Lauryl-gemcitabine loaded nanomedicine hydrogel for the local treatment of glioblastoma

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(1)3rd SFNano Annual Meeting Program & Abstract Book. December 12-14, 2016 Hotel IBIS - Clichy-Batignolles, Paris | France.

(2) CONTENTS. WELCOME LETTER. 3. LOCAL ORGANIZING COMMITTEE & SCIENTIFIC COMMITTEE. 5. PROGRAM IN A GLANCE. 6. MONDAY, DECEMBER 12, 2016. 7. SESSION #1 – Design & Characterization of Nanosystem. 8. TUESDAY, DECEMBER 13, 2016. 20. SESSION #2 – Macromolecule Delivery & Nanomedicine. 22. SESSION #3 – Nanomedicine & Imagering. 31. SFNano Young Session. 39. WEDNERDAY, DECEMBER 14, 2016. 47. SESSION #4 – Nanomedicine & Cancer. 48. SESSION #5 – Nanomedicine & Severe Infectious Diseases. 58. POSTER LIST. 65. LIST OF PARTICIPANTS. 136. GTRV & SFNano Awards. 147. 2.

(3) WELCOME LETTER. Société Française de Nanomédecine. Dear colleagues, dear friends, It is my great pleasure to welcome you to this third annual meeting of the French Society for Nanomedicine. It is our constant prospect to provide you with the best quality event in order to gather our forces and strengths in nanomedicine. Your presence in this event shows that the nanomedicine in France is strong, with diverse skills. Thanks to all our members, we can be heard on the French scientific landscape and reinforce the potential of nanomedicine. SFNano proposed you to gather in Paris and to organize conference and hosting at the same place, a hotel was chosen, which allows fostering interactions and collaborations. Together with Cnano Ile de France, we chose to dedicate the first session to the fundamentals of nanomedicine, the nanoobjects conception and characterization. The sessions on macromolecule delivery, cancer therapy and imaging which had received a great success last year were continued. A new session on the interest of nanomedicine for infectious disease was proposed by Dr R. Gref, who participates in the European Marie Curie ITN Cyclon Hit based on multifunctional cyclodextrin-based nanocarriers to overcome therapeutic drawbacks. You have been 227 to respond and join this program, and we received 119 abstracts from which 34 were selected for short oral communications. You will understand that we have shortened the duration of the oral presentations in order to allow many of you to communicate on your work and facilitate the discussions. We are very honored that speakers from Merck, Sanofi and Guerbet accepted our invitation to give us their feeling on the future of nanomedicine. We also dedicated a time for biotech presentations. If you do not know them, we will discover Carlina Technologies, Theraguix, Life Scientis, Atlangram, Bioaxial and Cell Constraints. Moreover, as you all know, nanomedicine characterization remains an issue, so do not miss the opportunity to talk with the representatives of the up-to-date equipment providers Malvern Instruments, Beckman Coulter, Izon, Schaefer, Anton Paar, Mettler Toledo who are thrilling to help you performing the best-quality research. For imaging, iThera and Visualsonics accepted our invitation and will be there to provide you with their best advices for experimental design. To further enrich our nanomedicine community, we are very attentive to young scientists and their future. Therefore, SFNano has accepted to support the European Nanomedicine Master which has been created by Prof K. Andrieux in Paris Descartes University and will start in September 2017. Moreover, the SFNano has missioned three young scientists together with Dr Sancey from the board, to propose means to gather young scientists working in nanomedicine and foster a network profitable for all. Therefore, SFNano is proud to launch the Young SFNano group. Young people will. 3.

(4) have opportunities to create their own session in our next meeting in Bordeaux. Moreover, as you might have seen, SFNano has created a linked group called SFNano group to provide this networking for all of us. SFNano Linked in group is not only for SFNano board, it is for all of you, to ask questions to the community, to post a new article that you published or a great review or article of interest, a post-doc or PhD position that you would like to fill, a job search…. This is your group, made for you, to favor your networking within the community. I think this is a very simple tool that all of us can use, and I would really like to thank Dr Hoarau and Dr Couffin for their strong contribution to SFNano communication. SFNano is doing his best to represent our community, but we really need you to participate in this dynamic of the nanomedicine community. Next year, we will organize a nanomedicine session within the congress RITS (Research in Imaging and Technology for Health of the French Society of Biological and Medical - SFGBM) which will be held in Lyon from March 27-29th, 2017. We will continue to be associated to the European Nanomedicine Meeting together with the British Society for Nanomedicine. It will be in London from April 3rd to 4th. For those of you working on nucleic acid delivery, Prof Lebleu is currently organizing an excellent summer school in La Grande Motte from June 7-9th, 2017. The highest research leaders within universities and companies have been invited. This promises to be an excellent quality summer school which should not be missed. Finally, the fourth annual meeting will be held in Bordeaux in December 5-7th, 2017. Please, save the dates. Tremendous speakers have been invited for all sessions and I would like to thank them to have accepted our invitation and all our partners for the different sessions, Cnano for the session synthesis and characterization, the European Society for Molecular Imaging (ESMI) and FLI for the imaging session, the American Metastasis Research Society (MRS) and the Canceropole Grand SudOuest for the cancer session. I also would like to thank the financial support from Sanofi, Bracco, Cnano, ESMI, FLI, GSO, Paris Descartes University, Paris Orsay University and Cyclon Hit. SFNano would like to thank its panel of scientific experts involved this year in program selection and abstract reviewing: Drs Ashford, Bazile, Couvreur, Menager and Scherman. SFNano would like to thank the local committee for the thorough organization of this annual meeting, many thanks to Drs Chaneac, Chauvierre, Richard, Greco, Gref, Tsapis, in particular to Dr Brun for the management of all the organization. I would also personally like to thank the people from the SFNano board constantly involved. We hope that our implication and devotion will be of great help for our community now and in the future.. Dr Nathalie MIGNET, President of SFNano. 4.

(5) LOCAL ORGANIZING COMMITTEE & SCIENTIFIC COMMITTEE LOCAL ORGANIZING COMMITTEE Last name. First name. Organization. BRUN. Amanda. Université Paris Diderot. GRECO. Stéphanie. Sanofi. RICHARD. Cyrille. CNRS/Université Paris Descartes. GREF. Ruxandra. Université Paris Sud. CHAUVIERRE. Cédric. Université Paris 7. MIGNET. Nathalie. CNRS/Université Paris Descartes. CHANEAC. Corinne. CNRS/université Paris 6. Last name. First name. Organization. COUVREUR. Patrick. Université Paris Sud. BAZILE. Didier. Sanofi. SCHERMAN. Daniel. CNRS/Université Paris Descartes. MENAGER. Christine. UPMC. ASHFORD. Marianne. Astrazeneca. SCIENTIFIC COMMITTEE. 5.

(6) PROGRAM IN A GLANCE DECEMBER 12, 2016. DECEMBER 13, 2016. DECEMBER 14, 2016. 8h30 Registration. SESSION 2 - Macromolecule delivery & Nanomedicine. SESSION 4 - Nanomedicine & Cancer. Chaired by: M-P. Rols and D. Letourneur. 9h00 Invited speaker Quentin Pankhurst, London (United Kingdom) and Claire Wilhelm, Paris (France). 9h00. Invited Speakers : Simon Geißler, Darmstadt (Germany), Rachel Auzély, Grenoble (France). Chairpersons: C. Wilhem and E. Fattal. 10h00 Keynote speakers. 10h00 Keynote speakers 10h30 Coffee break & Poster Session A. 10h40 Coffee break & Poster Session B. 11h30 Keynote speakers. 11h30 Invited speaker Louis Buscail, Toulouse (France). Biotech Corner. 12h30 Registration 13h00 Poster installation - Session A 13h30 Opening session Nathalie Mignet (SFNano) and Corinne Chanéac (CNano IdF). 12h00 Olivier Meyer, Carlina Technologies Géraldine Leduc, NHTheraguix Jean-Luc Bridot, Life Scientis Georges Tabary, Bioaxial Amokrane Reghal, Atlangram. 12h20 Biotech corner : Rémy Brossel, Cell Constraints and Cancer. 12h30. 12h50. Lunch break. SESSION 5- Nanomedicine & Severe Infectious Diseases. SESSION 3 - Nanomedicine & Imaging. Chairpersons: R. Gref and P.Couvreur. Mixed session with the ESMI. Mixed Session with CNano. 14h00 Invited Speakers Tony Lahoutte, Dr, Jette (Belgium), Hak Soo Choi, PhD, Charlestown (US). 14h00 Invited Speakers Giuseppe Mantovani, Nottingham (UK), Georges Favre, (France) 15h00 Keynote speakers 16h00. Coffee break. 16h30 Keynote speakers 16h50 Sponsor Presentations 17h50 Simon Baconnier, EU-NCL (France) 18h00 Didier Bazile, PhD, Sanofi (France) 18h30 Welcome Cocktail and. Lunch break. 13h45 Poster Installation - Session B. SESSION 1 – Design & Characterization of nanosystem Chairpersons: C. Chanéac & N.Tsapis. 12h30 General Assembly of SFNano (French Society for Nanomedicine). Chairpersons: E. Allémann and L. Sancey. 15h00 Keynote speakers 16h10 Coffee break & Poster Session B. SFNano Young Session. 14h00 Invited Speakers Dominique Duchene / Thorsteinn Loftsson, Galien Institute, ChâtenayMalabry, France/ University of Iceland, Reykjavik (Iceland) 14h30 Keynote speakers 15h20 SFNano/CNano, SFNano/ESMI poster awards and Sanofi Awards 16h00 Final conclusion and Meeting Closure. Chaired by C. Al-Sabbagh and J. Lavaud 17h00 Young Session 18h10 Invited Speaker : Jean-Marc Idée, Guerbet (France) 18h50 Departure for Gala Diner. Poster Session A. 6.

