pharmacotechnie et physico-chimie anné

249 Pages·2014·10.23 MB·English

by  Vianney Delplace

UNIVERSITÉ PARID-SUD 11

FACULTÉ DE PHARMACIE DE CHÂTENAY-MALABRY

ECOLE DOCTORALE : 425
INNOVATION THÉRAPEUTIQUE : DU FONDAMENTAL A L’APPLIQUÉ
PÔLE : PHARMACOTECHNIE ET PHYSICO-CHIMIE

ANNÉE 2014 – 2015 SÉRIE DOCTORAT N° 1300

THЀSE
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. Vianney DELPLACE
Titre de la thèse :
SYNTHESE DE POLYMERES VINYLIQUES PEGYLES DEGRADABLES PAR
POLYMERISATION RADICALAIRE CONTROLEE PAR LES NITROXYDES.

Soutenue le : 31 octobre 2014

JURY : Prof. Patrick Theato Rapporteur
Dr. Franck D’Agosto Rapporteur
Prof. Mathias Destarac Examinateur
Dr. Didier Gigmès Examinateur
Prof. Elias Fattal Examinateur
Dr. Didier Desmaële Examinateur
Prof. Patrick Couvreur Examinateur
Dr. Julien Nicolas Directeur de thèse
2
ACKNOWLEDGMENTS
First of all, many thanks should be addressed to Pr. Elias Fattal and Pr. Patrick
Couvreur for giving me the great opportunity to join the UMR CNRS 8612 and work in the
best conditions possible over the past three years and a half.
I would like to give very special thanks to my supervisor Dr. Julien Nicolas who offered
me a PhD position with trust and passed on all his knowledge. Julien, I certainly texted and
emailed you more than anybody else over the last three years, night and day, which made it
highly suspicious to my closest relations. You showed me the path to success and I’ll be
always grateful for that.
I would like to express my sincere gratitude to Pr. Patrick Theato and Dr. Franck
D’Agosto for having kindly accepted to review this manuscript, and Pr. Mathias Destarac for
joining as a member of the thesis jury.
I should also acknowledge our colleagues from the Institut de Chimie Radicalaire of
Aix-Marseille University, Dr. Yohann Guillaneuf, Dr. Didier Gigmès and the recently
promoted Dr. Antoine Tardy, who altogether made this project possible and pleasant. At both
the professional and personal levels, I wish this first collaboration will call some others.
The project associates from the Institut des Molécules et Matériaux du Mans of the
University of Maine, Dr. Sagrario Pascual, Pr. Laurent Fontaine and Dr. Hien The Ho, are
kindly acknowledged for sharing their experience on azlactone chemistry.
Many thanks to all the permanent members of the UMR CNRS 8612, among which
special thanks to Dr. Simona Mura for being so helpful anytime, Dr. Nicolas Tsapis for
trusting me the first and allowing me to join the group at the very beginning, and Dr Didier
Desmaële for being as nice and funny as good in organic chemistry (hexane d’Autriche
forever…).
Good science often requires meeting the good persons to do so. Among the people who
made my projects different and who generously shared their experience, I should mention and
3
especially thank Dr. Simon Harrisson for his support, his creativity and his duster coat. May
Slim Dusty be with you.
I would thank the students who had the courage to work under my supervision: Laëtitia
Korchia, Arlette El Asmar, Elise Guégain and Quentin Desguettes. Thank you for life lessons,
drinking, not drinking and looking like Julien Doré, respectively.
I would not have enjoyed these three years so much without “the crew”: Lilice Gaudin,
Mainro Canioni, Sabrinouille Valetti, Tanguy-Wesh Boissenot, Chaïch Leite, Nadou Handké,
Leticia “Sis” A. Santiago and Thomazzo Blino. So many nicknames for so many great people.
Romain, Tanguy, Thomas, thank you for having been the best buddies a man can dream of.
Thank you to Trung’s table, Hurling Pub’s pool, Resto U’s desserts, Beer Pong Bar’s
bathrooms, Piazza d’Oro’s cheap pizzas, Mad maker’s flaming shots, De Clercq’s French
fries and to the 350 Mystery Men from the past and the hundreds coming.
My last thought goes to the one I should be the most thankful to, for supporting me with
patience and affection all this time, my owl feather, Antonine.

