Polymeric nanoparticles as original theranostic approach for alzheimer‟s disease

249 Pages·2017·7.59 MB·English

by  Davide Brambilla

Polymeric nanoparticles as original theranostic approach
for alzheimer￿s disease
Davide Brambilla
To cite this version:
Davide Brambilla. Polymeric nanoparticles as original theranostic approach for alzheimer￿s disease.
Human health and pathology. Université Paris Sud - Paris XI, 2012. English. ￿NNT: 2012PA114801￿.
￿tel-00692581￿
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Submitted on 1 May 2012
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UNIVERSITÉ PARIS-SUD 11
FACULTÉ DE PHARMACIE DE CHÂTENAY-MALABRY
ECOLE DOCTORALE :
INNOVATION THÉRAPEUTIQUE : DU FONDAMENTAL A L‟APPLIQUÉ
PÔLE : PHARMACOTECHNIE ET PHYSICO-CHIMIE
ANNÉE 2011-2012 SÉRIE DOCTORAT N° 1149
THÈSE
Présentée
À L‟UNITÉ DE FORMATION ET DE RECHERCHE
FACULTE DE PHARMACIE DE CHÂTENAY-MALABRY
UNIVERSITÉ PARIS-SUD 11
pour l‟obtention du grade de
DOCTEUR DE L‟UNIVERSITÉ PARIS-SUD 11
par
M. Davide BRAMBILLA
POLYMERIC NANOPARTICLES AS ORIGINAL THERANOSTIC APPROACH
FOR ALZHEIMER‟S DISEASE
soutenue le : 11 Janvier 2012
JURY : Prof. Seyed Moein Moghimi Rapporteur
Prof. Claus Michael Lehr Rapporteur
Prof. Charles Duyckaerts Examinateur
Prof. Patrick Couvreur Co-directeur de Thèse
Dr. Julien Nicolas Co-directeur de Thèse
Dr. Karine Andrieux Co-directeur de Thèse

Table of contents
Sommaire / Table of contents

Introduction générale / General introduction ..................................................................................... 7
Partie 1: Etude bibliographique / Part 1: Bibliographic study ................................................... 11
Nanotechnologies for Alzheimer’s disease: diagnosis, therapy and safety issues ............................ 13
Résumé / Abstract ........................................................................................................................................ 14
1. Introduction .............................................................................................................................................. 15
2. Imaging-based nanotechnologies for AD diagnosis ................................................................................. 16
3. Nanotechnologies for detection of AD biomarkers in biological fluids ................................................... 20
4. Nanotechnologies for AD treatment ......................................................................................................... 25
5. Nanotechnologies for in vitro evaluation of drugs activity ...................................................................... 39
6. Safety issues ............................................................................................................................................. 39
7. Conclusions .............................................................................................................................................. 42
8. Acknowledgements .................................................................................................................................. 42
9. References ................................................................................................................................................ 43
Partie 2: Travail expérimental / Part 2: Experimental work ...................................................... 53
Chapitre 1: Nanoparticules polymériques et barrière hématoencéphalique .................................... 55
Chapter 1: Polymeric nanoparticles and blood-brain barrier .......................................................... 55
Section 1 ....................................................................................................................................................... 59
Implications dans le projet NAD / Implications in the NAD project ........................................................... 59
Design of fluorescently tagged poly(alkyl cyanoacrylate) nanoparticles for human brain endothelial cells
imaging ......................................................................................................................................................... 61
Résumé / Abstract .................................................................................................................................... 62
1. Communication .................................................................................................................................... 62
2. Acknowledgments ................................................................................................................................ 68
3. References ............................................................................................................................................ 68
Supporting Information ............................................................................................................................ 69
Section 2 ....................................................................................................................................................... 83
Implications dans le projet NAD / Implications in the NAD project ........................................................... 83
Quantum dots-loaded PEGylated poly(alkyl cyanoacrylate) nanoparticles for in vitro and in vivo imaging 85
Résumé / Abstract .................................................................................................................................... 86
1. Introduction .......................................................................................................................................... 87
2. Experimental part ................................................................................................................................. 88
3. Analytical techniques ........................................................................................................................... 89
Table of contents
4. Results and discussion ......................................................................................................................... 91
5. Conclusions .......................................................................................................................................... 97
6. Acknowledgements .............................................................................................................................. 97
7. References ............................................................................................................................................ 98
Supporting Information .......................................................................................................................... 101
Section 3 ..................................................................................................................................................... 103
Implications dans le projet NAD / Implications in the NAD project ......................................................... 103
Fine evaluation of polymeric nanoparticles endocytosis and transcytosis across a human Blood-Brain
Barrier in vitro model ................................................................................................................................. 105
Résumé / Abstract .................................................................................................................................. 106
1. Introduction ........................................................................................................................................ 107
2. Experimental section .......................................................................................................................... 109
3. Results and discussion ....................................................................................................................... 114
4. Conclusions ........................................................................................................................................ 123
5. References .......................................................................................................................................... 124
Supporting Information .......................................................................................................................... 126
Conclusions et perspectives futures / Conclusions and future perspectives ................................... 127
Chapitre 2 : Interaction de nanoparticules polymériques avec le peptide Aβ ............................ 129
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Chapter 2: Polymeric nanoparticle interaction with Aβ peptide ................................................ 129
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Section 1 ..................................................................................................................................................... 133
Implications dans le projet NAD / Implications in the NAD project ......................................................... 133
A new method based on capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) to
monitor interaction between nanoparticles and the amyloid-β peptide ...................................................... 135
Résumé / Abstract .................................................................................................................................. 136
1. Introduction ........................................................................................................................................ 137
2. Experimental Section ......................................................................................................................... 139
3. Apparatus ........................................................................................................................................... 139
4. Methods ............................................................................................................................................. 140
5. Results and Discussion ...................................................................................................................... 143
6. Conclusions ........................................................................................................................................ 148
7. Acknowledgments .............................................................................................................................. 149
8. References .......................................................................................................................................... 149
Supporting Information .......................................................................................................................... 152
Section 2 ..................................................................................................................................................... 155
Implications dans le projet NAD / Implications in the NAD project ......................................................... 155
Table of contents
Colloidal properties of biodegradable nanoparticles influence interaction with amyloid-β peptide .......... 157
Résumé / Abstract .................................................................................................................................. 158
1. Introduction ........................................................................................................................................ 159
2. Results and Discussion ...................................................................................................................... 160
3. Conclusions ........................................................................................................................................ 164
4. Acknowledgments .............................................................................................................................. 164
5. References .......................................................................................................................................... 164
Section 3 ..................................................................................................................................................... 167
Implications dans le projet NAD / Implications in the NAD project ......................................................... 167
Polyethylene Glycol at the surface of nanocarriers alters Aβ peptide conformation: toward a nanomedicine
for Alzheimer‟s disease .............................................................................................................................. 169
Resumé / Abstract .................................................................................................................................. 170
1. Introduction ........................................................................................................................................ 171
2. Results and discussion ....................................................................................................................... 173
3. Conclusions ........................................................................................................................................ 185
4. Experimental section .......................................................................................................................... 186
5. Acknowledgements ............................................................................................................................ 193
6. References .......................................................................................................................................... 193
Supporting Information .......................................................................................................................... 197
Section 4 ..................................................................................................................................................... 199
Implications dans le projet NAD / Implications in the NAD project ......................................................... 199
Selegiline-Functionalized, PEGylated poly (alkyl cyanoacrylate) Nanoparticles: Investigation of Interaction
with Amyloid-β Peptide ............................................................................................................................. 201
Résumé / Abstract .................................................................................................................................. 202
1. Introduction ........................................................................................................................................ 203
2. Materials and Methods ....................................................................................................................... 205
3. Results and discussion ....................................................................................................................... 210
4. Conclusion ......................................................................................................................................... 217
5. Acknowledgments .............................................................................................................................. 218
6. References .......................................................................................................................................... 218
Conclusions et perspectives futures / Conclusions and future perspectives ................................... 221
Discussion générale / General discussion ........................................................................... 223
Conclusion générale et perspectives futures ...................................................................... 239
General conclusion and future perspectives ...................................................................... 239
Table of contents
Abbreviations / Abréviations............................................................................................... 243
Aknowledgments / Remerciements ..................................................................................... 247
Introduction générale
General introduction