(7) MONDAY, DECEMBER 12, 2016 12h30. Registration. 13h00 13h30. Poster installation - Session A Opening session Nathalie Mignet (SFNano) and Corinne Chanéac (CNano IdF). SESSION 1 – Design & Characterization of nanosystem with CNano Chairpersons: Corinne Chanéac & Nicolas Tsapis 14h00 14h30 15h00 15h10 15h20. 15h30 15h40 15h50. Mixed Session Session sponsored by. Giuseppe Mantovani, PhD, Lecturer in Biologics Delivery, UoN, Nottingham (U Kingdom) Sweet Immunomodulation Georges Favre, PhD, Business Developper, LNE, Trappes (France) The metrological challenge of nanomaterials characterization Dan Elgrabli Paradigm shift: Carbon nanotubes can be degraded by macrophages Fanny D'Orlyé Physicochemical characterization and biocompatibility studies of persistent luminescent nanoparticles for medical and diagnosis applications Mathieu Varache Development and validation of characterization methods for Lipidots®: a critical step for Bench-to-Bedside translation Jean-Baptiste Coty Development of a high throughput immunoelectrophoresis for the characterization of complement activation by nanomedicines Louis Beauté Design of light-sensitive polymersome for spatial and temporal controlled release Emilie Secret Bio-functionalized magnetic nanoparticles for remote control of differentiation and oriented growth of neuronal cells. 16h00. Coffee break. 16h30. Sylvie Begin Design of smart anisotropic magnetic nano-objects towards an image-guided therapy Flavien Sciortino New Strategy to Self-assemble Nanoparticles Into Hollow Nanocapsules. Characterization and Applications of Hybridosomes® Sponsor Presentations Malvern Instruments; Beckman Coulter, Izon, Anton Paar; Mettler Toledo; Schaefer, Nanolane Visualsonics, iThera Medical Simon Baconnier, Laboratoire de Caractérisation Européen EU-NCL. Didier Bazile, PhD, Global Head of Drug Delivery Technologies and Innovation, Sanofi, Gentilly (France) Translation of nanomedicines to proof-of-concept in human - An update Welcome Cocktail and Poster Session A. 16h40 16h50. 17h50 18h00 18h30. 7.

(8) SESSION #1 – Design & Characterization of nanosystems. 8.

(9) Sweet immunomodulators Francesca Mastrotto,1,2 Alan Salama,3 Luisa Martinez-Pomares,2 Giuseppe Mantovani1 1 School of Pharmacy, University of Nottingham, Nottingham, UK ² School of Life Sciences, University of Nottingham, Nottingham, UK Giuseppe Mantovani obtained his PhD in Chemistry from Scuola Normale Superiore of Pisa (Italy) in 2002 after graduating as an Industrial Chemist from the University of Bologna (Italy). During his PhD he developed a new class of Co(II) and Fe(II) non-metallocene catalysts for the oligomerisation of ethylene to alpha-olefins. In 2002 he joined the Haddleton group at the University of Warwick where he worked at developing functional polymeric materials prepared by controlled radical polymerisation. Applications spanned from bioconjugate chemistry, to protein therapeutics and glycoscience. This theme continued after his appointment as Lecturer at the School of Pharmacy at the University of Nottingham in 2009. Current research in the Mantovani’s Group lies at the interface of organic/macromolecular synthesis and materials/soft matter science with particular emphasis in the development of lectin-recogniting glycopolymers, synthetic biology and polymer-(poly)peptide biohybrid materials. Recent research focussed on the development of carbohydrate-based materials and their application in immunology, both in vitro and in vivo. G Mantovani is the author of over 50 peer-reviewed papers, 7 patents, with an h-index of 30 In Nature, glycans mediate a range of key processes, which span from fecundation, to parasitisation, and the mounting of immune responses.1 Intriguingly, to exert these biological functions carbohydrates typically need to be presented in multiple copies and with precise spatial orientation onto oligo- or macromolecular scaffolds. This is required to bind specific sugar-binding proteins (lectins), which often exist themselves within oligomeric structure, thus engaging with their carbohydrate ligands in a multivalent manner.2 Synthetic glycopolymers – that is, synthetic polymers with carbohydrate pendant units – are increasingly utilised within this context as probes to dissect complex biological pathways involving sugar-protein recognition.3 In this work, we show that when appropriately designed, synthetic glycopolymers can be designed that engage lectin receptors on key immune cells, macrophages and dendritic cells, and modulate various aspects of their activity. Mechanistic aspect on this modulatory effects in vitro – e.g. receptor binding aviditiy and intracellular trafficking - and in vivo will be discussed, along with preliminary results on application in a preclinical model of ischaemia reperfusion injury (IRI). References 1. Schnaar, R. L., Glycobiology simplified: diverse roles of glycan recognition in inflammation. J Leukoc Biol 2016, 99 (6), 825-38. 2. Mammen, M.; Choi, S.-K.; Whitesides, G. M., Polyvalent Interactions in Biological Systems: Implications for Design and Use of Multivalent Ligands and Inhibitors. Angewandte Chemie International Edition 1998, 37 (20), 2754-2794. 3. Kiessling, L. L.; Gestwicki, J. E.; Strong, L. E., Synthetic multivalent ligands as probes of signal transduction. Angewandte Chemie - International Edition 2006, 45 (15), 2348-2368.. 9.

(10) The metrological challenge of nanomaterials characterization Georges Favre, PhD - Business Developper, LNE, Trappes (France) Georges Favre, 35, obtained his PhD in Analytical Chemistry from the University of Evry Val d'Essonne in 2008 after graduating as a chemical engineer from Ecole Centrale Paris in 2005. During his PhD, he worked at CEA (French Alternative Energies and Atomic Energy Commission, a French government-funded technological research organisation) on the isotopic characterizaton of nuclear fuel. He had intervened for three years with French SMEs as a consultant to develop advanced materials (fire or corrosion-resistant materials, …) based in particular on nanomaterial additives. He joined LNE in 2011 to coordinate the laboratory's R&D in the fields of metrology in chemistry and nanosciences (2011-2014). In addition to its commercial activities regarding testing and calibration services, LNE (Laboratoire national de métrologie et d’essais – www.lne.eu) is also the French National Metrology Institute. LNE has on this basis the responsibility to develop the French references (methods, standards, …) for all kinds of measurements in order to improve the reliability of data and therefore help public authorities and French industry in the decision-making process. Georges Favre is responsible since 2015 of the development of LNE’s services offer in the field of nanomaterials (calibration, standards, characterization, …). He is in particular involved in the identification of the needs and the dissemination of the scientific outputs on the topic of nanomaterials characterisation. Since 2012 he is involved in the Board of the nanoMetrology Club, a French network created in 2011 and coordinated by LNE, the aim of which is to gather people from public authorities, academia and industry in order to find solutions to nanometrology issues, in particular through R&D projects or the organization of workshops. He became its President in 2015 and is also the contact of industrial partners in the frame of the NANOMET project (www.nanomet.fr) funded by the French Ministry of Industry. Commercial nanomaterials containing products are already part of our everyday life and among them nanomedicines are expected as a big step forward in the treatment of serious pathologies. Efficiency of these emerging products, production processes upscaling, control of potential side effects and associated development costs are however fully dependent on a good characterization of nanoparticles’ properties. Nanometrology, science of the measurement at the nano-scale, is however still in its early stages, while it would have a determining catalytic effect on the growth of nanomedicines market. EU-NCL provides today different tools to help nanomedicines developers better understand their products and relies on NIST (US) expertise regarding metrology. But the scope of the work is huge as metrologists have to develop new reference materials and primary measuring methods in order to establish metrological traceability routes for each quantity (i.e. a link between the measurements and corresponding SI units). This is the only way to obtain comparability of phys-chem and biological data among various laboratories or different techniques. For instance, measuring nanomaterials’ size with an appropriate accuracy remains a real challenge despite the high number of techniques available on the market. This has been demonstrated in a recent proficiency testing proposed by the Club nanoMétrologie. Each step of the measurement process (sampling; calibration/instrument qualification; data acquisition & data processing) has to be carefully carried out to avoid bringing uncontrolled analytical bias. Moreover, as no single technique can be considered to be ideal to accurately measure the nanomaterials size, current trend is to combine complementary techniques. This hybrid metrology approach allows taking advantage of main strengths of each technique leading therefore to data with increased confidence level.. 10.

