Doc and Post-Doc Day Institut Galien - Paris-Sud th

Monday, 20 June 2016 9h – 18h

Sponsored by :

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Organizing Committee: Claire Albert Corentin Berardet Alexandre Bordat Jean-Baptiste Coty Elise Guegain Sophie Houvenagel Mathilde Lorscheider Julien Massiot Arnaud Peramo Marion Quaillet Marie Rouquette Coralie Ruel Johanna Tran

With great help of: Elias Fattal Dominique Martin Sylvie Zemmour

Sponsored by:

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Agenda 9h.

Opening. Prof. Elias Fattal – Director of Institut Galien Paris-Sud

Chairs: Felix Sauvage and Jean-Baptiste Coty 9h20-10h20

Dr. Romain Verpillot Alzohis, Paris (France)

Oral Presentations: Drug Delivery Chairs: Mathilde Lorscheider and Marion Quaillet 10h20-10h40

Jiao Feng Development of novel nano-biotechnology tools targeting opioid receptors

10h40-11h00

Poster flash presentation

11h00-11h30

Poster session and coffee break

11h30-11h50

Claire Albert Emulsion stabilized with biodegradable PLGA nanoparticles for pharmaceutical and cosmetic applications

11h50-12h10

Elise Guegain Design of Degradable Polymer Prodrugs by Nitroxide-Mediated Radical Ring-Opening Polymerization from the Drug-Initiated Method

12h10-12h30

Clémence N'Guessan Gnaman A promising vaginal gel containing lactobacillus crispatus for the prevention of the infection by neisseria gonorrhoeae

12h30-13h30

Lunch break

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

13h30-14h00

Poster session Chairs: Felix Sauvage and Jean-Baptiste Coty

14h00-15h00

Dr. Catherine Le Visage Laboratoire d'Ingénierie Ostéo-Articulaire et Dentaire (LIOAD), Inserm U791, Nantes (France)

Oral Presentations: Analysis, Diagnostic and Theranostic Chairs: Alexandre Bordat and Daniele Vinciguerra 15h00-15h20

Elodie Millart Lipid Janus nanoparticles with superparamagnetic compartment: from the implementation process to theranostic applications

15h20-15h40

Sophie Houvenagel Nanocapsules made of fluorinated polymers for tumor imaging by ultrasonography.

15h40-16h00

Poster Session and coffee break

16h00-16h20

Jeanne Bataille Toward the development of miniaturized analytical tool for the diagnosis of transthyretin familial amyloïdosis

16h20-16h40

Jean-Baptiste Coty Activation of complement system by nanomedicines: development of a high throughput method and determination of an indicator of activation

16h40-17h10

30 years of Institut Galien into a movie

17h10

Ending. Award for the best oral and poster communication

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Septembre 1985

Préambule du dossier déposé auprès du CNRS pour la création de l'unité « Au cours des dernières années, les progrès réalisés dans la connaissance de la physiopathologie ont conduit à de remarquables innovations thérapeutiques. Parallèlement, dans les grands secteurs de la Pharmacologie, les recherches ont souvent permis de préciser les modalités d’interactions entre les principes actifs et leur site d’action biologique ; de plus, le développement de la Biologie moléculaire a récemment autorisé l’identification de nouvelles cibles. Des processus originaux de création des médicaments ont ainsi été élaborés et ont permis de concevoir de nouveaux outils thérapeutiques. La façon de donner n’étant pas moins importante que ce que l’on donne (selon Saint Augustin), l’ensemble des observations précédentes fait clairement apparaître l’impérieuse nécessité d’innover aussi dans la façon d’administrer les médicaments, c'est-à-dire dans le domaine de la Pharmacotechnie : mise au point de vecteurs conduisant sélectivement les molécules actives dans la cible à atteindre ; recherche de nouveaux matériaux et de procédures rationnelles permettant d’optimiser la formulation ; conception et création de systèmes de délivrance originaux améliorant la biodisponibilité des médicaments. Dans la plupart des pays, l’innovation pharmacotechnique s’est considérablement développée au cours des dernières années. En France, elle a pris un retard certain qui, sans le soutien des Grands Organismes de Recherche risque de s’accentuer ». « Formée de Physicochimistes (ex ER 99 CNRS) et de spécialistes de Pharmacotechnie et de Biopharmacie (Laboratoire B1*), la formation souhaite aborder l’innovation pharmacotechnique selon une conception originale, en fondant ses travaux sur une double interface : - interface Physicochimie-Pharmacotechnie apportant l’assise fondamentale nécessaire à toute efficacité créatrice en pharmacotechnie ; - interface Pharmacotechnie-Biologie et Clinique, nécessaire à l’inventaire des besoins en thérapeutique et à l’évaluation de toute innovation pharmacotechnique. La première interface est assurée, au sein même de la forrmation, par la présence de Physicochimistes des surfaces et par celle de spécialistes de Pharmacotechnie et de Biopharmacie. La seconde interface sera envisagée, opération par opération, en recourant ponctuellement à des collaborations extérieures, notamment à celles que la formation s’est déjà assurée auprès du monde scientifique et médical ».

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Invited Speakers Presentations Dr. Romain Verpillot & Dr. Catherine Le Visage

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Dr. Romain Verpillot

Romain Verpillot is the CEO and co-founder of Alzohis diagnosis, a young innovative biotechnology company created in 2014, and which provides a new molecular diagnostic test for Alzheimer’s disease. After a Master’s degree at Université de Montpellier II, in biomolecules chemisty, R. Verpillot performed his PhD at Université Paris Sud in the Institut Galien Paris Sud between 2007 and 2010. He was a part of the team 4, “Proteins and Nanotechnology in Analytical Science”, under the supervision of Myriam Taverna. His work focused on the development of a novel diagnostic tool for Alzheimer disease. After graduation, he developed his managerial and entrepreneurial skills through different formations given by HEC, EM Lyon, Incuballiance, as well as a Ecole Centrale Paris. All along the development of his project, Romain Verpillot was laureate of several innovation prizes (C’Nano in 2011, Oséo in 2012, 1st “business model prize” for 2011-2012 HECChallenge+, Scientipôle initiative in 2013, and “création développement” from Bpifrance in 2014). All these encouragement led Dr. Romain Verpillot to found the company Alzohis, in March 2014, together with Pr. Myriam Taverna and Rudy Beaujean. As a CEO, he is notably in charge of strategy, partnership and financial aspects of the company.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Dr. Catherine Le Visage

Catherine Le Visage is a Research Director at the French National Institute of Health and Medical Research, in the University of Nantes, she is working in a group focusing on Biomaterials for Regenerative Medicine. She was first trained as a Pharmacist and she received, in 1999, her PhD in Pharmaceutical Technologies from the University of Paris 11; under the supervision of Patrick Couvreur, at that time director of the so-called URA-1218. Her PhD topic was related to polymer nanoand microparticles, especially in order to induce immune tolerance through oral delivery. She then performed a post-doctoral training at the Johns Hopkins School of Medicine (Baltimore, USA) in the Biomedical Engineering Department, where she was appointed as a Research Associate in 2003. Her research focused on tissue engineering projects, such as intervertebral disc regeneration and tracheal replacement. She joined the French National Institute of Health and Medical Research (Inserm) as a tenured Research Scientist in 2007 to investigate chemically cross-linked polysaccharides hydrogels as platforms for regenerative medicine. The driven point of her research was the versatile use of those solid hydrogels, with a wellcontrolled size and shape, using polysaccharides as building block which enables biodegradability and manipulation in aqueous conditions. Currently, Dr Le Visage’s interest lies in developing these biomaterials for enhanced bone repair, with a focus on long-term delivery of biochemical signals and addition of substituted nanoparticles. In 2016, Dr. Catherine Le Visage becomes the new president of the scientific council of la Fondation de l’Avenir, organization that aims to promote medical research, together with surgeons, by providing fundings to selected projects (up to 40 projects per year) on specific topics.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Oral Presentations: Drug delivery

Jiao Feng Development of novel nano-biotechnology tools targeting opioid receptors Claire Albert Emulsion stabilized with biodegradable PLGA nanoparticles for pharmaceutical and cosmetic applications Elise Guegain Design of Degradable Polymer Prodrugs by Nitroxide-Mediated Radical Ring-Opening Polymerization from the Drug-Initiated Method Clémence N'Guessan-Gnaman A promising vaginal gel containing lactobacillus crispatus for the prevention of the infection by neisseria gonorrhoeae

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Development of novel nano-biotechnology tools targeting opioid receptors Jiao Fenga, Sinda Lepetrea, Patrick Couvreura a

UMR-CNRS 8612, Faculté de Pharmacie, Université Paris-Sud, ChâtenayMalabry

Pain is a major health problem that significantly affects the quality of life of the patients. Some neuropeptide pharmaceuticals, have a great potential for the treatment of pain, but the blood-brain/spinal cord barrier (BBB／BSCB) limits their entry into the central neural system (CNS), where the vast majority of relevant receptors are located. Additionally, the half-lives of those peptides in plasma are very short. The bioavailability of the peptide-based drugs to the brain and spinal cord is therefore limited, due to poor metabolic stability, and/or inability to cross these barriers (because of their high hydrophilicity). What’s more, there also exist opioid receptors in peripheral nervous system, especially under inflammation condition, where the opioid receptors are up-regulated[1]. But the harsh environment (low pH, enzymes…) restricts the effect of neuropeptides on those sites.

