Research topics


Multifunctional polymer nanoparticles for imaging and cell/molecular targeting

The medical application of nanotechnology, usually termed nanomedicine, has given a crucial impulse to the development of various types of drug-loaded nanocarriers, such as liposomes, nanoparticles, micelles, etc. In our group, a great deal of effort is focused on the engineering of biodegradable polymer nanoparticles able to serve as efficient diagnostic and/or therapeutic tools against severe diseases, such as cancer, infectious or neurodegenerative disorders.

Poly(alkyl cyanoacrylate) nanoparticles

Poly(alkyl cyanoacrylate) (PACA) nanoparticles have opened exciting perspectives for drug delivery due to their biodegradability and their non-toxicity. For instance, doxorubicine-loaded poly(isobutyl cyanoacrylate) nanoparticles have reached phase III clinical trials against primary liver cancer (LIVATAG®). However, the design of functionalized PACA nanoparticles for imaging and/or targeting is still very challenging due to the high reactivity of alkyl cyanoacrylate monomers [WIREs Nanomed. Nanobiotechnol. 2009]. The suceptibility to hydrolysis of PACA polymers also prevents efficient post-functionalization approaches.

Fluorescent PACA nanoparticles. In order to circumvent the drawbacks usually encountered with the use of small fluorescent dyes encapsulated into nanoparticles (e.g., leakage), rhodamine B-tagged poly(alkyl cyanoacrylate) amphiphilic copolymer nanoparticles have been prepared by tandem Knoevenagel condensation-Michael addition from the corresponding cyanoacetates derivatives [Chem. Commun. 2010]. They have been used for human brain endothelial cell imaging, allowing their uptake and intracellular trafficking (see video) to be finely observed. We also developped PEGylated PACA nanoparticles loaded with visible- and near-infrared-emitting quantum-dots (QDs) with high encapsulation yields, suitable for in vitro and in vivo imaging, respectively [Soft Matter 2011].

Interaction with the amyloid-β 1-42 peptide. To detect interaction between functionalized nanoparticles and the amyloid-β 1-42 peptide (Aβ1-42), an important biomarker of the Alzheimer's disease (AD), we developped a reliable analytical tool based on capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) [Anal. Chem. 2010]. Unexpectedly, we discovered that the PEG corona of long-circulating PACA nanoparticles favors interaction with Aβ1-42 both in solution and in serum [ACS Nano 2012a]. In silico and modeling experiments highlighted the mode of PEG interaction with the Aβ1-42 peptide and its conformational changes at the nanoparticle surface (see figure). These nanoparticles might act as a peptide sequester in the bloodstream, carrying the peptide to the liver where it could be enzymatically cleaved and degraded.

Targeted PACA nanoparticles. We also designed a versatile and ligand-functionalized PACA nanoparticulate platform by 'click' chemistry [Macromolecules 2008], gathering all together crucial features required for active targeting and drug delivery (i.e., biodegradable, PEGylated, fluorescent and targeted). The multifunctional nanoparticles were successfully used to target two major pathologies; namely cancer and Alzheimer's disease, via their functionalization by appropriate biologically active ligands [ACS Nano 2012b]. Against cancer cells, we used biotin to target biotin receptor that are overexpressed at the surface of many cancer cells (see figure), whereras Aβ1-42 peptide was targeted by means of curcuminoid derivatives and an anti-Aβ1-42 peptide antibody. The antibody-decorated nanoparticles exhibited higher affinity toward Aβ1-42 species comparatively to other kinds of colloidal systems and led to significant aggregation inhibition and toxicity rescue of Aβ1-42 at low molar ratios.

Polyester nanoparticles

Polyesters are biodegradable FDA-approved polymers for use in Humans and have been widely used to prepare nanocarriers for drug delivery applications. However, the accurate functionalization of polyester nanoparticles to achieve active targeting is not straightforward and generally not orthogonal, which could result in multisite attachment [Chem. Soc. Rev. 2013].

In collaboration with Sanofi, we have been interested in applying 'click' chemistry to long-circulating poly(lactic acid)-block-poly(ethylene glycol) (PLA-b-PEG) copolymer nanoparticles (see figure) [WO 2013127949]. This strategy has been illustrated by the preparation of a large library of different nanoparticles, such as, ligand-decorated nanoparticles (with biotin, folic acid or anisamide), fluorescent nanoparticles (UV-Vis or near-infrared dyes), and multifunctional nanoparticles decorated with a targeting ligand and a fluorescent probe [Chem. Mater. 2014]. Optimal ligand availability was determined by surface plasmon resonance and successful targeting was demonstrated by in vitro experiments on different cancer cell lines.