(11) Paradigm shift: Carbon nanotubes can be degraded by macrophages Elgrabli Dan (1) 1 - MSC laboratory Paris Diderot University/CNRS (France) A plethora of applications based on carbon nanotubes (CNTs) in various fields such as electronics, energy storage and conversion, sensors, automotive or nanomedicine are being developed. Some CNTs have been associated with iron oxide nanoparticles for medical application such as magnetic resonance imaging, hyperthermia therapy, and magnetic manipulation. We previously reported the design of biocompatible functionalized multi-walled CNT (MWCNTs) filled with iron oxide NPs, the inner cavity of CNTs acting as a nanoreactor for in situ growth of ferrite nanoparticles.[1] In healthy animals and in vitro, CNTs are engulfed by macrophages but the macrophage capacity to degrade CNTs is still unclear.[2] In a recent study, we unraveled the molecular mechanisms leading to structural damages on MWCNTs engulfed by macrophages. We described a molecular pathway by which macrophages degrade functionalized MWCNTs designed for biomedical applications and containing, or not, iron oxide nanoparticles in their inner cavity (denoted Fe@MWCNTs or MWCNTs, respectively). Using in situ monitoring of ROS-mediated-MWCNT degradation by liquid-cell transmission electron microscopy, two degradation mechanisms induced by hydroxyl radical were extracted from these dynamic nanoscale investigations: a non-site-specific thinning process of the walls and a site-specific transversal drilling process on pre-existing defects of nanotubes.[3] Remarkably, similar ROS-induced structural injuries were observed on MWCNTs after aging into macrophages from one to seven days. Beside unraveling oxidative transformations of MWCNT structure, we elucidated a general biological pathway for MWCNT degradation by macrophages, involving NOX2 complex activation, superoxide production and hydroxyl radical attack, which highlights the critical role of oxidative stress in cellular processing of MWCNTs. Electron microscopy and Raman spectroscopy show that intracellularly-induced structural damages appear more rapidly for iron-free MWCNTs in comparison to Fe@MWCNTs, suggesting a role of iron in the degradation mechanism.[4] By comparing the molecular responses of macrophages derived from THP1 monocytes to both types of CNTs, we highlight a molecular mechanism regulated by Nrf2/Bach1 signalling pathways. In the light of the present studies, CNTs degradation in macrophages was clearly established and offer new possibilities to use these nanoparticles as therapeutic vector. References 1. Liu, X., Marangon, I., Melinte, G., Wilhelm, C., Ménard-Moyon, C., Pichon, B. P., Ersen, O., Aubertin, K., Baaziz, W., Pham-Huu, C., Begin-Colin, S., Bianco, A., Gazeau, F., Begin, D. (2014) Design of covalently functionalized carbon nanotubes filled with metal oxide nanoparticles for imaging, therapy, and magnetic manipulation. ACS Nano. 8, 11290-11304. 2. Elgrabli, D., Floriani, M., AbellaGallart, S., Meunier, L., Gamez, C., Delalain, P., Rogerieux, F., Boczkowski, J., Lacroix, G. (2008) Biodistribution and clearance of instilled carbon nanotubes in rat lung. Part Fibre Toxicol. 5, 20. 3. Elgrabli, D., Dachraoui, W., Ménard-Moyon, C., Liu, X. J., Begin, D., Begin-Colin, S., Bianco, A., Gazeau, F., Alloyeau, D. (2015) Carbon Nanotube Degradation in Macrophages: Live Nanoscale Monitoring and Understanding of Biological Pathway. ACS Nano. 9, 10113-10124. 4. Elgrabli, D., Dachraoui, W., de Marmier, H., Ménard-Moyon, C., Begin, D., Begin-Colin, S., Bianco, A., Alloyeau, D., Gazeau, F. (2016) Intracellular degradation of functionalized carbon nanotube/iron oxide hybrids is modulated by iron via Nrf2 pathway. Sci Rep. Submitted.. 11.

(12) Physicochemical characterization and biocompatibility studies of persistent luminescent nanoparticles for medical and diagnosis applications Ramírez-García Gonzalo (1) (2) (3), D'orlyé Fanny (1), Gutiérrez Silvia (2) (3), Martínez Minerva (2) (3), Mignet Nathalie (1), Richard Cyrille (1), Varenne Anne (1) 1 - Unite Technol Chim & Biol Sante (France), 2 - Univ Guanajuato, Dept Quim (Mexico), 3 - Univ Guanajuato, Dept Farm (Mexico) Zinc gallate nanoparticles doped with chromium: ZnGa1.995Cr0.005O4 (ZGO-NPs) are innovative persistent luminescence materials that can be re-excited through living tissues by visible photons, allowing in vivo imaging without any time constraints and avoiding auto-fluorescence signals. Modification of the ZGO-NP surface is a requisite for colloidal stability and particle stealthiness for proper in vitro studies and distribution in living organisms. Thus the characterization and optimization of a classic functionalization sequence, on going from hydroxyl groups to PEG chains at the nanoparticle surface, has been performed using dynamic light scattering, laser doppler electrophoresis and capillary electrophoresis. This thorough control in terms of hydrodynamic size, zeta potential, electrophoretic mobility, stability over time, and aggregation state guarantees ZGONP quality for subsequent studies on their biocompatibility and targeting properties. Actually it is necessary to develop new methodologies to characterize specific and non-specific protein/NP interactions in dynamic equilibrium (fast exchange kinetics). Thus, electrokinetic methodologies were evaluated on the study of a protein (albumin or apolipoprotein E)/ZGO-NPs model system and proved to be sensitive to slight changes in the association strength in terms of saturation degree, dissociation constant and degree of cooperativity at various ionic strengths. Concerning the toxicity studies, MDA-MB-231 and MCF-7 cells and mice were treated with a single administration of hydroxylated or PEGylated ZGO-NPs at various concentrations. In vitro studies with hydroxylated NPs revealed a dose-dependent inhibition of cell viability at high doses, which is well correlated with oxidative stress conditions and cellular uptake. Only very high concentrations of hydroxylated NPs after long-term exposition generated structural endoplasmic reticulum alterations and oxidative stress in mice liver as well as rising in white blood cell counts. No clinical response to PEGylated nanoprobe treatment was evidenced during our in vitro and in vivo experiments. This new generation of ZGO-NPs offers the promise of revolutionary tools for biological imaging and other biomedical applications. 12.

(13) Development and validation of characterization methods for Lipidots®: a critical step for Bench-to-Bedside translation Varache Mathieu (1), Ciancone Mathieu (1), Laffont Corentin (1), Navarro Fabrice (1), Couffin AnneClaude (1) 1 - Département Microtechnologies pour la Biologie et la Santé (France) For eight years, our research group has been developing lipid nanocarriers, known as Lipidots®, as a multifunctional platform for medical applications. A wide range of active pharmaceutical ingredients (APIs) and contrast imaging agents has been encapsulated into Lipidots®, enabling the delivery of therapeutic agents and the monitoring or diagnosis by fluorescence imaging. [1-2] As a nanocarrier, Lipidots® have been proved to possess a high colloidal stability over months. Particle size and viscosity of the core can be tuned, resulting in high payloads, controlled drug release profile and improved drug bioavaibility to appropriate biological sites. [3-4] In addition to size and surface charge of the nanomaterials, as key parameters requested by regulatory health agencies, a more deep understanding of nanoparticles composition, drug loading, drug localization and drug release kinetics is crucial to meet the requirements of final medicinal nanoproducts. The quantification and identification of Lipidots® components have been performed by using validated UPLC-ELSD methods and mass spectrometry. A « fingerprint approach » was established for Lipidots® nanoparticles based on lipid composition and content, enabling to follow up any evolution during manufacturing process and ageing of formulations. Moreover, in order to predict the in vivo behavior of the nanotherapeutics, we developed a suitable and transposable tool for separation and quantification of free and entrapped drugs/dyes. [5] Localization of different encapsulated drugs within the nanodroplets has been characterized by NMR spectroscopy and correlated with the loading profiles. HPLC quantification combined with spectroscopic techniques has been also routinely implemented and applied for the optimization of payload formulation and the localization of encapsulated drugs within the nanodroplets. Such extensive characterization in addition to the ability to scale up the manufacturing process of our carriers, allows identifying characterization criteria and specifications for Bench-to-Bedside translation of our nanotherapeutics. In this communication, we will present an overview of these characterization techniques, which represent a step forward industrial transfer, and an example of the development of standardized physical-chemical characterization methods for nanomedicines. References [1] Delmas T. et al., Journal of Colloid Science and Biotechnology 2012, 1, 1-10; [2] Navarro F. et al., Journal of Photochemistry and Photobiology B: Biology 2014, 130, 161169; [3] Delmas T. et al., Langmuir 2011, 27(5), 1683-1692; [4] Delmas T. et al., Journal of Colloid and Interface Science 2011, 360, 471-481; [5] Guillot A. et al., Pharmaceutical Research 2015, 32, 39994009.. 13.