The aim of this project is to create new nanomedicines targeting the pain, using squalene(SQ)[2]. By linking the neuropeptides with SQ, the corresponding bioconjugates have the ability of self-assembling into nanoparticles in water, with sizes around 100nm. The drug loading is much higher compared with other nano carriers, around 60%. Under this nanoparticle form, the peptides are protected from metabolism. And the free peptides can be released from the NPs in serum with sensitive linker compared with amide bond or diglycolic bond. The further pharmacological effect will be tested. References [1] Christoph Stein et al, Attacking pain at its source: new perspectives on opioids, NATURE MEDICINE, 2003. [2] Patrick Couvreur et al, Squalenoyl Nanomedicines as Potential Therapeutics, NANO LETTERS, 2006.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Formulation of emulsions stabilized with biodegradable PLGA nanoparticles: co-encapsulation of two active ingredients C. Albert1, N.Huang1, N.Tsapis1, V. Nicolas2, L. Michel3, G. Mekhloufi1, E.Fattal1, F. Agnely1 1

Institut Galien Paris-Sud, UMR 8612 CNRS, Univ. Paris-Sud, Université Paris-Saclay, Faculté de Pharmacie, 92296 Châtenay-Malabry, 2 Microscopy facility, FR141-IPSIT, Univ. Paris-Sud, Université Paris-Saclay, Faculté de Pharmacie, 92296 Châtenay-Malabry, France 3 INSERM U976, Pavillon Bazin, Hôpital Saint-Louis, 75010 Paris, France

The main objective of this study is to formulate, for therapeutic or cosmetic purposes, an emulsion stabilized by biodegradable and biocompatible nanoparticles. Indeed, our nanoparticles (NPs) of poly (lactic-co-glycolic) acid (PLGA) allow us to formulate long-term stable, biodegradable and biocompatible emulsions with the advantage of being potentially less toxic and irritant compared to emulsions stabilized with synthetic surfactants. Furthermore, it is possible to co-encapsulated two active ingredients in the same formulation: a first one in the NPs and a second one in the oil droplets. Co-encapsulation leads to an improvement of the observance of the patient and can lead to a synergistic effect between the two active pharmaceutical ingredients. To clarify the mechanisms and the kinetics of stabilization with PLGA nanoparticles, a thorough physical chemistry study is performed. The interface between the oil and the aqueous phase is characterized using interfacial rheology, interfacial and surface tensions, nanoparticles adsorption and three-phase contact angle measurements. A stability study of the emulsions is also performed, as well as a study of the contribution of the polymer stabilizing the NPs on the emulsion stability. A second part of the study consists in the co-encapsulation of the two active ingredients in the emulsion. The kinetics of release and the cutaneous penetration of the active ingredients, as well as the cutaneous penetration of the nanoparticles, are assessed. Two types of PLGA nanoparticles (NPs) are used in this study: PLGA-PVA NPs with Poly(Vinyl Alcohol) (PVA) as polymer stabilizing the nanoparticles and PLGA NPs without any other polymer or surfactant. The emulsions obtained with both NPs are radically different: for example, a 10% w/w oil phase leads to a much thicker emulsion phase with PLGA NPs than with PLGA-PVA NPs. The emulsions stabilized with PLGA-PVA NPs are also less stable than those stabilized with PLGA NPs which are stable for at least 55 days. These Pickering emulsions using PLGA nanoparticles are indeed very promising systems for the formulation of stable and bi-compartmentalized emulsions.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

The Drug-Initiated strategy: A convenient way to design polymer nanoparticles for anticancer therapy Elise Guégain, Johanna Tran, Quentin Deguettes, Julien Nicolas Institut Galien Paris-Sud, UMR CNRS 8612, Université Paris-Sud, Faculté de Pharmacie, 5 rue Jean-Baptiste Clément, 92290 Châtenay-Malabry, France

Drug-loaded polymer nanoparticles hold great hope for the treatment of severe diseases including cancer. However, important limitations still remain (e.g., burst release, poor drug loading). Polymer prodrug nanoparticles, for which the drug is covalently linked to a polymer scaffold, can circumvent those limitations.1 Among the different strategies, the ‘druginitiated’ approach2 is particularly promising and little-studied compared to the others. In this strategy, the drug is covalently linked to an initiator and a polymer chain is grown from it, leading to a polymer bearing one drug at the chain-end. By combining controlled radical polymerization (CRP) techniques and the prodrug approach, the ‘drug-initiated’ strategy was successfully applied to hydrophilic gemcitabine (Gem) as the anticancer drug, and isoprene (I) as hydrophobic vinyl monomer.3 The resulting well-defined Gem-PI conjugates led to narrowly dispersed nanoparticles upon nanoprecipitation with significant anticancer activity in vivo. Our aim was to develop a strategy to confer degradability to similar polymer prodrugs through the use of the copolymerization between 2-methylene-4-phenyl-1,3-dioxolane (MPDL), a cyclic ketene acetal monomer which is a precursor of main-chain ester moieties and methacrylic esters;4 either methyl methacrylate (MMA) or PEG methacrylate (PEGMA) to yield conjugates of different self-assembling properties in water. PEGylated prodrug nanoparticles Hydrophilic

Hydrophilic

rROP MPDL

Hydrophilic Gem

Nanop°

PEGMA

Hydrophobic

MPDL Hydrophobic

?

PEGMA Hydrophilic

PEGylated prodrug

Hydrophilic

rROP

Nanop°

MPDL

Hydrophilic Gem

MMA

MPDL Hydrophobic

Hydrophobic chain

MMA Hydrophobic

Prodrug nanoparticles

References 1. Delplace, V.; Couvreur, P.; Nicolas, J. Polym. Chem. 2014, 5, 1529-1544. 2. Nicolas, J. Chem. Mater. 2016. 3. Harrisson, S.; Nicolas, J.; Maksimenko, A.; Bui, D. T.; Mougin, J.; Couvreur, P. Angew. Chem., Int. Ed. 2013, 52, 1678-1682. 4. Delplace, V.; Guegain, E.; Harrisson, S.; Gigmes, D.; Guillaneuf, Y.; Nicolas, J. Chem. Commun. 2015, 51, 12847-12850.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

A promising vaginal gel containing lactobacillus crispatus for the prevention of the infection by neisseria gonorrhoeae N’Guessan Gnaman K.C.a,b, Aka-Any-Grah S.a, Geiger S.b, Bouttier S.c, Yeo A.d, Villebrun S.b, Huang N.b, Faye-Kette H.d, Ponchel G.b, Koffi A.A.a, Agnely F.b a.

Laboratoire de Pharmacie Galénique, Cosmétologie et législation pharmaceutique, UFR des Sciences Pharmaceutiques et Biologiques d’Abidjan, 01 BP V 34 Abidjan 01, Côte d’Ivoire. b. Institut Galien Paris-Sud, CNRS UMR 8612, Univ. Paris-Sud, Université Paris-Saclay, 92290 Châtenay-Malabry, France. c. EA 4043, Unité des bactéries pathogènes et Santé, Univ. Paris-Sud, Université Paris-Saclay, 92290 Châtenay-Malabry, France. d. Institut Pasteur, Abidjan, Côte d’Ivoire.

Infection by Neisseria gonorrhoeae (NG) is mainly sexually transmitted. It causes infertility both in men and women, severe damage in newborns and can multiply by three or more the risk of contracting HIV [1]. Drug resistance against the last intention treatment is increasing [2]. Moreover, there are currently no vaccine nor new drug emerging on the market. This infection has thus become a major public health issue. Our purpose was to develop a vaginal preventive gel containing Lactobacillus crispatus (LBc). Indeed, LBc is the predominant microorganism of the vaginal flora, and it could inhibit NG [3-6]. First, its capacity to inhibit NG was confirmed. Then, gels containing different ratios of poloxamer 407, a temperaturesensitive polymer, and sodium alginate, a mucoadhesive, biocompatible polymer, were formulated with or without LBc. Their rheological properties were characterized in physiological conditions (37°C with simulated vaginal fluid (SVF)) or in conditions of administration (25°C). The most promising gel was the one containing 21.5 wt% of poloxamer 407, 1 wt% of sodium alginate and 2.108 LBc/g of gel. Its pH, stability, seringability and adhesivity were evaluated. It was active on NG and remained thermogelling after dilution by SVF. LBc were viable in this formulation at 4°C. References 1. OMS, Aide-mémoire N°110 de Novembre 2014, consulté le 12/05/2016. 2. OMS, Aide-mémoire N°194 de Mai 2015, consulté le 12/05/2016. 3. Antonio M.A.D. et al, The Identification of Vaginal Lactobacillus Species and the Demographic and Microbiologic Characteristics of Women Colonized by These Species, J Infect Dis., 1999. 4. Amant D. C. St. et al, Inhibition of Neisseria gonorrhoeae by lactobacillus species that are commonly isolated from the female genital tract, Infect Immun. , 2002. 5. Vasquez A. et al, Vaginal Lactobacillus Flora of Healthy Swedish Women, J Clin Microbiol. , 2002. 6. Yu-li S. et al, Identification of and Hydrogen Peroxide Production by Fecal and Vaginal Lactobacilli Isolated from Japanese Women and Newborn Infants, J Clin Microbiol., 1999.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Oral Presentations: Analysis, Diagnostic and Theranostic

Elodie Millart Lipid Janus nanoparticles with superparamagnetic compartment : from the implementation process to theranostic applications Sophie Houvenagel Nanocapsules made of fluorinated polymers for tumor imaging by ultrasonography Jeanne Bataille Toward the development of miniaturized analytical tool for the diagnosis of transthyretin familial amyloidosis Jean-Baptiste Coty Activation of complement system by nanomedicines: development of a high throughput method and determination of an indicator of activation

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Lipid-based Janus nanoparticles : overview and focus on ferrofluid incorporation for theranostic applications Elodie Millart a, Aude Michel b, Christine Ménager b, Sylviane Lesieur a, Vincent Faivre a a

Institut Galien Paris-Sud, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 5 rue Jean-Baptiste Clément, 92296 Châtenay-Malabry, France b Laboratoire PHENIX, Sorbonne Universités, UPMC, Université Paris 06, UMR CNRS 8234, 4 place Jussieu 75005 Paris, France

In recent years, the team Physico-Chimie des Systèmes Polyphasés (Institut Galien Paris-Sud) has developed original compartmented lipid nanometer-sized particles produced by high pressure homogenization, a scalable process, with marketed and pharmaceutically approved excipients [1]. The particles actually belong to the family of Janus nano-objects [2] as they are organized in two juxtaposed substructures : one half is a droplet of liquid-state lipids while the other half is vesicle-like and encloses an aqueous core delimited by a phospholipid-containing bilayer shell. Added to the intrinsic biocompatibility of the constituting lipids, such a system provides a potentially very valuable tool in pharmaceutical and biomedical fields, able to separately incorporate and co-convey hydrophilic and lipophilic substances with distinct activities, for example, a medical imaging agent and a drug for coupling diagnosis and therapy. Here, we are interested in loading Janus nanoparticles with a magnetic fluid composed of superparamagnetic iron oxide nanocrystals (ferrofluid, FF), indeed as efficient contrast agent for magnetic resonance imaging (MRI), being magnetically targetable and providing ability for hyperthermia treatment [3]. Alternately, hydrophilic or lipophilic FF compatible with the production process have been developed by investigating different stabilization pathways of the nanocrystals depending on the encapsulation compartment. Furthermore, some optical microscopy images and X-ray diffraction experiments have helped to better understand the formation mechanism of Janus nanoparticles and the influence of FF on it.

