Le rôle des ultrasons, de l’IRM et de l’imagerie optique dans le cadre de l’activation locale de gènes et du dépôt local de médicaments

N° d’ordre : 3757






THÈSE

PRÉSENTÉE A

L’UNIVERSITÉ BORDEAUX 1

ÉCOLE DOCTORALE DES SCIENCES à renseigner

Par Roel DECKERS

POUR OBTENIR LE GRADE DE

DOCTEUR
SPÉCIALITÉ : Lasers, matière, nanosciences

LE RÔLE DES ULTRASONS, DE L’IRM ET DE L’IMAGERIE
OPTIQUE DANS LE CADRE DE L’ACTIVATION LOCALE
DE GÈNES ET DU DÉPÔT LOCAL DE MÉDICAMENTS


Directeur de recherche : Chrit Moonen




Soutenue le : 19 décembre 2008

Devant la commission d’examen formée de :

M. TANTER, Mickael Directeur de Recherche Inserm Rapporteur
M. TAVITIAN, Bertrand Chef de Laboratoire CEA (HDR) Rapporteur
M. BARTELS, Wilbert Associate Professor UMC Utrecht Examinateur
M. BRISSON, Alain Professeur UB1 Président de jury
M. VOISIN, Pierre Maître de Conférences UB2 Rapporteur de soutenance
M. MOONEN, Chrit Directeur de Recherche CNRS Examinateur



Université Bordeaux 1
Les Sciences et les Technologies au service de l’Homme et de l’environnement

Université Bordeaux 1
Les Sciences et les Technologies au service de l’Homme et de l’environnement