(14) Development of a high throughput immunoelectrophoresis for the characterization of complement activation by nanomedicines Coty Jean-Baptiste (1), Varenne Fanny (1), Vachon Jean-Jacques (1), Vauthier Christine (1) 1 - Institut Galien Paris-Sud (France) Over the last few years, it was evidenced that further progress within nanomedicines could be made at the expense of a strong effort on the understanding of their interactions with biological entities [1]. Among them, the complement system represents the first barrier of immunity after intravenous injection. Thus, the evaluation of complement activation by nanomedicines has become an imperative step in order to better understand their biodistribution and safety profile [2]. Common methods used to evaluate the reaction of the complement system in presence of nanomedicines are either expensive (Elisa, Radioimmunoassay) or suffer from a certain variability of the response (CH50 assay) [3]. 2D-immunoelectrophoresis was suggested as a direct method to monitor complement protein C3 cleavage upon incubation with nanoparticles [4]. However, it was limited to comparative studies due to the low number of samples that could be run at once and the tediousness of the method (5 samples in 19h). We have revisited the classical immunoelectrophoresis method to propose a high-throughput analysis of the interaction between a nanomedicine and the protein C3. The new experimental modalities (MultiCrossed-IE) [5] 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 to characterize a nanomedecine towards its ability to trigger the activation of the complement system. References [1] S. Wilhelm et al. (2016), Analysis of nanoparticle delivery to tumours, Nature Reviews Materials 1, Article number: 16014. [2] M. A. Dobrovolskaia et al. (2008), Preclinical Studies To Understand Nanoparticle Interaction with the Immune System and Its Potential Effects on Nanoparticle Biodistribution, Molecular Pharmaceutics, 5, 487–495. [3] P. P. Wibroe et al. (2012), Complement Sensing of Nanoparticles and Nanomedicines, in Functional Nanoparticles for Bioanalysis, Nanomedicine, and Bioelectronic Devices. Washington DC: American Chemical Society. [4] C. Passirani et al. (1998), Interactions of nanoparticles bearing heparin or dextran covalently bound to poly(methyl methacrylate) with the complement system, Life Sciences, 62, 775–785. [5] J-B. Coty et al. (2016), Serial multiple crossed immunoelectrophoresis at a microscale: A stamp-sized 2D immunoanalysis of protein C3 activation caused by nanoparticles, Electrophoresis, 37, 17–18. Financial support: BPI France - Project NICE. 14.

(15) Design of light-sensitive polymersome for spatial and temporal controlled release. Beauté Louis (1), Macclenaghan Nathan (1), Lecommandoux Sebastien (1) 1 - Laboratoire de Chimie des polymères organiques (France) Polymersomes are robust artificial vesicles enclosing an aqueous solution. Over the past decade, polymersomes became a popular research topic in the fields of medicine and biotechnology [1,2]. They are engineered and customizable to meet various specifications for a variety of domain such as drug delivery, nanoreactors, or gene therapy. One major challenge is to design stimuli-responsible systems that can release loaded species (i.e. drugs) in a controlled manner, at a precise time and space. Among all the possible triggers that can be applied, light has been proposed as a versatile approach since many years, especially using conformational changes of some specific molecules [3]. Here, we present light-sensitive polymersomes which can undergo rupture upon irradiation, for specific cargo release. In the present work, we used a coumarin derivative molecule as a linker between two polymers to yield an amphiphilic diblock copolymer with photosensitive properties. Coumarin undergoes heterolytic bond cleavage after irradiation [4] leading to a separation of the two blocks causing the vesicle leakage or rupture. This photosensitive molecule has attractive features such as a large molar absorption coefficient, a good stability, fast release rates and fluorescent properties which allow facile monitoring. In addition, this coumarin derivative was designed in such as way that a wide variety of polymers could be conjugated, offering a large range of versatile light-sensitive block copolymers. We specifically report here on the coumarin synthesis followed by grafting of poly(trimethylene carbonate) (PTMC) and poly(ethylene oxide) (PEO). PTMC and PEO have been widely studied as materials for biomedical applications due to their low toxicity and biocompatibility. In addition, the self-assembly properties and the light-mediated rupture of the vesicles were studied. References [1] D. Discher Science, 2002, Vol 297 (5583), 967-973. [2] H. Oliveira, J. Thevenot, S. Lecommandoux, Nanomed. Nanobiotechnol, 2012, Vol 4, 525-546. [3] M-H. Li, P. Keller Soft Matter, 2009, Vol 5, 927-937 [4] P. Klán, T. Šolomek, C. Bochet, A. Blanc, R. Givens, M. Rubina, V. Popik, A. Kostikov, J. Wirz Chem. Rev, 2013, Vol 113, 119-191.. 15.

(16) Bio-functionalized magnetic nanoparticles for remote control of differentiation and oriented growth of neuronal cells Secret Emilie (1), Balloul Elie (2), Michel Aude (1), Monzel Cornelia (2), Dahan Maxime (2), Fresnais Jerôme (1), Ménager Christine (1), Siaugue Jean-Michel (1) 1 - PHysicochimie des Electrolytes et Nanosystèmes InterfaciauX (France), 2 - Laboratoire PhysicoChimie, Institut Curie, CNRS UMR168, Paris-Science Lettres, Université Pierre et Marie Curie-Paris 6, 75005 Paris (France) Neurodegenerative disorders, such as Parkinson's, Alzheimer's or Huntington's diseases are among the most common group of medical conditions in the world, and are expected to surpass cancer by 2040.[1] However no cure exists for such diseases at that time. Cell replacement therapy is among the most promising approach to treat neurodegenerative disorder. In the work presented here, part of the MAGNEURON European project,[2] we used magnetic nanoparticles that are biofunctionalized to trigger neurons' differentiation and growth along the direction of use of an external magnet gradient. Mature neurons would in term be re-implanted in the patient brain to replace degenerated neurons. To this goal, maghemite (γ-Fe2O3) nanoparticles were synthesized by an inverse co-precipitation process.[3] These nanoparticles were then used to synthesize γ-Fe2O3@SiO2 core-shell nanoparticles with size, charge and magnetization adjusted to obtain a good colloidal stability, render them injectable in cells, and facilitate intracellular motion. These magnetic nanoparticles (MNPs) were then functionalized with a HaloTag ligand in order to interact specifically with proteins able to trigger different pathways in the cell. MNPs were then microinjected in the cell and showed intra-cellular biased diffusion toward the micro-magnet. The magnet can then be used to displace target proteins attached to the MNPs inside the cell, and trigger signalling events such as actin polymerization at particular subcellular localizations.[4,5] References [1] World Health Organization, Neurological disorders: public health challenges (2006). [2] MAGNEURON : H2020 Future and Emerging Technologies (FET) project n°686841, http://magneuron.eu/ [3] Massart R., Preparation of aqueous magnetic liquids in alkaline and acidic media, IEEE Trans. Mag. (1981), 17, 1247–1248 [4] Etoc et al., Magnetogenetic control of protein gradients inside living cells with high spatial and temporal resolution, Nano Lett. (2015), 15, 3487−3494 [5] Etoc et al., Subcellular control of Rac-GTPase signalling by magnetogenetic manipulation inside living cells, Nat. Nano (2013) 8, 1937.. 16.