Figure 1 : CryoTEM image of Janus nanoparticles. References [1] Faivre V. et al, Patent application WO 2015/011419 [2] Tran L.T.C. et al, Janus nanoparticles : materials, preparation, and recent advances in drug delivery, Expert Opin. Drug Deliv., 2014 [3] Lesieur S. et al, Multifunctional nanovectors based on magnetic nanoparticles coupled with biological vesicles or synthetic liposomes, J. Mater. Chem., 2011

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Nanocapsules made of fluorinated polymers for tumor imaging by ultrasonography Sophie Houvenagel a, Guilherme Picheth a, Nicolas Huang a, Camille Dejean b, Laurence Moine a, Nicolas Tsapis a a

Institut Galien Paris-Sud, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 92296 ChâtenayMalabry, France b BioCIS, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 92296 Châtenay-Malabry, France

Ultrasound imaging is a non-invasive, cost-effective, and portable diagnostic modality which can be used in combination with therapy. However, differentiation of tissues requires the use of ultrasound contrast agents (UCAs). Nanocapsules (NCs) consisting of a liquid core of perfluorooctyl bromide (PFOB) surrounded by a biodegradable and biocompatible shell of poly(lactide-co-glycolide)-poly(ethylene glycol) (PLGA-PEG) have appeared as promising UCAs due to their stability and stealth properties1. Nevertheless, the acoustic response of these capsules revealed to be too weak at clinical frequencies, owing mainly to the important thickness of the capsule. The strategy of decreasing the polymer quantity in the formulation, which proved to be efficient to reduce the thickness of plain PLGA NCs2, failed using PLGAPEG. Indeed, this resulted in local dewetting and formation of non-encapsulated PFOB droplets along with thick-shell NCs. In order to reduce the capsule thickness while keeping its stealth properties with PEG chains, new fluorinated end-functionalized polymers were synthesized. These polymers significantly reduced the interfacial tension between PFOB and an organic solvent, suggesting that the fluorinated end chain interacts with PFOB and stabilize the interface. The PFOB encapsulation efficiency in NCs was also increased with these polymers. We present here NCs characterization in terms of size distribution, shell thickness and morphology.

1

(Diou et al, Long-circulating perfluorooctyl bromide nanocapsules for tumor imaging by 19F MRI, Biomaterials, 2012) 2

(Pisani et al, Polymeric nano/microcapsules of liquid perfluorocarbons for ultrasonic imaging: Physical characterization, Langmuir, 2006)

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Toward the development of miniaturized analytical tool for the diagnosis of transthyretin familial amyloïdosis J. Bataillea, A. Pallandreb, C.Smadjaa, M.Tavernaa, R.Danielc, Régis Danielc, Florence Gonnetc, Cédric Przybylskic, S.Descroixd, F.BuhlerVarennee, C.Poüsf, I.Le Potiera a

CNRS, UMR 8612, Equipe 4, Proteins and Nanotechonologies in Analytical Sciences, Faculté de Pharmacie, Université Paris-Sud, Châtenay-Malabry, France; b CNRS, UPR 20, Laboratoire de Photonique et de Nanostructures, Marcoussis, France ; c CNRS, UMR 8587, Laboratoire Analyse et Modélisation pour la Biologie et l’Environnement, Université d’Evry Val-d’Essonne, Evry, France ; d CNRS, UMR 168, Institut Curie, France ; e CNRS, UMR 8612, Equipe 6, Amélioration du Passage des Barrières par les Molécules Biologiquement Actives, Faculté de Pharmacie, Université Paris-Sud, Châtenay-Malabry, France ; f Hôpital Antoine Béclère (AP-HP), Service de Biochimie, Clamart, France.

Familial transthyretin amyloidosis (ATTR) is a degenerative disease characterized by the formation of amyloid fibril deposits, in different organs and tissues. These deposits are mainly composed of transthyretin aggregates (TTR)1. ATTR is associated with single amino acid substitutions in TTR sequence. The objective of this project is to develop a miniaturized "point of care" system able to detect the wild type and mutant forms of TTR. This microsystem will integrate two modules: one dedicated to the TTR tryptic digestion and the second to the separation and quantification of TTR fragments of interest. TTR digestion has been carried out using two methods: on-a-gel digestion and digestion with trypsinimmobilized magnetic beads2 in batch or in a microfluidized bed3,4. The digestion samples were analyzed by capillary electrophoresis (CE) and ultra performance liquid chromatography with mass spectrometry (UPLC-MS). The amount of grafted trypsin on the magnetic beads was optimized prior to digestion with (i) BCA protein assay kit, (ii) determination of the affinity of the enzyme for bapna substrate and (iii) measurement of beads surface charge. We are currently investigating the TTR digestion on-a-chip containing the trypsin-immobilized magnetic beads characterized above and preliminary results are quite promising. References 1 Sonia Korchane et al., Derivatization strategies for CE-LIF analysis of biomarkers: Toward a clinical diagnostic of familial transthyretin amyloidosis, Electrophoresis, (2014). 2

Virginie Audonnet et al., Polymeric coating of magnetic microparticles and PDMS microchannels for online detection of prion protein in a microfluidic chip, ACS, (2009). 3

Mai Thanh Duc et al., Magneto-immunocapture with on-bead fluorescent labeling of amyloid-β peptides: towards a microfluidized-bed-based operation, The Analyst (2015). 4

French “Microfluidic system having a magnetic particle bed” WO2014037674 A1, (2012) Figure 1: Electropherograms represent peptides mapping obtained by CE WP

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Activation of complement system by nanomedicines: development of a high throughput method and determination of an indicator of activation Jean-Baptiste Coty, Fanny Varenne, Jean-Jacques Vachon, Christine Vauthier Institut Galien Paris-Sud, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 92290 Châtenay-Malabry,

France Interactions between nanomedicines and complement system become an unavoidable property of nanomaterial, as it gives information about nanomedicine potential toxicity and biodistribution1. Common methods used to deal with it are either expensive (Elisa) or variable from one experiment to another (CH50 assay)2. In our lab, 2D-immunoelectrophoresis was used as a direct method to monitor complement protein C3 cleavage upon incubation with nanoparticles3. However, this method was limited to comparative studies due to the low number of samples that could be run and the tediousness of the method (5 samples in 19h). We propose an optimized method to study at a high-throughput rate the interaction between a given nanomedicine and the protein C3, implementing a novel experimental procedure (MultiCrossed-IE). The new modalities make possible the analysis of 35 samples at once, while reducing handling complexity and electrophoresis time (5h). These improvements allow the screening of concentrations for a given nanomedecine and the establishment of a pattern of activation. The surface concentration (in cm2/mL) of nanomaterial that induces a cleavage of 50% of the C3 protein can be determined (C3A50) and used as a new indicator characterizing a nanomedecine.

References 1

M. A. Dobrovolskaia et al., Preclinical Studies To Understand Nanoparticle Interaction with the Immune System and Its Potential Effects on Nanoparticle Biodistribution, Molecular Pharmaceutics, 2008. 2

P. P. Wibroe et al., Complement Sensing of Nanoparticles and Nanomedicines, in Functional Nanoparticles for Bioanalysis, Nanomedicine, and Bioelectronic Devices, 2012. 3

C. Passirani et al., Interactions of nanoparticles bearing heparin or dextran covalently bound to poly(methyl methacrylate) with the complement system, Life Sciences, 1998.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Posters: N°1 Corentin Berardet Study of the oligomerization and aggregation of hIAPP by CE-UV and IMS-MS N°2 Alexandre Bordat Thermoresponsive polymer nanoparticles for the hyperthermia-triggered release of anticancer drugs N°3 Cédric Crosnier New preconcentration strategies for the development of diagnostic tools based on microsystems or mass spectrometry N°4 Gianpiero Lazzari Shedding lights on cancer cells and their microenvironment: in vitro 3D tumor model of pancreatic cancer for preclinical prediction of in vivo behavior of nanomedicines N°5 Zeineb Maaroufi Valorization of essential oils of some aromatic and medicinal plants from Tunisia as new antileishmaniasis substances N°6 Sophia Malli New insights on Trichomonas vaginalis mobility into pharmaceutical hydrogels by multiple particle tracking N°7 Julien Massiot Phototriggered drug release from liposomes: a mechanistic study N°8 Julie Mougin Contribution to physico-chemical studies of squalenoylated nanodrugs for cancer treatment