Controlled radical polymerization: mechanism and synthesis of biomaterials

Controlled radical polymerization (CRP) techniques, such as nitroxide-mediated polymerization (NMP), atom-transfer radical polymerization (ATRP) and reversible addition-fragmentation chain transfer (RAFT) polymerization, enable the synthesis of well-defined, complex and functional macromolecular architectures. We are excited to explore new areas of CRP techniques and to take the most of them to design innovative structures and novel biomaterials.

Nitroxide-mediated polymerization

Our group has a strong expertise in CRP techniques and especially in nitroxide-mediated polymerization (NMP) [Prog. Polym. Sci. 2013]. Among our different achievements in this field, we proposed an simple yet efficient method for the synthesis of alkoxyamine initiators mediated by copper metal [Polym. Chem. 2010] and reported optimized conditions for the successul NMP of isoprene [Macromolecules 2011]. We are also very interested by the NMP of methacrylic esters through the copolymerization approach. It consists in adding a very small amount of a comonomer such as styrene (typically 4-9 mol.%) during the NMP of methacrylic esters to obtain all the features of a controlled/living system [Macromolecules 2009]. We have extended this concept to other monomers such as oligo(ethylene glycol) methacrylate [Macromolecules 2008] to prepare PEG-based materials and also discovered that acrylonitrile can be used an efficient 'controlling' comonomer [JPSA 2010].

Controlled radical ring-opening polymerization

The crucial need for more environmentally friendly plastic materials is currently stimulating the development of degradable vinyl polymers [Nature Chem. 2015]. A significant part of our research efforts is also focused on controlled radical ring-opening polymerization (rROP) as a way to produce (bio)degradable vinyl polymers for biomedical applications and sustained development [Chem. Rev. 2017]. In this context, we reported the first synthesis of degradable PEG-based copolymers by NMP [Biomacromolecules 2013] using different cyclic ketene acetals (CKA) as precursors of main-chain labile moieties. We have shown that the structure of the CKA is a crucial parameter to reach high monomer conversion, good control of the polymerization and adjustable insertion of CKA in the copolymer. This resulted in tunable degradation of the copolymer, up to complete degradation for a sufficient amount of inserted CKA. The copolymers were show to be not cytotoxic. We also extended this approach to the deisgn of block copolymers with a degradable block [Macromol. Rapid Commun 2014], to the synthesis of dedradable, alternating copolymers based on the RAFT copolymerization between CKA and maleimides [ACS Macro. Letters 2017]. In collaboration with the group of Didier Gigmes, we also developped a new copolymerization system between CKA and vinyl ethers to afford degradable, functional polymers for various applications, including functional polymers, PEGylated nanoparticles, antibacterial surfaces and bioelastomers [Angew. Chem., Int . Ed. 2017].

Polymer-protein/peptides bioconjugates by CRP

The design of polymer-protein/peptide bioconjugates by CRP techniques is an important area of research [Polym. Chem. 2010]. NMP is perhaps the most adapted technique for biomedical applications due to its simplicity and the absence of toxic reagents. In this view, we reported the synthesis of innocuous PEG-based copolymers in water-rich solutions intended to be used for bioconjugation purposes [Macromolecules 2010]. N-succinimidyl ester-functionalized PEG-based copolymers were then linked to various amine-containing substrates such as a neuroprotective tripeptide and lysozyme [Polym. Chem. 2011]. This represents the first example of peptide/protein PEGylation by the NMP technique. Azlactone-functionalized SG1-based alkoxyamine for NMP and protein bioconjugation has also been developped by our group [Macromolecules 2015].


Prodrug nanoparticles

Prodrugs are inactive, bioreversible derivatives of active drug molecules that must undergo an enzymatic and/or chemical transformation in vivo to release the active parent drug. Inspired by this concept, we are currently working on the design of innovative polymer prodrug nanoparticles to tackle the common limitations of traditional nanoparticulate systems in which drugs are physically encpasulated (e.g., burst release, poor drug loadings, etc.) [Polym. Chem. 2014].

Squalene-based prodrug nanoparticles

A major breakthrough in this field has recently been reported by our group and was termed 'squalenoylation' . It consists in the coupling of squalene (Sq), a natural lipid, to a variery of different drugs to prepare high drug loading squalene-drug nanoparticles by self-assembly in aqueous solution of the corresponding conjugates. Among the different systems, Sq-gemcitabine (Gem) nanoparticles afforded remarkable anticancer activity against many different solid tumors in vivo. In the past few years, we focused our efforts on the PEGylation of Sq-Gem nanoparticles to confer them with long-circulating abilities [Adv. Funct. Mater. 2008]. Also, we developped the first Sq-based multifunctional system (i.e., therapeutic, traceable and targeted) by a simple co-self-assembly of the corresponding building blocks (i.e., Sq-Gem, Sq-dye and Sq-ligand), that demonstrated enhanced uptake by cancer cells [Chem. Commun. 2014].