(17) Design of smart anisotropic magnetic nano-objects towards an image-guided therapy Begin Sylvie (1), Cottin Geoffrey (1), Mertz Damien (1), Pichon Benoit (1), Meyer Florent (1), Felder Delphine (1) 1 - Institut de Physique et Chimie des Matériaux de Strasbourg (France) In the field of the synthesis and functionalization of inorganic nanoparticles (NPs) for biomedical applications, most researches aim at developing multifunctional theranostic NPs which can both identify disease states and deliver therapy and allow thus following the effect of therapy by imaging. The current challenge for iron oxide based NPs is the design of NPs able to combine in one nanoobjects both magnetic hyperthermia (MH) and MRI with the best efficiency in order to reduce the dose injected in the patient. Ultra small iron oxide NPs are already commercially used as T2 contrast agent for MRI. The use of MH as a stand-alone or an adjacent therapy for cancer is closer to be a reality in every hospital thanks to the positive results achieved by the clinical trials carried out by MagforceTM(Germany) treating glioblastoma. Nonetheless, the need for direct intratumoral injection of large amounts of NPs to achieve a therapeutic effect only points out at the need of improving the available nanomaterials for MH. Different parameters may be varied to increase the effective heat loss of a ferrofluid such as size, shape anisotropy or composition, among others. Shape and aspect ratio may offer interesting possibilities as chain formation has been previously reported to increase heat loss. Furthermore there is also a need for evaluating the heating efficiency in cellular media as it may be different from that in solution. On that basis, plate-like, cubic and octopod shape NPs were prepared by thermal decomposition of home-made iron stearate, functionalised with dendron ligands to achieve aqueous suspensions and proved suitable for in vivo injection. MH performance was found to be shape-dependent with octopod-shaped NPs exhibiting the highest SAR values of 260 W.g-1 (f = 579 kHz, 8 kAm-1, ILP = 7.1 nHm2kg-1) or 960 W.g-1 (f = 796 kHz, 16 kAm-1, ILP = 4.8 nHm2kg-1). At the same time, their performance in MRI was investigated leading to relaxivity values of 16.9 and 405.5 mM-1 s-1 for r1 and r2, respectively, which was superior to that of commercial products like Resovist®. Finally, cell response was studied as a function of NP concentration and morphology, as well as under MH treatment. Therapeutic properties were also obtained by filling carbon nanotubes (CNTs) with ferrite NPs as heat mediator for photothermal ablation and as contrast agent for MRI respectively. They were capable of absorbing and efficiently converting NIR light into heat to generate thermoablative temperatures and cell lysis. They can be used as T2 agents for MR image-guided photothermal therapy. The obtained results open the possibility of using these systems as theranostic platform thanks to the exhibited performance in hyperthermia and MRI at the cell level.. 17.

(18) New Strategy to Self-assemble Nanoparticles Into Hollow Nanocapsules. Characterization and Applications of Hybridosomes® Sciortino Flavien (1), Thivolle Maxime (2), Jacobzcyk Hélène (3), Casterou Gerald (4), Eliat PierreAntoine (5), Troadec Marie-Bérengère (3), Kahn Myrtil (6), Chevance Soizic (1), Gauffre Fabienne (1) 1 - Institut des Sciences Chimiques de Rennes (France), 2 - Ecole Nationale Supérieure de Chimie de Rennes (France), 3 - Institut de Génétique et Développement de Rennes (France), 4 - Laboratoire de chimie de coordination (LCC) (France), 5 - PRISM-Biosit (France), 6 - Laboratoire de chimie de coordination (France) We will present a new and versatile strategy to self-assemble nanoparticles into hollow nanocapsules. This strategy is based on the existence of large scale (100nm) inhomogeneities existing in macroscopically miscible mixtures of organic solvent and water. We hypothesized that such droplets could template the formation of nanoparticle shell similarly to Pickering emulsion and that nanoparticles could stabilize this interface between liquids. After addition of a crosslinking and biocompatible polymer, removal of the solvent core, hollow capsules of diameter ca 150nm, coined Hybridisomes® are obtained. They were characterized using a full set of techniques including nanoparticle tracking analysis, transmission and scanning electron microscopy and liquid phase atomic force microscopy. Nanocapsules were easily obtained from Quantum Dots, superparamagnetic iron oxide and gold nanoparticles as well as mixture of it. The entrapment of a fluorescent dye was also demonstrated leading. Interestingly magnetic Hybridosmes® enable MRI contrast enhancement of tumors in vivo. The high versatility of these multifunctional platforms could have many interests in multimodal imaging, photodynamic therapy and lead to several applications in the field of nanomedicine.. 18.

(19) Translation of nanomedicines to proof-of-concept in human - An update Didier Bazile, Ph. D. Head of External Innovation - Sanofi (integrated CMC – New Products Program) As Head of CMC(1) External Innovation at Sanofi, Didier Bazile is in charge of the initiation and follow-up of programs and projects based on innovative Drug Delivery approaches (nanotechnology driven accumulation in tumors, oral delivery of peptides, nucleic acids delivery, depot formulations), in close collaboration with the Therapeutic Units, the CMC functions, DMPK and Safety. Didier Bazile held different positions in Drug Delivery Technologies and Pharmaceutical Engineering, in medium size and big pharmaceutical companies, from Discovery to filing and launch. He is the author/co-author of innovative patents and pioneering articles on nanotechnologies. Didier Bazile was an alumnus of the Ecole Normale Supérieure de Cachan in Biochemistry, made a Ph.D. in Pharmacology at the Institut Gustave Roussy (University of Paris VI) and received an Accreditation to Supervise Research in Pharmaceutical Sciences (University of Paris XI). 1: Chemistry Manufacturing and Control A wide variety of nanotechnology applications have been reported, from the simple increase of the surface/volume ratio inherent to the high level of dispersion of nano-materials, to the sophisticated design of multifunctional nano-carrier constructs delivering drugs at the specific tissular, cellular and sub-cellular levels (D. Bazile, 2014). Considering the risks associated with the innovative nanotechnology-based products, the development of nano-objects should be consistent with a translational approach (S. Stern et al., 2010), i.e. the ability to build in parallel (i) the product design and early development plan, (ii) the quality by design management principles and (iii) the regulatory strategy. This is particularly true when the time and space controlled drug release principles of nanotechnology-based products are significantly different from those described for standard pharmaceutical dosage forms. In such an instance, the anticipation and management of the risks, based on a robust multidisciplinary scientific rationale, are the backbone of the translational process. The understanding and control of the drug/nano-carrier association appeared as key to properly extrapolate preclinical data to human, as recently described for the taxanes formulations (L. Harivardhan Reddy and D. Bazile, 2014) and for PLA-PEG nanoparticles (L. Harivardhan Reddy and D. Bazile, 2016). In this context, we proposed a methodology to anticipate the fraction of nano-encapsulated drug after manufacturing, and after the dilution in blood following the administration (O. Diou et al., 2015). The proper evaluation of this fraction is of prior importance since its fate and biodistribution is thought to be significantly different from the free and protein-bound fractions investigated with standard sterile solutions. From an efficacy standpoint, the drug should stay associated to the carrier to be co-delivered in the tumor, and released according to a timescale consistent with the inhibition of cancer cells multiplication. At the same time, from a safety standpoint, the nano-encapsulated drug should not accumulate in undesired organs, in particular those known to capture the nano-carriers, such as the liver. The anticancer drug cabazitaxel encapsulated in PLA-PEG nanoparticles was chosen as a case study. We showed that the association of the drug to the nano-carrier can be satisfactorily assessed from a partition coefficient (Kp) of cabazitaxel between the PLA matrix and the suspending medium. The overall objective of this presentation is to propose an updated and integrated vision of the translation of nanomedicines to proof-of-concept in human and to illustrate how the integration of the physico-chemical aspects of the nano-assembly (including drug loading) and of the biopharmaceutical aspects of the drug routing is a funding principle prone to bring nanotechnology to the next level. D. Bazile, Nanotechnologies in drug delivery — an industrial perspective, J. Drug Delivery Sci. Technol. 24 (2014) 12–21. O. Diou, S. Greco, T. Beltran, D. Lairez, J.-R. Authelin, D. Bazile, A method to quantify the affinity of cabazitaxel for PLA-PEG nanoparticles and investigate the influence of the nano-assembly structure on the drug/particle association, Pharm. Res. 32 (2015) 3188–3200. L. Harivardhan Reddy, D. Bazile, Drug delivery design for intravenous route with integrated physicochemistry, pharmacokinetics and pharmacodynamics: illustration with the case of taxane therapeutics, Adv. Drug Deliv. Rev. 71 (2014) 34–57. L. Harivardhan Reddy, D. Bazile, Building the design, translation and development principles of polymeric nanomedicines using the case of clinically advanced poly(lactide(glycolide))–poly(ethylene glycol) nanotechnology as a model: An industrial viewpoint, Adv. Drug Deliv. Rev. (2016), http://dx.doi.org/10.1016/j.addr.2016.08.012. S. Stern, J. Hall, L. Yu, L. Wood, G. Paciotti, L. Tamarkin, S. Long, S. McNeil, Translational considerations for cancer nanomedicine, J. Control. Release 146 (2010) 164–174.. 19.