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

N°9 Marline N'Diaye Targeting of retinoblastoma cells with glycoconjugated tetraphenylporphyrin dimers for twophoton photodynamic therapy N°10 Arnaud Peramo Palladium nanoparticles for intracellular modification of proteins N°11 Marie Rouquette Development of squalenoyl adenosine nanoparticles for treating hepatic ischemia N°12 Coralie Ruel Mass spectrometry-based glycomics of biological fluids: towards the identification of a novel class of disease biomarkers N°13 Johanna Tran Design of a novel class of biodegradable and functionalized polyesters for cardiac therapy N°14 Daniele Vinciguerra Synthesis and evaluation of dual-functionalized polymer prodrug nanoparticles for anticancer therapy N°15 Xiao Wu Assessment of the mechanism of interaction of a hydrophobic photoactivatable drug with phospholipid bilayers, and formulation of nanocarriers for anticancer therapy

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°1 Study of the oligomerization and aggregation of hIAPP by CE-UV and IMS-MS Corentin Berardet a, Julia Kaffy b, Sandrine Ongeri b, Myriam Taverna a a

Institut Galien Paris Sud, UMR8612, Protein and Nanotechnology in Analytical Science (PNAS), CNRS, Univ. Paris-Sud, Université Paris-Saclay, 5 rue Jean-Baptiste Clément, 92290 ChâtenayMalabry, France, b Biomolécules : Conception, Isolement, Synthèse, UMR 8076, Molécules Fluorées et Chimie Médicinale, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 5 rue Jean-Baptiste Clément, 92290 Châtenay-Malabry, France

Type II diabetes (T2D) and associated cardiovascular risks are now considered as major concerns of public health, affecting more than 300 million people in the world. Human Islet Amyloid Polypeptide (hIAPP) is a 37 amino acids peptide co-secreted with insulin by pancreatic  cells and involved in glucose homeostasis. Its aggregation under certain conditions leads to  cells degeneration and promotes T2D by inhibiting insulin secretion. Both mechanisms and concerned species of hIAPP toxicity are still unknown. 1 Here we propose to develop a capillary electrophoresis method to monitor in vitro oligomerization and aggregation of hIAPP in order to get more insight on this pathological process and also to develop a screening method to test for new therapeutic agents. Due to its high pI (8.59), hIAPP is cationic under physiological conditions which leads to its adsorption onto the silica capillary walls and reproducibility issues. A polybrene coating was developed in order to inhibit this phenomenon. Identification of most of the different species involved in this process will be done by Ion Mobility Spectroscopy-Mass Spectrometry. 2 This method will allow us in the future to test peptidomimetics inhibitors of hIAPP aggregation in order to target specifically toxics species for  cells. Preliminary tests to select candidates are conducted with Thioflavin-T fluorescence assays.

hIAPP aggregation pathway 3 1

Mukherjee et al, Type 2 diabetes as a protein misfolding disease, Trends in Molecular Medicine, 2015. Young et al, Ion Mobility Spectrometry–Mass Spectrometry Defines the Oligomeric Intermediates in Amylin Amyloid Formation and the Mode of Action of Inhibitors, Journal of the American Chemical Society, 2014. 3 Wu and Shea, Structural Similarities and Differences between Amyloidogenic and Non-Amyloidogenic Islet Amyloid Polypeptide (IAPP) Sequences and Implications for the Dual Physiological and Pathological Activities of These Peptides, Plos Computational Biology, 2013. 2

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°2 Thermoresponsive polymer nanoparticles for the hyperthermia-triggered release of anti-cancer drugs Alexandre Bordat, Tanguy Boissenot, Imen Ben Chraït, Julien Nicolas, Nicolas Tsapis Institut Galien Paris-Sud UMR CNRS 8612

To improve the targeting of tumors by nanoparticles, we wish to develop thermoresponsive nanoparticles. The first step of the project consists in designing a thermoresponsive polymer which will be the building material for the nanoparticles. Previous work by Imen Ben Chraït has shown that the diblock copolymer P(AAm-co-AN)-b-PEGMA1–3 (poly(acrylamide-coacrylonitrile)-b-(poly(ethylene glycol) methacrylate)) is thermoresponsive in an acceptable temperature range (between 10 °C and 60 °C). The monomer composition of the P(AAm-coAN) block determines the temperature at which the polymer completely solubilizes in water or PBS (named upper critical solution temperature, UCST). We are thus optimizing the polymer composition to yield a UCST of 42 °C, to release the active pharmaceutical ingredient (API) upon mild hyperthermia. The PEGMA block is used to obtain colloidal stability without using surfactant and to enable longer circulation of the nanoparticles in the blood stream. We use the reversible addition-fragmentation chain transfer (RAFT) technique to control the polymerization, namely the size of the polymer and we can tune the length of each block. We have started formulating nanoparticles using the solvent displacement technique and are currently characterizing them. The next steps are going to be encapsulating a cytotoxic drug (doxorubicin), and evaluating the drug loading of our nanoparticles as well as the drug release with and without hyperthermia. Formulated in thermoresponsive nanoparticules

Doxorubicin (DOX)

Mid hyperthermia

100 nm

Controlled release of DOX

References 1.

Zhang, H., Tong, X. & Zhao, Y. Diverse thermoresponsive behaviors of uncharged UCST block copolymer micelles in physiological medium. Langmuir 30, 11433–11441 (2014).

2.

Huang, G. et al. Self-assembled UCST-Type Micelles as Potential Drug Carriers for Cancer Therapeutics.

3.

Li, W. et al. Angewandte Antitumor Drug Delivery Modulated by A Polymeric Micelle with an Upper Critical Solution Temperature **. (2015). doi:10.1002/anie.201411524

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°3 New preconcentration strategies for the development of diagnostic tools based on microsystems or mass spectrometry Cédric Crosnier, Myriam Taverna Institut Galien Paris Sud, UMR8612, Protein and Nanotechnology in Analytical Science (PNAS), CNRS, Univ. Paris-Sud, Université Paris-Saclay, 5 rue Jean Baptiste Clément, 92290 ChâtenayMalabry, France

35 million people in the world suffer from Alzheimer’s disease (AD). There is currently no treatment for AD due to the difficulty to diagnose this disease on the one hand, and to enrol well diagnosed patient for testing new treatment protocols during clinical trials on the other hand. The cognitive impairment can only be detected at a late stage of the pathology where the brain damages are irreversible. In this context, scientists are developing new methods of molecular diagnostics that allow diagnosing AD at an earlier stage, and also to distinguish it from other neurodegenerative diseases. Thus the objective of the thesis consists to develop analytical setups based on either lab-on-achip or mass spectrometry to diagnose different neurodegenerative diseases. One important aspect of the project is the realization of new strategies for sample enrichment and matrix removal as an alternative to the conventional preconcentration methods such solid phase extraction. Today we present the first results of preconcentration method using isotachophoresis (picture below) which can allow the detection of amyloid βeta peptides at a concentration of 1 nM. This strategy was applied to conventional capillary electrophoresis system and downscaling to a microchip platform is under progress.

Isotachophoresis in capillary electrophoresis of Aβ1-40

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°4 Shedding lights on cancer cells and their microenvironment: in vitro 3D tumor model of pancreatic cancer for preclinical prediction of in vivo behavior of nanomedicines. Gianpiero Lazzari, Simona Mura, Patrick Couvreur a

Institut Galien Paris-Sud, UMR 8612, CNRS, Univ. Paris-Sud, Université Paris-Saclay, Faculté de Pharmacie, 5 rue JB Clément, 92296 Châtenay-Malabry, France

Nanomedicines offer the possibility to improve the efficacy of anticancer drugs, however progresses in pancreatic cancer therapy have remained exceedingly slow mainly as consequence of an inefficient drug delivery to cancer cells. 1 The extensive desmoplastic reaction is the hallmark of this tumor and acts as a physical barrier sequestrating nanomedicines, blocking their diffusion and limiting the effectiveness of the treatment. 2,3 Thus, in vitro models, which reliably mimic the clinical conditions, are highly required for an appropriate preclinical screening of nanomedicines. Herein attention focuses on 3D in vitro tumor spheroids, which have been already successfully constructed using the Panc-1 cell line. Viable spheroids have been maintained in culture up to 17 days while only aggregates of poorly viable cells that quickly disassembled were obtained using the BxPc3 cells. Co-culture of Panc1 cells and fibroblasts has been also achieved. Multiple culture with endothelial and stellate cells is currently investigated in order to reproduce the structural and functional integration of the various cell types constituting the pancreatic tumor. 4 Once validated, the model will be used for screening nanomedicines. Information gathered by this research will enable to identify the strategies that would prompt their penetration in the tumor mass making shorter the step for their clinical translation.

Figure 1. Panc-1 spheroids at d4, 7, 13 and 17 post seeding. Scale bar: 200µm

References 1

Torosean et al, Nanoparticle uptake in tumors is mediated by the interplay of vascular and collagen density with interstitial pressure, Nanomedicine, 2013. 2 Kadaba et al, Imbalance of desmoplastic stromal cell numbers drives aggressive cancer processes, J Pathol, 2013. 3 Sutherland et al, Cell and environment interactions in tumor microregions: the multicell spheroid model, Science, 1988.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°5 Valorization of essential oils of some aromatic and medicinal plants from Tunisia as new antileishmaniasis substances. Zeineb Maaroufi1,2, Olfa Bachrouch2, Sandrine Cojean3, Philippe Loiseau3, Manef Abderrabba2, Florence Agnely1, Ghozlene Mekhloufi1 1

Institut Galien Paris-Sud, UMR 8612, CNRS, Univ. Paris-Sud, Université Paris-Saclay, Faculté de Pharmacie, Châtenay-Malabry, France 2 Laboratoire Matériaux, Molécules et Applications, IPEST, La Marsa, Tunisie 3 Laboratoire de Chimiothérapie Antiparasitaire, UMR 8076, CNRS BioCIS, LabEx LERMIT, Univ. Paris-Sud, Université Paris-Saclay, Châtenay-Malabry, France

Leishmaniasis is a vector parasitic disease caused by Leishmania, a protozoan parasite. It is considered as a major public health concern, as 12 million people are infected worldwide with 1.3 million new cases each year1. Leishmaniasis causes high rate morbidity and mortality mainly in the poorest regions on earth. Most current chemotherapies based on the antimony have several limitations such as toxic side effects, high cost, relapse, parasite resistance, etc. So, it is urgent to develop effective new drugs to replace or supplement those in current use. The plants are undoubtedly a valuable source of new bioactive molecules. This work aims to find alternatives in the treatment of cutaneous leishmanianis based on plant extracts, such as essential oils (EOs) and organic extracts (OEs). Based on data in traditional medicine, we chose Pisctacia lentiscus, Citrus Limon and Artemisia Herba-Alba, from 2 different regions for each. After harvesting and drying, the extraction yield was 0.0066 to 1.3% for EOs and 2.7 to 35% for OEs. The next step is to determine the antiparasitic activity of EOs and OEs. Then, in order to improve the therapeutic efficacy of these bioactive molecules, they will be encapsulated in nanoemulsions for the treatment of cutaneous leishmaniasis.