4
SOMMAIRE

ACKNOWLEDGMENTS .............................................................................................................. 3
SOMMAIRE ................................................................................................................................... 5
ABBREVIATIONS ...................................................................................................................... 13
INTRODUCTION ........................................................................................................................ 17
STATE OF THE ART – BREAKING GOOD: (BIO)DEGRADABLE VINYL POLYMERS
FOR BIOAPPLICATIONS .......................................................................................................... 21
I. Introduction ....................................................................................................................... 23
II. Degradation of the polymer backbone ............................................................................ 24
1. Discrete insertion of main-chain degradable groups .................................................. 25
2. Multiple insertions of main-chain degradable functionalities ................................... 28
3. Copolymers with non-vinyl degradable blocks. ......................................................... 33
III. Degradation of the polymer side chain ....................................................................... 35
1. Degradation leading to water-solubility ...................................................................... 35
2. Cleavage triggering colloidal disassembly ................................................................. 36
IV. Connecting polymer chains via degradable junctions ................................................ 37
1. Degradable crosslinkers ............................................................................................... 37
2. Other strategies to insert degradable junctions ........................................................... 40
V. Cleavable conjugates ........................................................................................................ 41
1. The “grafting to” technique ......................................................................................... 41
2. The “grafting from” technique..................................................................................... 42
VI. Summary and outlook .................................................................................................. 43
SYNTHESIS TOOLBOX............................................................................................................. 57
I. Controlled radical polymerization ................................................................................... 57
II. Nitroxide-mediated polymerization................................................................................. 59
5
III. Radical ring-opening polymerization of cyclic ketene acetals .................................. 61
PREAMBLE – SCOPE AND LIMITATIONS OF THE NITROXIDE-MEDIATED
RADICAL RING-OPENING POLYMERIZATION OF CYCLIC KETENE ACETALS ..... 65
I. Introduction ....................................................................................................................... 67
II. Experimental Section ....................................................................................................... 69
1. Materials........................................................................................................................ 69
2. Polymerization .............................................................................................................. 69
3. Characterization ............................................................................................................ 69
4. Computational details ................................................................................................... 70
III. Results and Discussion ................................................................................................. 71
1. BMDO Polymerization ................................................................................................ 71
2. MPDL Polymerization ................................................................................................. 75
3. Block copolymerization ............................................................................................... 76
4. Mechanism and side-reaction ...................................................................................... 78
5. Conclusions ................................................................................................................... 88
CHAPTER 1: DEGRADABLE AND COMB-LIKE PEG-BASED COPOLYMERS BY
NITROXIDE-MEDIATED RADICAL RING-OPENING POLYMERIZATION .................. 93
I. Introduction ....................................................................................................................... 95
II. Experimental section ........................................................................................................ 99
1. Materials........................................................................................................................ 99
2. Analytical Techniques .................................................................................................. 99
3. Methods ....................................................................................................................... 100
a. Bulk Polymerizations. Synthesis of P(MeOEGMA-co-AN) Copolymer............ 100
b. Synthesis of P(MeOEGMA-co-S-co-CKA) Copolymer ................................... 100
c. Solution Polymerizations. Synthesis of P(MeOEGMA-co-AN) Copolymer ... 100
d. Synthesis of P(MeOEGMA-co-AN-co-CKA) Copolymer................................ 101
e. Hydrolytic Degradation ....................................................................................... 101
6
f. Cell Culture .............................................................................................................. 102
g. Cytotoxicity Studies ............................................................................................. 102
III. Results and discussion................................................................................................ 103
1. Bulk Copolymerizations ............................................................................................ 103
2. Solution Copolymerizations ...................................................................................... 104
a. Copolymerizations in Toluene ............................................................................ 104
3. Livingness ................................................................................................................... 109
4. Hydrolytic Degradation .............................................................................................. 110
5. Cytotoxicity Studies ................................................................................................... 113
6. Cell Morphology ........................................................................................................ 115
IV. Conclusion .................................................................................................................. 117
CHAPTER 2: NITROXIDE-MEDIATED RADICAL RING-OPENING
COPOLYMERIZATION: CHAIN-END INVESTIGATION AND BLOCK COPOLYMER
SYNTHESIS ............................................................................................................................... 125
I. Introduction ..................................................................................................................... 127
II. Experimental Section ..................................................................................................... 128
1. Materials...................................................................................................................... 128
2. Analytical Techniques ................................................................................................ 129
3. Methods ....................................................................................................................... 129
a. Synthesis of P(MeOEGMA-co-AN) Copolymer ............................................... 129
b. Synthesis of P(MeOEGMA-co-AN-co-CKA) Copolymer................................ 130
c. Chain Extension from P(MeOEGMA-co-AN-co-CKA) Macroinitiator .......... 130
d. Synthesis of P(MMA-co-AN-co-CKA)-b-PS Diblock Copolymer .................. 131
e. Synthesis of PS Homopolymer............................................................................ 131
f. Hydrolytic Degradation ........................................................................................... 131
III. Results and Discussion ............................................................................................... 131