(20) TUESDAY, DECEMBER 13, 2016 8h30. Registration. SESSION 2 - Macromolecule delivery & Nanomedicine Chairpersons: Marie-Pierre Rols and Didier Letourneur 9h00 9h30 10h00. 10h10 10h20. Simon Geißler, Dr., Director Drug Delivery & Innovation, Merck, Darmstadt (Germany) Nanomedicine for therapeutic vaccination Rachel Auzély, Prof., Professor of Polymer Chemistry, CNRS, Grenoble (France) Smart delivery systems based on polysaccharide hydrogels and nanogels Jørgen Kjems Self-assembled nucleic acid-peptide-lipid devices with improved cell targeting and uptake, intracellular trafficking and in vivo pharmacokinetics for delivery of biologics Viorel Simion microRNA delivery using lipo-polyplexes: a comprehensive overview of intracellular trafficking, endosomal escape, RISC loading, P-body processing and gene repression kinetics Cédric Chauvierre Thrombolytic therapy based on P-selectin targeted polymer nanoparticle. 10h30. Coffee break & Poster Session A. 11h30. Dario Carradori New drug delivery system to target neural stem cells Emilie Bayon Vectorization of Protein Antigens by Lipid Nanocarriers for Vaccine Formulation Anaïs Vaissière RICK, an improved CPP for siRNA delivery via nanoparticle formation. 11h40 11h50. Biotech Corner Session co-organised with 12h00. Olivier Meyer, Carlina Technologies Jean-Luc Bridot, Life Scientis Amokrane Reghal, Atlangram. 12h30. Lunch break. 13h45. Poster Installation - Session B. SESSION 3 - Nanomedicine & Imaging session with the ESMI Chairpersons: Eric Allémann and Lucie Sancey 14h00 14h30 15h00 15h10. 15h20. Géraldine Leduc, NHTheraguix Georges Tabary, Bioaxial. Mixed Session sponsored by. Tony Lahoutte, Dr, Head of Department, Nuclear Medicine, Jette (Belgium) Molecular targeting using nanobodies Hak Soo Choi, PhD, Associate Professor, MGH, Charlestown (United States) Targeted Contrast Agents for Bioimaging and Nanomedicine Raphaël Lévy Silica-coated gold nanorods enhance the sensitivity of photoacoustic detection of labelled stem cells in vivo Christophe Mingotaud Assembly of Double-Hydrophilic Block Copolymers triggered by Gadolinium Ions: New Colloidal MRI Contrast Agents Jonathan Lavaud Multispectral photoacoustic imaging of tumors in vivo. 20.

(21) 15h30. 15h40 16h10. Jonathan Pansieri Gd-nanoparticles functionalization with Pittsburgh Compound B, antibody fragment B10AP or specific peptides for amyloid plaques detection Sylvain Gioux, PhD, Professor, UNISTRA, Illkirch (France) Real-time Quantitative Optical Imaging for Surgery Coffee break & Poster Session B. DECEMBER 13, 2016 SFNano Young Session Chairpersons: Chantal Al-Sabbagh and Jonathan Lavaud 17h00 17h10 17h20 17h30 17h40 17h50. 18h00. Young Session Introduction Max Piffoux Monitoring microvesicles and exosomes dynamics at the nanoscale by liquid-cell TEM Sébastien Piant How to actively target Melanoma in vivo with dendronized iron oxide nanoparticles? Alberto Curcio Shape controlled Fe2O3@CuS nanohybrids: a multi-functional platform combining magnetic hyperthermia, photothermia and photodynamic therapy Jonathan Bruniaux In vitro transfection of targeted stealth magnetic siRNA nanovectors (TS-MSN) in breast cancer cells Pancani Elisabetta Development of an advanced label-free technique to study nanoparticles internalization in cells to fight intracellular bacteria Charles Skarbek New oxazaphosphorine prodrug's as nanomedicine against cancer. 18h10. Jean-Marc Idée, Dr, Guerbet (France) Development of contrast agents from the industrial point of view. 18h40. Departure for Gala Diner. 21.

(22) SESSION #2 – Macromolecule Delivery & Nanomedicine. 22.

(23) Nanomedicine for therapeutic vaccination Simon Geissler Director Drug Delivery & Innovation, Merck, Darmstadt (Germany). Simon Geissler, 36, joined Merck in 2006.His work focused on preclinical pharmaceutical development and the translation of research into technologies addressing drug delivery needs of Merck pipeline comounds. Since 2015 he is leading a R&D group for drug delivery for small and large molecules.. Therapeutic vaccines present a promising option for advancing the therapy of cancer. The talk presents several examples of nanoparticulate platforms, namely liposomes and polymeric nanoparticles, and the use in vaccine applications. Apart from s.c. injection also other routes of administration, such as inhalation and peroral administration, are highlighted. Available preclinical data support further efforts in these therapeutic platforms.. 23.

(24) Smart delivery systems based on polysaccharide nanogels Rachel Auzély-Velty Grenoble Alpes University, CERMAV-CNRS, F-38000 Grenoble, France Rachel Auzély-Velty, 44, obtained her Ph.D. in Organic Chemistry from the University of Rennes I in 1997 after graduating as a chemical engineer from Ecole Nationale Superieure de Chimie de Rennes (ENSCR) in 1994. During her Ph.D., she worked at the laboratoire de Chimie des Substances Naturelles (ENSCR) under the supervision of Prof. Daniel Plusquellec. She developed the synthesis of analogues of glycolipids found in archaebacteria and investigated their self-organization properties in water. During a two-year post-doctoral fellowship at the Commissariat à l’Energie Atomique (CEA) in Saclay under the supervision of Dr. Bruno Perly, she acquired further knowledge in amphiphilic carbohydrate derivatives and liposomes by designing cholesterylcyclodextrin conjugates and investigating their incorporation into phospholipid membranes. Based on her expertise in carbohydrate chemistry and oligosaccharides-based amphiphiles, she was recruited as a CNRS researcher in CERMAV-CNRS (Grenoble) in 1999 to develop smart biomaterials based on chemically modified water-soluble polysaccharides. In 2005, she became a full Professor at the University in Grenoble (University Grenoble Alpes, UGA), while continuing her research on the development of well-defined biomaterials based on polysaccharides, especially hyaluronic acid. Potential target applications include controlled and targeted drug delivery, surface modification, tissue engineering and cell biology. Self-assembled gel nanoparticles (also called nanogels) made of hydrophobically modified polysaccharides are particularly attractive for drug delivery. Their size can be easily varied, their interior network comprising hydrophobic domains can be used to incorporate poorly water-soluble drugs and their hydrophilic shell can be exploited to control their biological fate and targeting ability. In this regard, several studies have focused on the use of hyaluronic acid (HA) for the design of anticancer drug carriers due to the interesting biological properties of this natural polysaccharide.1,2 Indeed, HA is a glycosaminoglycan specifically recognized by the CD44 receptor that is overexpressed by several cancer cells. Our group previously demonstrated that grafting onto hyaluronic acid (HA) thermoresponsive ethylene-glycol based copolymers allowed temperature-triggered assembly of HA into nanogels.3,4 These gel nanoparticles possess many interesting features for drug delivery, like: facile formation, easy loading of hydrophobic molecules, and degradation behavior due to the inherent biodegradability of HA. In addition, these platforms can be carefully tuned for drug delivery by tailoring the thermosensitive copolymer architecture and the degree of substitution of the modified HA. In this respect, we recently focused our efforts on engineering new HA-copolymer conjugates to produce nanogels exhibiting a favorable balance between long circulation and moderate stability to release their payload at the targeted location (i.e. tumor site). We will show how precise control over HA functionalization and copolymer architecture can induce original physico-chemical and biological properties of HA-based nanogels. 1. F. Dosio, S. Arpicco, B. Stella, E. Fattal. Adv. Drug Delivery Rev. 2016, 97, 204-236. 2. N. V. Rao, H. Y. Yoon, H. S. Han, H. Ko, S. Son, M. Lee, H. Lee, D.-G. Jo, Y. M. Kang, J. H. Park. Expert Opin. Drug Delivery 2016, 13, 239-252. 3. T. Fernandes Stefanello, A. Szarpak-Jankowska, F. Appaix, B. Louage, L. Hamard, B. G. De Geest, B. van der Sanden, C. V. Nakamura, R. Auzély-Velty. Acta Biomater. 2014, Ahead of Print. 4. J. Jing, D. Alaimo, E. De Vlieghere, C. Jerome, O. De Wever, B. G. De Geest, R. Auzély-Velty. J. Mater. Chem. B 2013, 1, 3883-3887.. 24.

(25) Self-assembled nucleic acid-peptide-lipid devices with improved cell targeting and uptake, intracellular trafficking and in vivo pharmacokinetics for delivery of biologics Kjems Jørgen (1) 1 - Interdisciplinary Nanoscience Center (Denmark) Non-viral delivery of oligo nucleotide and protein based drugs in vivo is a challenging task that can be significantly improved by engineered lipid or polymer based nanocarrier systems. However, the assembly process is usually empirically optimized and without rigid control of size and composition. The application of self-assembled chemically modified nucleic acid structures driven by predictable Watson and Crick base pairing has enabled construction of highly defined nanodevices that in a robotic fashion can sense the environment, compute the signal and adjust a functional responds. Spatial nanometer control of oligonucleotides fused to peptides, sugars, lipids or small molecules has enables us to study multivalency and pattern recognition for improved delivery. In particular, we have constructed and studied self-assembled structures capable of sensing complex mixtures of microRNA, delivering functional nucleic acids to specific cells and compartments and control activity of enzymes. This field opens up for “intelligent” diagnostic robots that for instance can diagnose complex diseases at cellular level and impose a tailor made treatment at the cellular level. To achieve clinical relevance in animals and humans we have introduced chemical modifications to improved stability and pharmacokinetics and obtained successful targeted delivery to organs and diseased tissue.. 25.