Reference 1 WHO, Leishmaniasis. Fact sheet n°375, February 20 15.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°6 New insights on Trichomonas vaginalis mobility into pharmaceutical hydrogels by multiple particle tracking Sophia Malli a,b, Christian Bories b, Valérie Nicolas c, Tiphany Grisin a, Christine Vauthier a, Gilles Ponchel a, Philippe M. Loiseau b, Kawthar Bouchemal a a

Institut Galien Paris Sud, UMR CNRS 8612, School of Pharmacy, University Paris Sud, ChâtenayMalabry, France; bBioCis, UMR CNRS 8076, Antiparasitic Chemotherapy, School of Pharmacy, University Paris Sud, Châtenay-Malabry, France

T. vaginalis is a protista parasite responsible for the urogenital trichomoniasis, one of the most common STI in the world. T. vaginalis is implicated in the upper reproductive tract postsurgical infections, infertility, premature rupture of the placental membranes, premature labor, low-birth-weight infants and neonatal morbidity and mortality. Conventional treatment of urogenital trichomoniasis relies on metronidazole (MTZ), a 5nitroimidazole drug. Although oral metronidazole is an effective therapy for T. vaginalis infection, treatment failure is reported due to significant gastrointestinal adverse effects. In this context, we designed new formulations composed of hydrogels containing mucoadhesive chitosan, thermosensitive pluronic F127 and MTZ used as antiparasitic drug. Also, we investigated the effect of different formulation components on T. vaginalis mobility by multiple particle tracking.1 The results of trajectories and mean square displacement (MSD) were analysed and compared to the ones obtained with chitosan, chitosan/F127, chitosan/MTZ, F127, HEC, HEC/MTZ and MTZ solution. Chitosan, F127 and MTZ were used at concentrations of 1wt%, 20wt% and 7 mg/mL, respectively. Acting as a trap, the formulation could be able to immobilize T. vaginalis and, in turn, to reduce parasite infectivity. This study highlighted for the first time the behaviour of T. vaginalis trajectories in liquid solution and in the presence of hydrogels. We demonstrated that thermosensitive hydrogel F127 (20wt%) was able to hinder T. vaginalis mobility. The investigation of anti-T. vaginalis activity2 suggests that there is no synergy between formulation components and MTZ. The overall results therefore highlight the ability of the formulation F127/MTZ to form a double barrier against T. vaginalis: a physical and a pharmacological barrier. 1

(Bouchemal K et al, Thermosensitive and mucoadhesive pluronic®/ hydroxypropylmethylcellulose hydrogel containing the mini CD4 M48U1 is a promising efficient barrier against HIV diffusion through macaque cervicovaginal mucus, Antimicrobial Agents and Chemotherapy, 2015). 2 (Pradines B et al, Drug-free nanoparticles are active against Trichomonas vaginalis and non-toxic towards pig vaginal mucosa, Pharmaceutical Research, 2015).3 (Zhang M et al, Nanostructured fluids from pluronic mixtures, International Journal of Pharmaceutics, 2013).

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°7 Phototriggered drug release from liposomes: a mechanistic study Julien Massiot, Ali Makky, Véronique Rosilio Université Paris-Sud, Faculté de Pharmacie, Institut Galien Paris Sud - UMR CNRS 8612, 92296 Châtenay-Malabry cedex, France

Phototriggered release of encapsulated drugs from liposomes is currently considered as a potential and an interesting modality for drug delivery in a controlled manner1. Indeed, phototriggered liposomes are nanocarriers that can be activated by light at a specific wavelength to release their cargo upon illumination. Such a drug delivery system has been explored since decades and potentially presents a versatile tunable method since the features such as the source of wavelength, duration, and intensity of light treatment can be easily controlled as desired2. Typically a phototriggered liposome includes a photosensitive molecule embedded in the lipid bilayer matrix namely with unsaturated fatty acyl chain. Upon their illumination, photosensitive molecules will generate reactive oxygen species (ROS), such as singlet oxygen, that oxidize the phospholipids unsaturated chains leading thus to defects in the lipid bilayer and increase in the membrane permeability3. However, although the potentiality of such systems exists, their drug release efficiency is limited to small molecules (Mw < 1 KDa). In addition, it depends on several factors such as the lipid composition, the penetration depth of the photosensitizers in the lipid bilayer, and their singlet oxygen quantum yield. Hence, in the aim of developing a new drug delivery system allowing the release of macromolecules with anti-cancerous activity, it is extremely interesting to investigate the impact of the illumination on liposome membrane permeability, lipid peroxidation, membrane thermodynamical behavior and mechanical properties. To do so, liposomes with different lipid composition incorporating different photosensitizers (meso-tetra(3hydroxyphenyl)porphyrin (m-THPP), verteporfin and pheophorbide A) were chosen as drug delivery systems. The influence of light illumination on the membrane permeabilization was studied by monitoring the release of a fluorescent probe encapsulated in their aqueous core. The efficiency of bilayer lipids oxidation in the presence of photosensitizers was monitored using methyl linoleate as a target of singlet oxygen. The thermodynamic behavior of lipid bilayers upon illumination was studied using differential scanning calorimetry, and changes in membrane mechanical properties were assessed by AFM-nanoindentation experiments on supported planar bilayers. Our results showed quantitatively that the illumination of the studied systems altered significantly their lipid bilayer properties, and that the extent of the system efficiency was dependent upon a photosensitizer / phospholipid combination. References 1

2 3

Lovell, J. F et al., Porphysome Nanovesicles Generated by Porphyrin Bilayers for Use as Multimodal Biophotonic Contrast Agents. Nat. Mater. (2011) Shum, P. et al., Phototriggering of Liposomal Drug Delivery Systems. Adv. Drug Deliv. Rev. (2001) Mojzisova, H. et al., Photosensitizing Properties of Chlorins in Solution and in Membrane-Mimicking Systems. Photochem. Photobiol. Sci. (2009)

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°8 Contribution to physico-chemical studies of squalenoylated nanodrugs for cancer treatment Julie Mougina, Andrei Maksimenko, Franco Dosiob, Barbara Stellab, Patrick Couvreura a

Institut Galien Paris-Sud, Châtenay-Malabry, France, bDipartimento di Scienza e Tecnologia del Farmaco, Torino, Italy

Doxorubicin is one of the most used anticancer drugs, showing efficacy in a series of adult and pediatric cancers such as solid tumors, leukemias…1 However, its severe toxicity induces limiting side effects, mainly irreversible cardiotoxicity. To overcome those limitations, vectorization strategies have been developed and some of them reached the market, like liposomal formulations Caelyx and Myocet.2 Lately, Pr. Couvreur’s group elaborated a new vectorization method. By coupling a drug to a natural and biocompatible lipid, the squalene, bioconjugates were formed, presenting self-assembling properties when nanoprecipitated in water.3 Applying the squalenoylation to doxorubicin chlorhydrate lead to the formation of nanodrugs showing an increased anticancer activity, a prolonged plasmatic circulation time and a strongly reduced cardiotoxicity compared with free doxorubicin. These improvements have been attributed to the particular “loop-train” structure of these nanoobjects, prolonging plasmatic half-life of doxorubicin by avoiding massive uptake by liver and spleen macrophages.2 Besides, the coupling of basic doxorubicin to squalene revealed the formation of spherical nanoparticles with a lesser anticancer activity than loop-train nanoparticles. We are now investigating more precisely the conditions to obtain these particular structures and the physico-chemical mechanisms involved, in order to explain in a more fundamental way the pharmacokinetic properties of the loop-train versus sphericle nanoparticles.