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General introduction
Recent statistics indicate that 35 million people worldwide are affected by dementia with 4.6
million new cases per year (one new case every 7 seconds). In Europe there are 5 million cases of
dementia, 3 million of which are classified as Alzheimer's disease (AD). Given the continuing increase
in life expectancy, these numbers are expected to rise dramatically. In 2040, cases are expected to
double in Western Europe and to triple in Eastern Europe. Despite great progresses in research,
Alzheimer‟s disease etiology remains a big question and its treatment an important challenge of
modern medicine.
A major limitation to the development of active drug against Central Nervous System (CNS)-
related diseases is represented by the Blood-Brain Barrier (BBB), a formidable gatekeeper in the body
towards exogenous substances. Generally, pharmaceuticals including most small molecules do not
cross the BBB. Therefore, an ideal device for treatment of AD should provide two main properties: (i)
be able to cross the BBB and (ii) to exert a beneficial effect by inhibiting the AD pathogenesis.
In this view, an original solution could arise from nanotechnologies. Indeed, during the last
decades, nanotechnology applied to medicine, refereed as nanomedicine, showed interesting results for
the treatment of numerous severe diseases leading to the development and commercialization of more
efficient drugs.
This PhD thesis has been performed in the framework of the NAD (Nanoparticles for therapy
and diagnosis of Alzheimer's disease) European research project aiming to develop nanoparticulate
systems for the diagnosis and the therapy of Alzheimer's disease. The research is founded by the
European Union's 7th Framework Program and includes 19 European research centers (Grant
agreement no: CP-IP 212043-2 NAD). Prof. M. Masserini from University of Milano-Bicocca is the
scientific coordinator of the project.
The goal of this study was to create nanoparticlate systems able to cross the BBB in order to
reach the brain, the main site of AD, and to capture the Amyloid peptide, especially A species, the
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aggregates of which are known to play a pivotal role in the etiology of the disease.
In particular, the PhD was oriented toward the design of polymeric nanoparticles and the study
of their crossing of a novel in vitro BBB model (provided by P.O. Couraud, Institut Cochin) and to
study the interaction between the polymeric nanoparticles and the A peptide. Different kinds of
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nanoparticles have been designed during the project, mainly based on poly(alkyl cyanoacrylate)
(PACA) and presenting various surface properties, in order to increase their affinity towards the
peptide. All the obtained particles have been screened in vitro for their ability to capture the peptide.

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