Polymer prodrug nanoparticles

We are also very interested in the design of polymer prodrug nanoparticles as a way to combine both the advantages of the prodrug strategy and the flexibility offered by macromolecular synthesis. We recently developped a new concept termed "drug-initiated" method [Chem. Mater. 2016] which consists in the synthesis of well-defined drug-polymer amphiphiles by CRP of a hydrophobic monomer from an hydrophilic drug [Angew. Chem., Int. Ed. 2013]. The flexibility of this approach was illustrated by the synthesis of different polymer promoeties such as polyisoprene (isoprene being the basic structural motif of many biocompatible natural terpenes) or a naturally occuring lipid-containing polymethacrylate by either NMP or RAFT polymerizations [Biomacromolecules 2013]. Biological evaluations successfully demonstrated both the in vitro and in vivo anticancer activity on solid pancreatic tumors in mice [Chem. Mater. 2014]. This approach was applied to other drugs like cladribine [Chem. Mater. 2016], and to a fluorescent dye [Chem. Commun. 2017] for the preparation of composite nanoparticles by co-nanoprecipitation.

Collaborations


International

  • Pr. Massimo Masserini. Department of Experimental Medicine, Univ of Milano-Bicocca, Italy
  • Pr. Francesco Nicotra. Department of Biotechnology and Biosciences, Univ of Milano-Bicocca, Italy
  • Pr. Moein Moghimi. School of Pharmaceutical Sciences, Univ of Copenhagen, Danemark
  • Pr. Wiep Scheper. AMC Amsterdam, Univ of Amsterdam, The Netherlands
  • Pr. Christophe Detrembleur. Center for Education and Research on Macromolecules (CERM), Univ de Liege, Belgium
  • Dr. Jorge F. J. Coelho. Department of Chemical Engineering, Univ of Coimbra, Portugal
  • Pr. Guido Ennas. Dipartimento di Scienze Chimiche, Univ Cagliari, Italy
  • Pr. Krzysztof Matyjaszewski. Department of Chemistry, Carnegie Mellon Univ, USA
  • Pr. David M. Haddleton. Department of Chemistry, Univ of Warwick, UK
  • Pr. Steve P. Armes. Department of Chemistry, Univ of Sheffield, UK

National

  • Drs. Yohann Guillaneuf and Didier Gigmes. ICR, UMR CNRS 7273, Aix-Marseille Univ, Marseille
  • Dr. Didier Letourneur. LVTS, INSERM U1148, CHU Xavier Bichat, Univ Paris Nord, Paris
  • Drs. Valérie Langlois and Benjamin Le Droumaguet. ICMPE, UMR CNRS 7182, Univ Paris-Est, Thiais
  • Pr. Laurent Fontaine and Dr. Sagrario Pascual. Institut des Molécules et des Matériaux du Mans, UMR 6283 CNRS, Université du Maine, Le Mans
  • Pr. Bernadette Charleux. Laboratoire de Chimie Catalyse Polymères et Procédés (C2P2), UMR CNRS 5265, CPE Lyon, Univ Lyon 1, Villeurbanne
  • Pr. Erwan Poupon. BioCIS, UMR CNRS 8076, Univ Paris-Sud, Châtenay-Malabry
  • Drs Benoit Dubertret and Thomas Pons. Laboratoire d'Optique Physique, ESPCI, Paris
  • Dr. Anne Valérie Ruzette. Laboratoire Matière Molle et Chimie, UMR ESPCI-CNRS 7167, ESPCI, Paris

Industrial

  • Sanofi. Drs Didier Bazile (Global Head of Drug Delivery Technologies and Innovation) and Harivardhan Reddy Lakkireddy (Head of Drug Delivery Technologies and Innovation)
  • Sanofi. Dr Dominique Lesuisse (Head of the ‘Aging’ Department)
  • LFB. Drs Patrick Santambien (Director of Technological Innovation) and Sami Chtourou (Executive vice-president in charge of innovation and scientific affairs)
  • Arkema. Drs Laurence Couvreur (Research chemist) and Stéphanie Magnet (Research chemist)
  • Unilever. Dr. Ezat Khoshdel (Senior scientist)

Facilities


Chemical characterization

  • Nuclear Magnetic Resonnance (NMR)

- Avance 400 MHz (H, C, N) spectrometer (Bruker)
- Avance 300 MHz (H, C) spectrometer (Bruker)
- ARX 200 MHz (H, C, F, P) spectrometer (Bruker)

  • Fourier Transform Infrared (FT-IR)

- Spectrum Two FT-IR spectrometer with UATR module (Perkin Elmer)
- Vector 22 spectrometer (Bruker)