Part I. Introduction 7
Chapter 1. General introduction 9
References 11
Chapter 2. Ultrasound 13
2.1. Basics of ultrasound 13
2.2. Focused ultrasound 14
2.3. Interaction of ultrasound with tissue 15
2.4. Radiation force 20
2.5. References 21
Chapter 3. MRI 25
3.1. Introduction 25
3.2. Quantum-mechanical description 25
3.3. Classic-mechanical description 26
3.4. Relaxation and signal detection 27
3.5. Image acquisition 28
3.6. MR thermometry 29
3.7. Automatic control of temperature 31
3.8. References 32
Chapter 4. Optical imaging 35
4.1. Introduction 35
4.2. Origin of light 36
4.3. Interaction light-tissue 40
4.4. Light measurement 42
4.5. References 44
Part II. Spatio-temporal control of gene activation 47
Chapter 5. Regulatable gene expression systems 49
5.1. Introduction 49
5.2. References 51
3 Chapter 6. Heat shock proteins 53
6.1. Introduction 53
6.2. Hsp promoters in gene therapy 54
6.3. References 55
Chapter 7. In vitro characterization of Hsp70 promoter 57
7.1. Introduction 57
7.2. Materials & methods 60
7.3. Results 62
7.4. Discussion 67
7.5. Conclusions 69
7.6. Reference 70
Chapter 8. In vivo characterization of the Hsp70 promoter 73
8.1. Introduction 73
8.2. Material & methods 74
8.3. Results 76
8.4. Discussion 84
8.5. Conclusion 85
8.6. References 86
Chapter 9. Local gene activation using MR guided HIFU 89
9.1. Introduction 89
9.2. Material & methods 90
9.3. Results 93
9.4. Discussion 98
9.5. Conclusions 102
9.6. References 102
Part III. Local drug delivery 105
Chapter 10. The role of ultrasound and molecular imaging in local drug delivery 107
10.1. Introduction 107
10.2. Ultrasound facilitated local drug delivery 109
10.3. Imaging of drug delivery 111
4 10.4. References 117
Chapter 11. MRI monitoring of ultrasound mediated drug delivery 125
11.1. Introduction 125
11.2. Materials and methods 127
11.3. Results 130
11.4. Discussion 133
11.5. References 135
Part IV. Summaries and perspectives 139
Summary 141
Perspectives 145
Résumé 147
Perspectives 151
Part V. Word of thanks and publications 153
Word of thanks 155
List of publications 157
5
6 Part I. Introduction
7
8 Chapter 1. General introduction
Historically, image guided therapy has combined advances in imaging and therapeutic
technology to develop minimally invasive surgical and interventional techniques. Different
imaging modalities, mainly CT and ultrasound, but also MRI and PET/SPECT are used in the
preparation phase, during treatment and for post-treatment evaluation. The different imaging
modalities are increasingly used during minimally invasive procedures for real-time guidance
of instruments. Ultrasound is for example the primary modality for guiding radio-frequency
needles [1] and laser fibers used in tumor ablation [2]. Local injections of anti-inflammatory
drugs (steroids) for herniated disks [3] and the injection of blood cloth removal drugs are
performed with CT guidance [4]. The latest developments in image guided therapy such as
the fusion of imaging technologies with robotics and multi-modality imaging increase even
further the available information and the precision of the intervention [5,6].
MRI guided high intensity focused ultrasound (HIFU) is a completely non-invasive form of
image guided therapy and entered the clinical environment only recently for treatment of
uterine fibroids [7]. Further clinical trials are underway for treatment of cancers in breast,
liver and prostate [8-10].
New non-invasive image guided molecular therapies are evolving. These novel therapies use
molecular imaging techniques for dynamically monitoring of cellular function and molecular
processes in living animals [11]. Molecular imaging allows for (early) identification of
diseased tissue using imaging probes targeted to disease specific markers on cell membrane
[12,13] and tracking cell migration [14]. Combining molecular imaging with nanomedicine
can further improve the efficacy of molecular and cellular therapy. Monitoring of
nanomedicine such as genes, antibodies, chemotherapeutic drugs and drug carriers with
molecular imaging techniques gives insight in the local interaction of the drug with the tissue
and allows for the development of more specific nanomedicine. In this thesis two different
applications areas of image guided molecular therapy using MRI, HIFU and optical imaging
are exploited: local gene activation and local drug delivery.
Gene therapy is an experimental technique that uses genes to treat or prevent disease instead
of using drugs or surgery. With gene therapy it is possible to add a missing gene in the
genome and replace or repair a gene that malfunctions. Although gene therapy is a promising
treatment option for a number of diseases (including inherited disorders, some types of
cancer, and certain viral infections), the technique remains risky (e.g. auto-immune reponse to
9 introduced gene and toxicity of viral vector) and is still being investigated to make sure that it
will be safe and effective. One part of the study is based on the delivery of the therapeutic
gene. Autoimmune reactions, toxicity and the low efficacy of gene delivery are here the major
problems to conquer. Secondly, a tight control of gene activation is necessary, because the
objective of gene therapy is to express a therapeutic gene in the region where therapy is
required and for the duration necessary to achieve a therapeutic effect and to minimize
systemic toxicity. This implies the need for spatial and temporal control of gene activation in
vivo, which will be the subject of Part II in this thesis. Part II starts with an overview of the
different approaches for spatial and/or temporal control of local gene activation. In this thesis
local hyperthermia in combination with a temperature sensitive heat shock protein (Hsp)
promoter will be used for local gene activation. In Chapter 6 a biological background on heat
shock proteins is provided, including a discussion on the possible applications of the Hsp
promoter in gene therapy. In the following chapters the influence of different heating
protocols on the promoter activity are determined in order to be able to do treatment planning.
First the characteristics of the Hsp promoter are analyzed in vitro (Chapter 7) and then in vivo
(Chapter 8). Finally, we demonstrate in Chapter 9 the use of MR guided HIFU for the local
activation of a transgene.
Not only gene therapy, but also drug delivery may take advantage of the local deposition of
thermal and/or mechanical energy by means of ultrasound and this is the subject of Part III in
this thesis. In chemotherapy anti-cancer drugs are often administrated systemically resulting
in low concentration in the tumor, hence low treatment efficacy and significant toxic side
effects. Local increase of the cytotoxic drug concentration would ameliorate the efficacy of
the therapy and reduce systemic toxicity. Ultrasound can play an important role in facilitating
local drug delivery, which is explained in Chapter 10. Another important facet of local drug
delivery is monitoring non-invasively the drug’s pharmacokinetics and pharmacodynamics,
giving real time insight in the position and concentration of the drug and its influence on the
system. An overview of the different imaging modalities and their strengths and weakness are
also given in chapter 10. In Chapter 11 we demonstrate that we can improve the deposition of
a macromolecule in the liver with a clinical echograph and that we can follow the process of
delivery dynamically with a clinical MR scanner.
The thesis will begin with an introduction of the most important techniques (ultrasound, MRI
and optical imaging) used for the research per