References 1

Octavia Y., Tocchetti C. G., et al,. Doxorubicin-induced cardiomyopathy : From molecular mechanisms to therapeutic strategies, Journal of Molecular and Cellular Cardiology, 2012. 2

Maksimenko A., Dosio F., et al., A unique squalenoylated and nonpegylated doxorubicin nanomedicine with systemic long-circulating properties and anticancer activity, Proceedings of the National Academy of Sciences, 2014. 3

Couvreur P., Stella B., et al., Squalenoyl nanomedicines as potential therapeutics, Nano Letters, 2006.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°9 Targeting of retinoblastoma cells with glycoconjugated tetraphenylporphyrin dimers for two-photon photodynamic therapy Marline N’DIAYE, Véronique ROSILIO Université Paris-Sud, Faculté de Pharmacie, Institut Galien Paris Sud - UMR CNRS 8612, Université Paris-Saclay, 92296 Châtenay-Malabry cedex, France

Retinoblastoma is a malignant retina tumor affecting 1 child in 15.000 births. It is currently treated by surgery and chemotherapy. Photodynamic therapy (PDT) is considered as a promising alternative1 to these treatments. It consists in intravenous injection of a photosensitizer (PS) such as a tetraphenyl porphyrin (TPP), which accumulates in the tumor, and then illumination of the tumor area with a laser of appropriate wavelength. The porphyrin is non-toxic in darkness, but when excited by light in the presence of oxygen, it generates singlet oxygen. To be efficient, the porphyrin must penetrate into cells. However, TPPs are hydrophobic and nonspecific. Four series of glycoconjugated TPP derivatives have been synthesized by Dr. Philippe Maillard’s group at Institut Curie in order to get more hydrophilic compounds, sufficiently amphiphilic to cross the cell membrane through passive diffusion, and sufficiently specific to penetrate into cells via a mannose receptor overexpressed at retinoblastoma cell surface. The last series of TPPs includes TPP dimers for two-photon absorption. However these molecules are even more hydrophobic than monomeric ones, aggregate in the aqueous medium and do not penetrate into cells through passive diffusion or receptor-mediated transport. Our project aims at developing polymer nanoparticles covered by TPP-bearing erythrocyte membranes for increased biocompatibility and specificity 2,3. The specific interaction will be checked using giant unilamellar vesicles (GUV) expressing the MRC2 receptor by a cell-free method. The lipid composition of the GUVs will be deduced from a lipidomic analysis4. Recognition activity of glycoconjugated TPPs by the receptor will be evaluated by competition experiments. Endocytosis and photocytotoxic effect will be evaluated in vitro in cell cultures, and if successful in vivo on orthotopic retinoblastoma models developed in mice (Institut Curie).

References 1 Teixo R et al. Cancer Metastasis Rev. doi : 10.1007/s10555-014-9544-y, 2015 2 Zhang L, and Zhang LF, Nano LIFE 1, 163, 2010 3 Hu C-MJ et al. Proc Natl Acad Sci USA 108, 10980, 2011 4 Essaid D. et al. submitted, 2016.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°10 PLGA-PEG-supported Pd nanoparticles as efficient catalysts for Suzuki-Miyaura coupling reactions in water Peramo Arnaud, Anaëlle Dumas, Desmaële Didier, Pr. Couvreur Patrick

Chemical transformations that can be performed selectively under physiological conditions are highly desirable tools to track biomolecules and manipulate complex biological processes. Here, we report a new nanocatalyst consisting of small palladium nanoparticles stabilized on the surface of PLGA-PEG nanoparticles that show excellent catalytic activity for the modification of biological building blocks through Suzuki-Miyaura cross-coupling reactions in water. Brominated or iodinated amino acids were coupled with aryl boronic acids in phosphate buffer in good yields. Interestingly, up to 98% conversion into the coupled amino acid could be achieved in 2 h at 37 °C using the stable, water-soluble cyclic triolborate as organometallic partner in the presence of only 1 mol% of palladium. These results pave the way for the modification of biomolecules in complex biological systems such as the intracellular space. References 1J. A. Prescher, C. R. Bertozzi, Nat. Chem. Biol., 2005, 1, 13-21 2 C. D. Spicer, B. G. Davis, Nat. Commun., 2014, 5, 4740-4754 3E. M. Sletten, C. R. Bertozzi, Angew. Chem. Int. Ed., 2009, 48, 6974-6998 4J. M. Chalker, C. S. C. Wood, B. G. Davis, J. Am. Chem. Soc., 2009, 131, 16346-16347 5A. Dumas, C. D. Spicer, Z. H. Gao, T. Takehana, Y. Y. A. Lin, T. Yasukohchi, B. G. Davis, Angew. Chem. Int. Ed. 2013, 52, 3916-3921 6A. Dumas, A. Peramo, D. Desmaële, P. Couvreur, Chimia, 2016, 70, 252-257

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°11 Squalenoyl adenosine nanoparticles effect on hepatic ischemia-reperfusion injury Marie Rouquettea, Sinda Lepêtre-Mouelhia, Karine Ser-Lerouxb and Patrick Couvreura a

Institut Galien Paris-Sud, Université Paris-Sud, UMR CNRS 8612, Faculté de Pharmacie, 5 rue JeanBaptiste Clément, 92296 Châtenay-Malabry, France b Plateforme d’évaluation pré-clinique, Institut Gustave Roussy, Pavillon Recherche 1, Niveau 1, 114 rue Edouard-Vaillant, 94805 Villejuif, France

Ischemia-reperfusion injury causes severe liver damage during surgical procedures like liver transplantation or tissue resection and represents the major complication of a hemorrhagic shock. Adenosine was shown to have a protective effect against ischemia in the liver1,2, however its clinical use is currently limited by its very short half-life3. This limit can be overcome by the bioconjugation of adenosine with squalene - a natural triterpene - which leads to the formation of stable nanoparticles. Recently, our team demonstrated that the squalenoyl adenosine nanoparticles (SQ-Ad NPs) had a beneficial effect on cerebral ischemia4. Moreover, the biodistribution profile of these nanoparticles in mice showed their accumulation in the liver and revealed the release of adenosine in this organ for as long as 24h after injection5. Hence we try to validate the hypothesis that SQ-Ad NPs will provide hepatoprotection before induction of ischemia-reperfusion. We use a mouse model of global ischemia (described by the scheme hereunder) and present the results in terms of survival, histology and ASAT and ALAT quantitative analysis.

References 1. Lappas, C. M., Day, Y.-J., Marshall, M. A., Engelhard, V. H. & Linden, J. Adenosine A2A receptor activation reduces hepatic ischemia reperfusion injury by inhibiting CD1d-dependent NKT cell activation. J. Exp. Med. 203, 2639–2648 (2006). 2. Peralta, C. et al. The protective role of adenosine in inducing nitric oxide synthesis in rat liver ischemia preconditioning is mediated by activation of adenosine A2 receptors. Hepatology 29, 126–132 (1999). 3. Turnover of adenosine in plasma of human and dog blood | Cell Physiology. Available at: http://ajpcell.physiology.org/content/256/4/C799. (Accessed: 7th April 2016) 4. Gaudin, A. et al. Squalenoyl adenosine nanoparticles provide neuroprotection after stroke and spinal cord injury. Nat. Nanotechnol. 9, 1054–1062 (2014). 5. Gaudin, A. et al. Pharmacokinetics, biodistribution and metabolism of squalenoyl adenosine nanoparticles in mice using dual radio-labeling and radio-HPLC analysis. J. Controlled Release 212, 50–58 (2015).

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°12 Mass spectrometry-based glycomics of biological fluids: towards the identification of a novel class of disease biomarkers Coralie Ruel a,b, Myriam Tavernaa, Christophe Junotb, François Fenailleb, Thuy Tran a a

Institut Galien Paris Sud, UMR8612, Protein and Nanotechnology in Analytical Science (PNAS), CNRS, Univ. Paris-Sud, Université Paris-Saclay, 5 rue Jean Baptiste Clément, 92290 ChâtenayMalabry, France, b CEA, iBiTec-S, Service de Pharmacologie et d’Immunoanalyse, Laboratoire d’Etude du Métabolisme des Médicaments, Gif-sur-Yvette 91191, France

Glycosylation is one of the most complex types of post-translational modifications of proteins1. Disease-associated modifications in protein glycosylation constitute a major source of biomarkers and are often exploited for diagnosis and monitoring in various pathologies2. The goal of my PhD thesis is to implement an analytical platform for the analysis of free oligosaccharides in urine and/or plasma protein-bound oligosaccharides. Starting from biological fluids, the first step will be the on-line enrichment of glycoproteins on a functionalized monolithic column. Then an enzymatic deglycosylation of preconcentrated glycoproteins followed by the derivatization of released glycans with a fluorophore will be performed. The obtained glycans will be separated on-line using orthogonal techniques such as high-performance liquid chromatography and capillary electrophoresis for the efficient resolution of isomeric oligosaccharide structures. Such separation system will be coupled to high resolution mass spectrometry (Orbitrap or Q-TOF). I will present here the development and the optimization of the separation method of derivatized N-glycans or O-glycans using capillary electrophoresis coupled to mass spectrometry (Q-TOF). For this purpose, different conditions (background electrolyte, sheath liquid, mass parameters…) were tested. References 1 S. C. Bunz et al, Capillary Electrophoresis/Mass Spectrometry of APTS-Labeled Glycans for the Identification of Unknown Glycan Species in Capillary Electrophoresis/Laser-Induced Fluorescence Systems, Analytical chemistry, 2013 2 S. Bekesova et al, Nglycans in liver-secreted Enrichment on monolithic and immunoglobulinderived protein fractions, Deglycosylation support Journal of Proteomics, 2012 Released glycans

Glycoproteins

Fluorescent derivatization

Complex mixture

Fluo

EC-LIF

Fluo

Fluo

EC-MS

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°13 Design of a novel class of biodegradable and functionalized polyesters for cardiac therapy J. Tran a, J-C. Honoré b, C. Lefay b, Y. Guillaneuf b, D. Gigmes b, J. Nicolas a a

Institut Galien Paris-Sud, UMR 8612, Faculté de Pharmacie, 5 rue Jean Baptiste Clément, 92290 Châtenay-Malabry b Institut de Chimie Radicalaire, UMR 7273, Université Aix-Marseille, 52 avenue Escadrille Normandie Niemen, 13013 Marseille

Considering that heart diseases are one of the leading causes of mortality in the world, it is urgent to develop efficient cardiac therapies. Classic therapy involves active molecules injected at high doses, leading to an uncontrolled distribution in the whole body that could lead to serious toxic side effects. Our approach consists in the design of novel degradable vinyl nanoparticles loaded with biologically active molecules in order to overcome the limitations associated with the traditional therapy. We propose to prepare degradable, functional copolymers by radical ring-opening copolymerization (rROP)1,2 between cyclic ketene acetal (CKA) monomers and vinyl ether derivatives (VE), further nanoprecipitated into nanoparticles. These new copolymers comprise a polyester-like backbone together with pending functional groups from VE units. The aim is to design degradable nanoparticles loaded with growth factor, such as IGF-1, to heal the injured cardiac tissue. Targeting abilities can also be conferred to the nanoparticles by surface functionalizing them with appropriate ligands that specifically bind to the cardiac tissue.3