(26) microRNA delivery using lipo-polyplexes: a comprehensive overview of intracellular trafficking, endosomal escape, RISC loading, P-body processing and gene repression kinetics Simion Viorel (1), Garcion Emmanuel (2), Pichon Chantal (1), Midoux Patrick (1), Baril Patrick (1) 1 - Centre de biophysique moléculaire (France), 2 - Micro et nanomédecines biomimétiques (France) Development of functional miRNAs monitoring systems is required for optimizing RNAi nanoformulations, miRNA chemical modifications, intracellular trafficking, RISC loading and activity. Here we developed a positive readout miRNA monitoring system called RILES, for RNAi-Inducible Luciferase Expression System, and stably integrated this bioluminescence probe in different cancer cell lines for monitoring the biological activity of transfected miRNAs. Using RILES cells, we determined the transfection kinetics of miRNA duplexes complexed with our home-made LPRi lipopolyplexes (which is ternary complexes formed between imidazole/imidazolium cationic liposomes, histidinylated-LPEI and miRNA mimics) and compare its efficiency with the golden standard Lipofectamine RNAiMax transfection reagent. We demonstrated that the LPRi formulation acts as a long-lasting miRNA releasing agent with low toxicity while the RNAi-Max formulation delivered quickly the miRNA mimics, with less efficiency. To get insight about the intracellular mechanisms responsible for different kinetics of miRNA transfection using the two nanoformulations, we examined the endocytosis pathways, intracellular trafficking, endosomal escape, RISC processing in the P-/GW-bodies and finally miRNA activity monitored by the RILES. Our confocal microscopy analysis demonstrated that LPRi entry into the cells through the caveolin-dependent endocytosis, with further trafficking to early and late endosomes, and especially to multivesicular bodies. We found intrinsic differences in term of endosomal escape between these two formulations, characterized by a fast endosomal release from RNAiMAx after just few hours, and a slow but constant release from LPRi, that correlate remarkably well with the biological activity of miRNA monitored by the RILES. We then examined for a potential colocalization of miRNA mimics with AGO2, a marker of RISC machinery and with P-/GW-body marker proteins such as GW-182, RCK-p54 and DCP1a. We found that miRNA mimics delivered byLPRi colocalized with these molecular partners of miRNA biogenesis, in a time-dependent manner, that correlated again well with their biological activity monitored by the RILES. To explore the functional implication of these proteins in the biological activity of transfected miRNAs, we silenced their expression and found that GW-182 and RCK-p54 were the two most implicated proteins, while DCP1a appeared less active. We are currently translating this RILES monitoring system in different animal tumor models to monitor the in vivo delivery and functionality of miRNAs using different lipo- and polyplexes nanoformulations. We believe that the present study provides a comprehensive understating on intracellular delivery of miRNA formulations which is relevant to optimize the design of novel miRNA nanomedicines with efficient trafficking to the RISCs machinery for optimal biological activity, and with obvious application in the field of miRNA based therapy.. 26.

(27) Thrombolytic therapy based on P-selectin targeted polymer nanoparticle Juenet Maya (1), Aid-Launais Rachida (1), Ollivier Véronique (1), Berger Alice (1), Li Bo Letourneur Didier (1), Chauvierre Cédric (1). (1),. 1 - INSERM U1148 (France) Injection of tissue plasminogen activator (tPA) remains the standard treatment for thrombolysis. However, high doses have to be administered with the risk to cause hemorrhages [1]. In our work, tPA-loaded nanoparticles are investigated to decrease the injected amount while keeping the same efficiency by ensuring a specific delivery. For this purpose, we synthesized polymer nanoparticles targeted against P-selectin. This protein is indeed a biomarker of activated platelets composing the thrombus. Fluorescent polysaccharide-poly(isobutylcyanoacrylate) nanoparticles were synthesized by redox radical emulsion polymerization. They were functionalized with fucoidan (Fuco-NPs) as this anionic polysaccharide shows a nanomolar affinity for P-selectin [2]. Carboxymethyl-dextran was used as control (Control-NPs). To validate the targeting strategy, an in vitro flow assay was set up. Human whole blood was injected into collagen coated micro-channels to induce platelets activation and aggregation. Adhesion of NPs onto aggregates in venous and arterial conditions was quantified under fluorescence microscopy. tPA (Actilyse®) was then loaded by adsorption. The drug-related activity of the suspensions was checked in vitro and used to normalize the doses. Finally, an in vivo study was performed in a mouse model of mesenteric thrombosis induced by iron chloride. Both NPs were similar in size, surface charge and fluorescence intensity. Fuco-NPs bound significantly more than Control-NPs to platelet aggregates in venous and arterial conditions confirming the potential of fucoidan as a targeting agent (Figure A). The preclinical study showed that only FucoNPs-tPA were able to induce complete thrombolysis (4 mice out of 11) at a dose four times lower than the recommended one for murine models (Figure B). This result reinforces the choice of an active targeting approach, compare to other delivery strategies [3]. Fucoidan-functionalized nanoparticles were efficient as carriers for thrombolysis enabling to decrease the injected tPA dose and therefore the risk of undesirable side effects. Although the variability between individuals is high in the developed model, this study underlines the great potential of targeted nanomedicine to fight thrombotic diseases References [1] J. Alvazer-Sabin et al. Lancet Neurology 12(7) 52013 [2] L. Bachelet et al. Biochimica et Biophysica Acta 1790(2) (2009) [3] M. Varna et al. Future Science OA 1(4) (2015).. 27.

(28) New drug delivery system to target neural stem cells Carradori Dario (1), Saulnier Patrick (3), Preat Veronique (2), Des Rieux Anne (2), Eyer Joel (3) 1 - Louvain Drug Research Institute (Belgium), 2 - Advanced Drug Biomaterials,Université Catholique de Louvain (Belgium), 3 – Université d’Angers. Delivery. and. The in situ differentiation of endogenous neural stem cells (NSC) represents a potential therapeutic strategy to replace injured neuronal cells and to treat neurodegenerative diseases. Nevertheless, no work based on this approach has yet reached the clinical phase. The lack of NSC-targeting molecules primarily promotes the development of non-selective systems with limited effects on NSC differentiation. The aim of our work was to produce a drug delivery system able to selectively target endogenous NSC. The peptide NFL-TBS.40-63 (NFL) shows specific interactions with brain NSC, where it affects their properties and induces their differentiation [1]. Consequently, we produced an NFL-based drug delivery system to target those cells. The peptide was adsorbed on DiD-labeled lipid nanocapsules (LNC, NFL-LNC) and characterized by dynamic light scattering. NFL-LNC targeting efficiency was evaluated on brain and spinal cord NSC. NFL-LNC were incubated with primary NSC cultures, in vitro, and injected either in adult rat's brain or spinal cord, in vivo. The determination of the targeting efficiency was performed by FACS for in vitro experiments, and by immunohistochemistry for in vivo study. Both in vitro and in vivo results show that NFL-LNC targeted brain NSC while they showed no affinity for spinal cord NSC [2]. While we are currently investigating the mechanisms involved in the preferential interactions of NFL-LNC with brain NSC, these data show that NFL-LNC is a promising therapeutic tool to selectively deliver bioactive molecules and to induce in situ NSC differentiation. References [1] Lépinoux-Chambaud, C., Barreau k., & Eyer J. The Neurofilament-Derived Peptide NFLTBS. 40-63 Targets Neural Stem Cells and Affects Their Properties. Stem Cells Transl Med (2016), 5: 901-913. [2] Carradori, D., Saulnier, P., Préat, V., des Rieux, A., & Eyer, J. (2016). NFL-lipid nanocapsules for brain neural stem cell targeting in vitro and in vivo. J. Control. Release, 238: 253262.. 28.