IV. Conclusions ................................................................................................................. 139
7
CHAPTER 3: A SINGLE MONOMER TO CONTROL THE NITOXIDE-MEDIATED
POLYMERIZATION OF METHACRYLATES AND CONFER DEGRADABILITY ....... 145
I. Introduction ..................................................................................................................... 147
II. Experimental section ...................................................................................................... 148
1. Material ....................................................................................................................... 148
2. Analytical techniques ................................................................................................. 149
a. Nuclear magnetic resonance spectroscopy (NMR) ............................................ 149
b. Size exclusion chromatography (SEC) ............................................................... 149
3. Methods ....................................................................................................................... 150
a. Synthesis of poly[oligo(ethylene glycol) methyl ether methacrylate]
(PMeOEGMA)................................................................................................................ 150
b. Synthesis of poly[(oligo(ethylene glycol) methyl ether methacrylate)-co-(2-
methylene-4-phenyl-1,3-dioxolane)] (P(MeOEGMA-co-MPDL) .............................. 150
c. Synthesis of polystyrene (PS) .............................................................................. 151
d. Hydrolytic degradation ........................................................................................ 151
4. Cytotoxicity study ...................................................................................................... 151
a. Cell lines and cell culture .................................................................................... 151
b. Cytotoxicity assay ................................................................................................ 151
III. Results and discussions .............................................................................................. 152
IV. Conclusions ................................................................................................................. 157
CHAPTER 4: ONE-POT SYNTHESIS OF AZLACTONE-FUNCTIONALIZED SG1-
BASED ALKOXYAMINE FOR NITROXIDE-MEDIATED POLYMERIZATION .......... 161
I. Introduction ..................................................................................................................... 163
II. Experimental part ........................................................................................................... 165
1. Materials...................................................................................................................... 165
2. Analytical Techniques ................................................................................................ 165
3. Methods ....................................................................................................................... 166
8
a. Synthesis of azlactone-functional SG1-based alkoxyamine (AzSG1).............. 166
b. Determination of the dissociation rate constant (k ) .......................................... 167
d
c. Polymerization of styrene (S) .............................................................................. 167
d. Polymerization of n-butyl acrylate (nBA) .......................................................... 167
e. Copolymerization of methyl methacrylate (MMA) with a small amount of
acrylonitrile (AN) ........................................................................................................... 168
f. Conjugation of benzylamine to AzPMMA ............................................................ 168
III. Results and discussion................................................................................................ 169
1. Synthesis of azlactone-functionalized SG1-based alkoxyamine (AzSG1) ............. 169
2. NMP of vinyl monomers initiated by the AzSG1 alkoxyamine .............................. 171
a. NMP of styrene (S) .............................................................................................. 171
b. NMP of acrylic esters........................................................................................... 172
c. NMP of methacrylic esters .................................................................................. 174
3. Determination of the living chain fraction ................................................................ 176
4. Availability of the Az functionality for coupling ..................................................... 177
IV. Conclusion .................................................................................................................. 179
OPENING CHAPTER: RECENT TRENDS IN THE DESIGN OF ANTICANCER
POLYMER PRODRUG NANOCARRIERS ............................................................................ 183
I. Introduction ..................................................................................................................... 185
II. General considerations ................................................................................................... 186
III. The ‘conjugation to’ approach ................................................................................... 187
1. Conjugation to amphiphilic copolymers ................................................................... 189
a. PEG-polyester block copolymers ........................................................................ 190
b. Amphiphilic vinyl copolymers. ........................................................................... 197
c. Other amphiphilic copolymers. ........................................................................... 198
2. ‘Drug-induced’ self-assembly ................................................................................... 199
a. Polypeptide-based nanocarriers........................................................................... 200
9
b. Vinyl polymer-based nanocarriers ...................................................................... 203
c. Other systems ....................................................................................................... 206
3. The ‘conjugation through’ strategy ........................................................................... 207
4. The ‘drug-initiated’ method ....................................................................................... 208
a. Hydrophobic polymer prodrugs .......................................................................... 208
b. Amphiphilic polymer prodrugs ........................................................................... 211
IV. Conclusions ................................................................................................................. 212
GENERAL DISCUSSION ......................................................................................................... 221
I. Project overview and comments .................................................................................... 221
1. Picking the good strategy ........................................................................................... 221
2. First degradable PEG-based copolymers by NMP ................................................... 222
a. Literature and preliminary works ........................................................................ 222
b. Copolymerization of methacrylates and CKAs by NMP................................... 224
c. Chemical hydrolysis of MPDL-containing copolymers .................................... 226
3. Combining NMP and rROP: a new understanding by 31P NMR ............................ 227
a. MDO/BMDO copolymerization mechanism ..................................................... 228
b. MPDL, the reason of success .............................................................................. 229
4. Toward degradable controlling comonomers ........................................................... 230
5. In vitro study of the cytotoxicity ............................................................................... 231
6. Azlactone-functionalized polymers ........................................................................... 232
a. Alkoxyamine synthesis by the copper (0) method ............................................. 233
b. AzPolymer synthesis and limitations .................................................................. 234
c. Conjugation to azlactone-functionalized polymers ............................................ 234
II. Critical insight, limitations and possibilities ................................................................. 235
1. Biomedical applications: CLRPs vs the rest of the world ....................................... 235
2. Synthesis and limitations ........................................................................................... 236
10