  • UV-Vis and fluorescence spectrometry

- LS 50B fluorescence spectrometer (Perkin Elmer)
-
Lambda 25 UV/Vis spectrometer (Perkin Elmer)

  • High performance liquid chromatography (HPLC)

- 4 complete HPLC systems (isocratic pump, 2-channel pump and 4-channel pumps) with UV and fluorescence detectors (Waters)

  • Misceallenous

- 3 R-210 Rotavapors equipped with a V-700 vacuum pump, a V-850 vacuum controller and a F-105 recirculating chiler (Büchi)
- DR 3003 heated vacuum dessicator (Vinci Technologies)

  • In-campus mass spectrometry platform (LC-MS and GC-MS)

- LTQ Orbitrap Velos Pro (Thermofischer scientific)
- ITQ900 (Thermofischer scientific)
- Quattro Ultima (Waters)
- Esquire-LC (Bruker)


Polymer characterization

  • Size exclusion chromatography (SEC)

- Complete, temperature controlled, quadriple-detection (RI, viscometer, laser light scattering LALLS/RALLS and UV) GPC/SEC system in DMAc (Viscotek)
- Temperature controlled, dual detection (RI and UV) GPC/SEC system in chloroform (Waters/Viscotek)

  • Differential Scanning Calorimetry (DSC)

- DSC 7 differential scanning calorimeter (Perkin Elmer)


Nanoparticle formulation and characterization

  • Formulation

- DVX-2500 digital multi-tube vortexer (VWR)
- Optima LE-80K ultracentrifuge (Beckman Coulter, Inc)
- Branson 5200 ultrasonic bath (Bransonic)
- Alpha 1-2 LD Plus freeze dryer (Christ)

  • Dynamic light scattering (DLS)

- Zetasizer Nano ZS (Malvern)
- Mastersizer 2000 (Malvern)
- Ultracentrifuge (Beckman Coulter, Inc)

  • Near-campus facilities

- Scanning Electron Microscopy (SEM)
- Atomic Force Microscopy (AFM)
- Transmission Electron Microscopy (TEM), TEM after freeze-fracture and Cryomicroscopy (Cryo-TEM)
- SOLEIL synchrotron (SAX, WAX)


Biological evaluations

  • Flow cytometry (FACS)

- Accuri C6 flow cytometer (BD Biosciences)

  • In-campus imaging platform

- Confocal microscopy (Zeiss LSM 510 Meta confocal microscope equipped with different argon and helium lasers, LSM 510 for image analysis and treatment, attofluor cell chamber, cell culture facilities, etc.)
- Videomicroscope Axio-Observer Z1 (Zeiss)

  • In-campus proteomic platform

- Analysis of proteins by bi-dimensional electrophoresis, MALDI-ToF mass spectrometry and Nano-LC

  • Surface plasmon resonnance (SPR)

- BIAcore T100 (GE Healthcare Life Sciences)

  • Isothermal titration calorimetry (ITC)

- VP-ITC system (MicroCal)

  • Cell culture department

Modern cell culture department containing all necessary immortalized and primary cell lines such as healthy cells (e.g., J774, NIH/3T3, HUVEC, hCMEC/D3, etc.) and cancer cells (e.g., KB-3-1, PC3, MiaPaca, HeLa, M109, MCF-7, A549, P388, L1210, CCRF-CEM, SH-SY5Y, NIH OVCAR-3, Panc-1, etc.) and in which advanced methodologies are routinely performed: toxicity assays (e.g., MTT, MTS, LDH, etc.), cell cycle (flow cytometry), apoptosis studies, etc. Molecular Biology and transcription regulation studies include: genomic footpriniting in vivo or in vitro, reporter genes (CAT and luciferase), PCR, RT-PCR and Q-PCR, Northern blots, Nuclear run on, Southern blots, bandshifts, gene arrays, nucleosome positioning, studies by micrococcal nuclease digestion, chromatin remodeling study by DNASe I hypersensitivity, enzymatic digestions, cloning, sub-cloning, CHIP (chromatin immunoprecipitation). Concerning cellular imaging, the Institute is equipped with optical microscopes and video cameras. HES coloration, immunochemistry (paraffin embedded tissues or tumors) and TUNEL analysis are also available.

  • Radioactivity department

Our institute has a radioactivity department with all the facilities to perform pharmacokinetic and tissue distribution studies (e.g., scintillation counting of tissues, beta imager, radioactivity scan of electrophoresis gels, etc.).

  • Laboratory animal facilities

An in-campus modern animal facility is at the disposal of our group. It is conform to the French and European ethical rules concerning animal experimentations. It contains up to ca. 4,000 animals including mice, rats and rabbits, and also handles 40 strains of transgenic mice.

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