References 1 Delplace, V.; Nicolas, J. Nature Chem. 2015, 7, 771 2 Agarwal, S. Polym. Chem. 2010, 1, 953 3 Dvir, T.; Bauer, M.; Schroeder, A.; Tsui, J. H.; Anderson, D. G; Langer, R.; Lia, R.: Kohane, D. S. Nano Lett. 2011, 11, 4411

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°14 Synthesis and evaluation of dualfunctionalized polymer prodrug nanoparticles for anticancer therapy Daniele Vinciguerra , Patrick Couvreur and Julien Nicolas Institut Galien Paris-Sud, Université Paris-Sud, UMR CNRS 8612, Faculté de Pharmacie, 5 rue JeanBaptiste Clément, 92296 Châtenay-Malabry, France

Drug-loaded polymer nanoparticles are meant to increase the therapeutic efficacy of the drugs and to decrease their toxicity.[1]However, they also have some disadvantages, including the "burst release" and poor drug loadings. Performing a covalent linkage between the polymer scaffold and the drug (the so-called "prodrug approach"), aims to solve those issues.[2] Controlled/living radical polymerization (CLRP) was performed in our group to synthesize Gemcitabine-polyisoprene (Gem-PI) prodrug nanoparticles using the "drug-initiated method",[3] where the drug is linked to a CLRP initiator prior polymerization.[4] . Polyisoprene has been chosen due to its similarity with natural terpenoids and Gemcitabine for its significant anticancer activity. The aim of the project is to design dual-functionalized polymer prodrug nanoparticles bearing two biologically active drugs per polymer chain (one at each polymer chain-end) for combination therapy. A small library of nanoparticles will be prepared and evaluated with several drug combinations. Note that the second molecule of interest can also be a fluorescent dye or a ligand for diagnostic or targeting purposes, respectively. As a first attempt, the nitroxide exchange reaction was selected to perform this construction.[5]

References 1.

2. 3. 4.

5.

Nicolas, J., et al., Design, functionalization strategies and biomedical applications of targeted biodegradable/biocompatible polymer-based nanocarriers for drug delivery. Chem Soc Rev, 2013. 42(3): p. 1147-235. Delplace, V., P. Couvreur, and J. Nicolas, Recent trends in the design of anticancer polymer prodrug nanocarriers. Polym. Chem., 2014. 5(5): p. 1529-1544. Nicolas, J., Drug-Initiated Synthesis of Polymer Prodrugs: Combining Simplicity and Efficacy in Drug Delivery. Chemistry of Materials, 2016. Harrisson, S., et al., Nanoparticles with In Vivo Anticancer Activity from Polymer Prodrug Amphiphiles Prepared by Living Radical Polymerization. Angewandte Chemie International Edition, 2013. 52(6): p. 1678-1682. Guillaneuf, Y., et al., Radical Chain End Chemical Transformation of SG1-Based Polystyrenes. Macromolecules, 2010. 43(1): p. 91-100.

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Poster N°15 Assessment of the mechanism of interaction of a hydrophobic photoactivatable drug with phospholipid bilayers, and formulation of nanocarriers for anticancer therapy. Xiao Wu and Véronique Rosilio Université Paris-Sud, Faculté de Pharmacie, Institut Galien Paris Sud - UMR CNRS 8612, Université Paris-Saclay, 92296 Châtenay-Malabry cedex, France

β-lapachone (β-lap) is a semi-synthetic drug obtained from the tree lapachol. It has attracted increasing attention due to its antibacterial, antifungal, and anticancer properties. Indeed, βlap presents antiproliferative activity at micromolar concentrations against a variety of cancer cells. Its physicochemical characteristics could be of interest for penetration into cells by permeation through the plasma membrane, such as its low molecular weight (242.3 g/mol) and its moderately high log P (2.5). However, β-lap therapeutic applications are limited due to its low water solubility (0.038 mg mL-1 or 0.16 mM) and its hepatotoxicity. It is thus necessary to solubilize it, and to control its pharmacokinetics. Some approaches such as polymer micelles, pegylated liposomes, and cyclodextrins have been proposed, in order to improve the therapeutic activity and reduce the side effects of β-lapachone. However, attempts to form complexes with cyclodextrins have not greatly improved its solubilisation. Calvacanti et al.1 have shown that β-lapachone was incorporated into liposomes with the same loading efficiency, in the presence and absence of β-cyclodextrins. In fact, few systems have taken into account the physicochemical properties of this drug, except for its poor water solubility. Our preliminary study by surface pressure and surface potential measurements, and atomic force microscopy has shown that β-lap has a very unusual interaction with phospholipids. Our goal is thus to elucidate the mechanism of β-lap interaction with cell membranes that controls its photocytotoxicity, and to propose appropriate nanocarriers (liposomes, lipid nanoparticles) for its solubilisation and delivery to cells with a specific localization. Our methodology includes (i) the physicochemical characterization of β-lap interactions with potential excipients for nanocarriers preparation and with model membranes, (ii) the study of its photoactivatable properties in giant vesicles mimicking cells, (iii) the conception of appropriate β-Lap-loaded nanocarriers, and (iv) the evaluation of their efficiency on human breast and prostate cancer cells. Reference: 1.

I.M.F. Cavalcanti et al., European Journal of Pharmaceutical Sciences 44 (2011) 332–340

_______________________

- Angelina ANGELOVA (CR) - Gillian BARRATT (DR) - Claudie BOURGAUX (CR) - Vincent FAIVRE (MC) - François-Xavier LEGRAND (MC)

_______________________

- David CHAPRON (IE) - Ali MAKKY (MC) - Jean-Philippe MICHEL (MC) - Catherine RINGARD (MC)

- Elodie MILLART (Doc) - Marlon NUNES DA SILVA (Post-Doc)

Chef d’équipe Sylviane LESIEUR (DR)

Chef d’équipe Véronique ROSILIO (PR)

- Julien MASSIOT (Doc) - Marline N’DIAYE (Doc) - Xiao WU (Doc)

Physico-chimie des Systèmes Polyphasés Chef d’équipe Elias FATTAL (PR) _________________________

- Nicolas TSAPIS (DR) Chef d’équipe adjoint - Amélie BOCHOT (PR) - Hervé HILLAIREAU (MC) - Laurence MOINE (CR) - Juliette VERGNAUD(MC)

Chef d’équipe Myriam TAVERNA (PR) _____________________________

- Marie-Claude ANMELLA (T) 50% - Isabelle LE POTIER (MC) - Claire SMADJA (MC) - Thuy TRAN-MAIGNAN (MC)

- Nacera ABOUD (Doc) - Jeanne BATAILLE (Doc) - Corentin BERARDET (Doc) - Cédric CROSNIER (Doc) - Claire GALLAND (Post-Doc) - Emmanuel JACCOULET (Doc) - Coralie RUEL (Doc) - James STOCKMANN (Post-Doc)

Chef d’équipe Florence PETIT-AGNELY (PR) _______________________

- Sandrine GEIGER (MC) - Nour-Eddine GHERMANI (PR) - Nicolas HUANG (MC) - Ghozlène MEKHLOUFI (MC) - Claire ALBERT (Doc) - Ali ALI (Doc) - Clémence GNAMAN N’GUESSAN (Doc) - Thomas ISOARDO (IE) - Zeineb MAAROUFI (Doc)

-Anne BELVERT (IE) - Tanguy BOISSENOT (Post-Doc) - Alexandre BORDAT (Doc) - Federica COSTAMAGNA (IE) - Sophie HOUVENAGEL (Doc) - Emilie LANGLOIS (IE) - Mathilde LORSCHEIDER (Doc) - Ludmila NASIMENTO (Doc) - Marion QUAILLET (Doc) - Emeline SERVAIS (IE) - Félix SAUVAGE (Doc)

Ingénierie Particulaire et Cellulaire à visée thérapeutique

Equipe 5

- Jean-Baptiste COTY (Doc) - Sophia MALLI (Doc) - An TAYLOR CASTILLO (Doc)

- Kawthar BOUCHEMAL (MC) - Christine VAUTHIER (DR)

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Chef d’équipe Gilles PONCHEL (PR)

Amélioration du passage des barrières par les molécules biologiquement actives

Equipe 6

- Guillaume BORT (Post-Doc) - Anaelle DUMAS (Post-Doc) - Jiao FENG (Doc) - Sandrine GOUAZOU (IE) - Elise GUEGAIN (Doc) - Gianpiero LAZZARI (Doc) - Julie MOUGIN (IE - Doc) - Arnaud PERAMO (Doc) - Marie ROUQUETTE (Doc) - Teresa SIMON YARZA (Post-Doc) - Dunja SOBOT (Doc) - Johanna TRAN (Doc) - Daniele VINCIGUERRA (Doc)

- Didier DESMAELE (DR) - Sinda LEPETRE (MC) - Simona MURA (MC) - Julien NICOLAS (DR) - Mariana VARNA (MC)

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Chef d’équipe Patrick COUVREUR (PR)

Nanomédicaments innovants pour le traitement des maladies graves

Equipe 7

Sarah VILLEBRUN (ASI) 50%

Magali NOIRAY (IE) Claire MERLET (ADT) 50 %

Physique pharmaceutique

Equipe 4

Service Rhéologie

Fatiha HAMZA (T) Patricia LIVET (ADT) Marie Claude ANMELLA (T) 50%

Equipe administrative et financière

Service des interactions moléculaires

Protéines et Nanotechnologies en Sciences analytiques

Equipe 3

Jean-Jacques VACHON (AI)

