitful collaboration and interesting insights into physical chemistry.
Furthermore I would like to thank Elke Kleemann, Ulrich Westedt, Florian
Unger and all other members of the Kissel laboratory for the nice time in
Marburg. A special acknowledgement belongs to Lea Ann Dailey for her skillful
revisions of my English manuscripts. Thank you, as well, to my laboratory
colleague, Christine Oster, whose devotion to science always impressed and
motivated me.
Furthermore, I thank my friends at the University of Utah Aparna Nori, Jon
Callahan, Keith Jensen, Nobuhiro Nishiyama, Pad Chivukula, Arshan Amiri,
Vaikunth Cuchelkar and all the others. Thank you for the friendly reception into
the Kopecek research group and for the great time in the lab, in the pubs and on
the slopes.
Thank you to Prof. Udo Bakowsky from the Saarland University and to James
Ellis from the University of Nottingham for performing atomic force
microscopy experiments for my projects.












Another major thanks belongs to Gudrun Hohorst, Eva Mohr, Nicole
Bamberger, Klaus Keim and Klaus Guckelsberger for excellent help with the
animal and cell culture experiments, as well as the graphics design.
I also want to acknowledge our precision mechanic Lothar-Walter Kempf, for
the skillful production of a heating device, which enabled us to observe living
cells under the confocal microscope.
Andreas Mitsch, Peter Nold, Margit Janat and Markus Kleinschmidt thank you
for the nice lunch breaks and interesting discussions.
Most of all I would like to thank Sabine Rückauer for her patience in all phases
during the past three years.
TABLE OF CONTENTS
CHAPTER 1 INTRODUCTION
Aims of this dissertation…………………………………………………….. 14
PROSPECTS FOR CATIONIC POLYMERS IN GENE AND
OLIGONUCLEOTIDE THERAPY AGAINST CANCER
Summary………………………………………………………………… 18
1. Introduction ………………………………………………………….. 18
2. Non Viral vectors…………………………………………………….. 22
2.1. Polyethylenimine………………………………………………… 22
2.2. Poly(L-lysine)……………………………………………………. 24
2.3. Imidazole containing polymers………………………………….. 24
2.4. Chitosans………………………………………………………… 25
2.5. Dendrimers………………………………………………………. 27
2.6. Comparison of transfection agents………………………………. 27
3. Hurdles in polymer based nucleic acid delivery……………………... 29
3.1. Hurdles on the systemic level……………………………………. 29
3.2. Hurdles on the cellular level……………………………………... 39
4. Gene/nucleic acid therapy towards cancer…………………………… 64
4.1. Antisense oligonucleotides and ribozymes……………………… 64
4.2. Knockout gene therapy…………………………………………... 69
4.3. Gene replacement/gene augmentation…………………………… 70
4.4. Suicide gene therapy…………………………………………….. 71
4.5. Cancer vaccination using gene therapy approaches…………….. 73
4.6. Bystander effect………………………………………………….. 74
4.7. Clinical trials……………………………………………………. 75
5. Conclusions…………………………………………………………... 78
6. References……………………………………………………………. 81
CHAPTER 2 INTRACELLULAR PROCESSING OF POLYETHYLEN-
IMINE/RIBOZYME COMPLEXES CAN BE OBSERVED IN LIVING
CELLS USING CONFOCAL LASER SCANNING MICROSCOPY AND
INHIBITOR EXPERIMENTS
Summary………………………………………………………………… 106
Introduction……………………………………………………………… 107
Materials and methods…………………………………………………... 108
Results…………………………………………………………………… 112
Discussion……………………………………………………………….. 119
References……………………………………………………………….. 123
CHAPTER 3 PEGYLATED POLYETHYLENIMINE – FAB’ ANTIBODY
FRAGMENT CONJUGATES FOR TARGETED GENE DELIVERY TO
HUMAN OVARIAN CARCINOMA CELLS
Summary………………………………………………………………….128
Introduction……………………………………………………………… 129
Materials and methods…………………………………………………... 131
Results and discussion…………………………………………………... 137
References……………………………………………………………….. 149
CHAPTER 4 COMPARISON OF IN VITRO AND IN VIVO
PROPERTIES OF ELECTROSTATIC COMPLEXES PREPARED WITH
EITHER POLYETHYLENIMINE OR PEGYLATED POLYETHYLEN-
IMINE AND PLASMID DNA
Summary………………………………………………………………… 154
Introduction……………………………………………………………… 155
Materials and methods………………………………………………….. 157
Results and discussion………………………………………………….. 162
References……………………………………………………………….. 176