(29) Vectorization of Protein Antigens by Lipid Nanocarriers for Vaccine Formulation Bayon Emilie (1) (2) (3), Morlieras Jessica (1) (2), Courant Thomas (1) (2), Gonon Alexis (2) (3), Marche Patrice (2) (3), Navarro Fabrice (1) (2) 1 - CEA, LETI, MINATEC Campus (France), 2 - IAB, Université Grenoble Alpes (France), 3 - INSERM U1209 (France) The development of vaccines was one of the major health advances of the last century. However, new vaccine systems are still highly anticipated in the near future for facing today's health challenges. In this context, we have designed a novel biocompatible nanocarrier for inducing potent and specific immune responses. These particles are less than a hundred nanometers in size and made of a lipid core surrounded by a shell of pegylated surfactants1. Besides being biocompatible and highly stable, they can also be chemically grafted with protein antigens for delivery to the antigen presenting cells (APC). Different bioconjugation strategies have been explored to graft the model antigen ovalbumin (OVA) and P24 (a protein from HIV capsid) onto the particle shell. The safety of the resulting particles was determined by a cytotoxicity assay on dendritic cells (DC) and fibroblast cell lines. Next, we assessed in vitro the ability of primary DC to present the antigen to T lymphocytes (TL) after exposition to the antigen-bearing nanoparticles, by measuring secreted cytokines. Then, we evaluated in vivo the immunogenicity of these nanoformulations combined with immunostimulants. BALB/c mice were immunized with two intra-peritoneal injections at a 3-weeks interval. Following administration, we determined the antigen-specific antibodies in the mice sera and we collected the spleen cells in order to challenge them in vitro to the antigen (OVA or P24). A panel of secreted cytokines was analyzed in the supernatants, especially IFNγ, which expresses the strength of the induced cell-mediated immune response. The nanoparticles obtained after the grafting of OVA and P24 were very well tolerated by DC and fibroblasts. The vectorization of the antigen (OVA or P24) by the nanocarrier definitely improved its delivery to the immune cells, enabling DC to efficiently present the antigen to TL, which lead in vivo to the production of high titers of specific antibodies. In addition, the high levels of IFNγ secreted by TL from the mice give a strong indication for cell-mediated immune responses. Our results highlight the great benefit from the use of such nanostructured lipid carriers for delivering protein antigens to APC, by inducing higher immune responses. In perspectives, we plan to further explore the engineering of these nanoparticles, by incorporating additional functions such as the active targeting of dendritic cells, or by developing other dosage forms enabling oral delivery or nasal administration. 1 Delmas et al, Langmuir 27(5) 2011.. 29.

(30) RICK, an improved CPP for siRNA delivery via nanoparticle formation Vaissière Anaïs (1), Aldrian Gudrun (2), Konate Karidia (1), Lindberg Mattias, Jourdan Carole, Telmar Anthony, Seisel Quentin (1), Fernandez Frédéric, Viguier Véronique, Germain-Genevois Coralie, Couillaud Franck, Boisguerin Prisca (1), Deshaye Sébastien (1) 1 - Centre de Recherches de Biologie cellulaire de Montpellier (France), 2 - Sysdiag-Modélisation et Ingénierie des Systèmes Complexes Biologiques pour le Diagnostic (France) Cancer treatments are of a major interest for many years, all existing therapy are very aggressive and just efficient enough to delay tumor progression. We are mostly interested by brain cancer treatment and more specifically glioblastomas which represent the most aggressive brain tumor (Furnari et al, 2007). Indeed, once the diagnosis of glioblastoma established the prognosis is less than a year. The need of developing an efficient and non-aggressive therapeutic cancer treatment is crucial. On this purpose, we choose to develop a strategy based on proteins silencing which are crucial for cell cycle progression (Cycline and/or CDK). For many years now, siRNA molecules are widely investigated because they represent a therapeutic tool of major interest. Indeed, siRNA are able to bind and to degrade their mRNA complementary target leading to the silencing of the corresponding protein expression (Kathryn et al, 2009). The main challenge is the delivery of intact siRNA, which are easily degraded without protection, inside the cell. Cell penetrating peptides (CPPs) are of major interest as smart delivery tools for pharmacological molecules. Amphipatic CPPs have the ability to form nanoparticles by self-assembling around the oligonucleotide. One of the most advantage of CPPs is their ability to promote cellular uptake of therapeutic molecules like siRNA. Recently, we reported based on a comparative study of secondary amphipatic peptides that CADY-K could be used as efficient peptide-based nanoparticle for the delivery of siRNA in neuronal and melanoma cell lines (Konate et al, 2016). Based on these results, we also developed a carrier peptide, named RICK, composed of D-amino acids in the retro-inverso form. The advantage of this strategy is to obtain a vector which retains the delivery abilities and biological activity of its parental analogue (CADY-K) since its spatial orientation of the side chains remains unchanged. Furthermore, RICK should also not be recognized by proteases. We used different biophysical and biological techniques to study and to compare the nanoparticle assembly of RICK, CADY-K and D-cady-k. In details, we investigated RICK behavior and its ability to cross plasma membrane and to deliver intact siRNA inside the cell. This work is divided in two distinct part: the first part concerns the structural characterization of RICK peptide and its ability to form stable nanoparticles (in vitro). The second part is an investigation of the peptide ability to deliver siRNA inside the cell and induce biological knockdown activity (in cellulo).. 30.

(31) SESSION #3 – Nanomedicine & Imaging. 31.

(32) Molecular targeting using nanobodies Tony Lahoutte, MD, PhD Department of nuclear medicine at UZ Brussel. Prof Dr Tony Lahoutte is head of the department of nuclear medicine at UZ Brussel and head of the molecular imaging research unit at the Vrije Universiteit Brussel (VUB) in Belgium. He obtained his medical degree in 1998 and started his research activities in combination with a residency program in nuclear medicine. His current research is focused on the development and clinical translation of molecular imaging probes and targeted radionuclide therapies for the detection and treatment of cancer.. Positron emission tomography (PET) imaging of cancer is routinely performed using radiolabeled glucose for imaging cancer. This is a sensitive method to map all cancer lesions throughout the body at the time of diagnosis but also during follow up to measure response to treatment or relapse. The imaging technique also allows the detection of cancer related antigens when combined with specific targeting probes. The presence of certain antigens is a requirement for effective treatment with novel targeted anti-cancer treatments. Our research is focused on the development of radiolabeled cancer targeting probes to measure these cancer antigens non-invasively. We apply probes that are based on single domain antibody fragments derived from heavy chain only antibodies also called nanobodies. We developed nanobodies against the human epidermal growth factor receptor subtype 2 (HER2) for imaging breast cancer. Recently these imaging probes were evaluated in a phase I clinical trial. The presentation will give an overview of the development of the anti-HER2 nanobodies and their translation into a clinical application.. 32.

(33) Targeted Contrast Agents for Bioimaging and Nanomedicine Hak Soo Choi, Ph.D. Gordon Center for Medical Imaging, Department of Radiology, Massachusetts General Hospital and Harvard Medical School. Dr. Hak Soo Choi is Associate Professor of Radiology at Harvard Medical School, and faculty of Dana Farber/Harvard Cancer Center (DF-HCC) and Harvard Medical Faculty Physicians (HMFP) at BIDMC. Dr. Choi is a graduate of the Polymer-Nano Science Program at Chonbuk National University and earned his advanced degree in medicinal chemistry and nanomedicine from Japan Advanced Institute of Science and Technology (JAIST) in 2004. After experiencing gene and drug delivery at Pharmaceutical College of Hokkaido University, he extended his research into molecular cancer imaging and joined the Center for Molecular Imaging at BIDMC and MGH in Boston. Since 2008, his laboratory focuses on the development of novel contrast agents for tissue- and organ-specific targeting and diagnosis. Of particular interest is targeted fluorophores, which can be used for image-guided surgery by specifically visualizing target tissue with high optical properties and by avoiding nonspecific uptake in normal background tissues. Two fundamental and unsolved problems facing biophotonics and nanomedicine are nonspecific uptake of intravenously administered diagnostic and/or therapeutic agents by normal tissues and organs, and incomplete elimination of unbound targeted agents from the body. To solve these problems, we have synthesized a series of indocyanine near-infrared (NIR) fluorophores that varied systematically in net charge, conformational shape, hydrophilicity/lipophilicity, and charge distribution. Using 3D molecular modeling and optical fluorescence imaging, we have defined the relationship among the key independent variables that dictate biodistribution and tissue-specific targeting such as lung and sentinel lymph nodes (Nat Biotechnol. 2010), human prostate cancers (Nat Nanotechnol. 2010), and human melanomas (Nat Biotechnol. 2013). Recently, we have developed new pharmacophore design strategy “structure-inherent targeting,” where tissue- and/or organspecific targeting is engineered directly into the non-resonant structure of a NIR fluorophore, thus creating the most compact possible optical contrast agent for bioimaging and nanomedicine (Angew Chem. 2015, Nat Med. 2015). The biodistribution and targeting of these compounds vary with dependence on their unique physicochemical descriptors and cellular receptors, which permit 1) selective binding to the target tissue/organ, 2) visualization of the target specifically and selectively, and 3) provide curing options such as image-guided surgery or photo dynamic therapy. Our study solves two fundamental problems associated with fluorescence image-guided surgery and lays the foundation for additional targeted agents with optimal optical and in vivo performance.. KEY WORDS: Nanotechnology; Optical imaging; Diagnostic imaging; Tumor targeting; Near-infrared fluorophore; Biodistribution; Clearance; Image-guided surgery. 33.