Stéphanie DENIS (AI)

Claire GUEUTIN (MC)

Equipe 2

Service de prototype

Service de culture cellulaire

Dominique MARTIN (T)

Secrétariat de direction

Sylvie ZEMMOUR (AI) (Responsable)

Administration et finances

Service d’analyse HPLC

Physico-chimie des Surfaces

Equipe 1

Hélène CHACUN (IE)

Service de radioactivité manipulation de produits à risques

Services communs

Directeur Pr Elias FATTAL

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

INSTRUMENTS OF UMR8612

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 1: TOOLS FOR THE ANALYSIS OF FLUID INTERFACES Supervisor: Pr. Véronique ROSILIO ([email protected])

Wilhelmy plate Tensiometer Surface and interfacial tension measurements

Langmuir trough – Brewster Angle Microscopy Interfacial behaviour of poorly soluble amphipathic compounds

Pendant drop tensiometer Calculation of surface energies

Quartz crystal microbalance (QCM-D) Evidence and quantification of interfacial interactions

Contact angle measurement Calculation of surface energies

Atomic force microscopy (AFM) Analysis of morphologies and physical properties of material surfaces

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 2: VARIOUS TOOLS Supervisor: Dr. Sylviane LESIEUR ([email protected]) Differential Scanning Calorimetry (DSC) Measurement of the amount of energy absorbed or released by a sample as it is heated, cooled or held in isothermal conditions. Scan rate: 0.05°C.min-1 →200°C.min-1 Determination of the temperature at which a reaction of a phase transition occurs. Determination of the associated enthalpy variation. High sensitivity thermal analysis.

Microcalix Coupled DSC with Small and Wide Angle X-ray Scattering Temperature range: - 20°C→ +180°C Scan rate: 0.01°C.min-1 →10°C.min-1 Can be used online with a laboratory/a synchrotron Xray source

Densimeter Liquids and colloidal dispersions Temperature control ±0.01°C Non destructive Need 1ml of sample Precision (10-6 g/ml)

Size Exclusion Chromatography (SEC) Aqueous phase Photodiode array detector Refractive index detector Water soluble polymers sizing Small colloids analysis  Micelles  Vesicles  Virus Others UV-Visible double-beam sceptrophotometer SPEX fluorescence spectroscopy Dynamic light scattering Technologies applied to lipid-based colloids (ultrasonication, extrusion, high-pressure homogenizer, freeze-drying) Ultra-centrifugation device

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 3: TOOLS FOR RHEOLOGY Supervisor: Pr. Florence AGNELY ([email protected]) Volume and interfacial rheology Viscosity as a function of share rate Material evolution over time Transition studies (i.e. gel) Capillary viscometry Specific or intrinsic viscosity

Texture analyzer Syringeability Adhesion to (mucous) membrane Penetrometry

Atomic Force Microscopy (S. Geiger, I. Canero Infante) High-resolution imaging of surface materials:  Topographyin air and liquid  Quantitative local mechanic , electric, magnetic properties

Drop tensiometer (TECLIS, tracker) interfacial tension, interfacial rheology, contact angle

Laser diffraction Material size: suspensions, emulsions, dry powders Material sizing technique for materials ranging from hundreds of nanometers up to several millimeters

X-ray diffraction (NE Ghermani) Crystal structure and high resolution crystallography for electron density studies Others Osmometer Conductimeter

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 4: VARIOUS TOOLS Supervisor: Pr. Myriam TAVERNA ([email protected])

NanoDrop Spectrophotometer 0.7 μL to 5 μL sample volume Suitable for wavelength range from 190nm to 1100nm

Absorbance microplate reader ELISA assays, protein and nucleic acid quantification or enzyme activity assays

Fluorescence microplate reader Protein and nucleic acid quantification or enzyme activity assays

Capillary electrophoresis  Electrophoretic mobility  Electroosmotic flow High separation efficiency Determination of apparent complexation constant by capillary affinity electrophoresis (ACE) Application to the determination of binding constants of vasoactive intestinal peptide to poly(amidoamine) dendrimers

Capillary electrophoresis - Q-TOF mass spectrometry Hyphenation High mass accuracy and high resolution instruments Ideal for peptide sequencing, determination of charge state and protein mass measurements

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 5: VARIOUS TOOLS Supervisor: Pr. Elias FATTAL ([email protected])

Spray drying Therapeutic applications for lung delivery Fast drying of droplets obtained by atomization of a solution/suspension

Multi-stage Liquid Impiger (MSLI) Aerodynamic characterization of dry powder Air flow-rate60 L/min, for 4 s, equivalent to 4L Liquid on each stage mimicks lungs humidity and solubilizes the dry-powder

Liposome extruder Liposomal formulation Several of different cut-off size filters, tune the number of passages

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 6: TOOLS FOR NP PREPARATION AND STUDY OF BIOLOGICAL INTERACTIONS Supervisor: Pr. Gilles PONCHEL ([email protected])

Scale-up of colloidal system preparation by nanoprecipitation  Liposomes  Nanocapsules  Nanoemulsions  Nanospheres Responsible Dr Kawthar Bouchemal ([email protected])

Immunoelectrophoresis: 2D gel electrophoresis Tools for investigating C3 complement activation:  First dimension: Molecular weight  Second dimension: Antibody affinity Responsible Dr Christine Vauthier ([email protected])

Ussing chamber Measuring membrane properties



Pharmacology Study of the passage of ions, water, electrolytes, nutrients and xenobiotics through barriers (intestinal, pulmonary or mucous membranes, skin, cornea…) Study of drug metabolism

 

Biopharmacy Evaluation of drug formulation (permeability) Study of drug vectors (nanoparticles,



Others Turbiscan Optical microscopy

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TEAM 7: VARIOUS TOOLS Supervisor: Pr. Patrick COUVREUR ([email protected])

Non-Invasive In Vivo Imaging Non-invasive - Bioluminescence Imaging and Fluorescence Imaging  No need to sacrifice the animal to perform the measurements  Longitudinal studies on the same mouse  Increase of statistical power of the model: each animal serves as its own control Significant reduction of the number of animals required to complete the study allowing to abide by the 3Rs (reduction, refinement and replacement) guiding principles for the use of animals in research. Not available for rats Not available for infected animals (No possibility to decontaminate the instrument) System available at the Animal facility, Located in the Immunodepressed room (Z18) Responsible Dr Simona MURA ([email protected])

Size Exclusion Chromatography (SEC) Organic phase (Chloroform, Dimethylacetamid) Refractometer detector, UV-detector and Multi-angle light scattering detector Polymer analysis  Molar mass  Dispersity Responsible Dr Julien Nicolas ([email protected])

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

MUTUAL INSTRUMENTS Zetasizer Nano Size and Zeta potential measurements Fixed scattered angle at 173° vs 90° Electrophoteric mobility and electroosmotic flow Supervisor: Dr. Gillian BARRATT and Dr Sylviane LESIEUR ([email protected] and [email protected])

Laser diffraction Material size: suspensions, emulsions, dry powders Material sizing technique for materials ranging from hundreds of nanometers up to several millimeters High sensitivity (only 100 mg are needed to perform size measurements) Supervisor: Dr. Nicolas TSAPIS ([email protected])

Microscopy Direct or Fluorescence DAPI (UV)/Fluorescein/Rhodamin Useful for observing systems before confocal microscopy Supervisor: Dr. Nicolas TSAPIS ([email protected])

Isothermal Titration Calorimetry Binding constant (Kb) Stoechiometry of binding (n) Thermodynamic parameters of binding ΔH and ΔS Supervisor: Magali NOIRAY ([email protected])

HPLC (4 chains -Waters) Reverse Phase (C18 column ) UV or Fluo detector Possibility of normal phase (anticipation of manipulations and crucial wash !)  Encapsulation rate  Release kinetics  Behavior of new forms for animal models Supervisor: Claire GUEUTIN ([email protected])

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

Ultracentrifuge Speed range 10000to 40000 rpm Room temperature or 4°C rotor Purification, concentration Supervisor: Stéphanie Denis ([email protected])

UV spectrometer analytical chemistry for the quantitative determination of different analytes, such as transition metal ions, highly conjugated organic compounds, and biological macromolecules. Spectroscopic analysis is commonly carried out in solutions

Fluorescence spectrometer Quantification of fluorescent organic compounds A complementary technique of absorption spectroscopy Supervisor: Kawthar Bouchemal ([email protected])

FTIR spectrometer Used to obtain an infrared spectrum of absorption or emission of a solid, liquid or gas Supervisors: Hervé Hillaireau and Julien Nicolas ([email protected] and [email protected])

Freeze dryer wide spectrum of potential applications: Preserving the characteristics of the original substances Preserving the original form Conditioning the material Sample preparation for chemical or biochemical analysis Supervisor: Stéohanie Denis ([email protected])

Access to regulated experiments Animal responsible: Dr Gillian BARRATT Radionuclide responsible: Hélène CHACUN Cell culture responsible : Stéphanie Denis

Doc and Post-Doc Day – Institut Galien Paris-Sud – June, 20th 2016

TECHNICAL

PLATEFORMS: IPSIT

CIBLOT Screening to identify small molecules that could be used as molecular probes for studying and/or modulating biological functions and to discover drug candidates

PHIC Immunopathological analysis of genetically modified mice and inflammatory animal models of human diseases and/or haematological

PLAIMMO Phenotype of cells by detection of intracellular and membrane molecules using flow cytometry

INTERMOL Molecular interactions

SAMM Mass spectrometry for structural analysis and quantification of small molecules and lipidomic profiling

ANIMEX Lodging and functional exploration of small animals

TRANS-PROT Qualitative and quantitative analysis of the transcriptome and proteome from cells and tissues of various origins

MIPSIT Imaging with confocal microscopy