nanoBioMed2015 Conference Bbook by Phantoms Foundation

nov. 18-20, barcelona (spain) www.nanobiomedconf.com

abstracts book organisers

On behalf of the Organizing Committee, we take great pleasure in welcoming you to Barcelona (Spain) for the NanoBio&Med2015 International Conference.

Foreword Organising

Committee

Sponsors

Exhibitors

Speakers

Posters

This event, after successful editions organized within ImagineNano in Bilbao 2011 & 2013, and in Barcelona in 2014, is going to present the most recent international developments in the field of Nanobiotechnology and Nanomedicine and will provide a platform for multidisciplinary communication, new cooperations and projects to participants from both science and industry. Emerging and future trends of the converging fields of Nanotechnology, Biotechnology and Medicine will be discussed among industry, academia, governmental and non-governmental institutions. NanoBio&Med2015 will be the perfect place to get a complete overview into the state of the art in those fields and also to learn about the research carried out and the latest results. The discussion in recent advances, difficulties and breakthroughs will be at his higher level. Considering the significant potential of graphene for applications in biomedicine and biotechnology, a specific session on Bio-graphene will be organised for the first time during nanoBio&Med2015. As last edition, an industrial forum will also be organized to promote constructive dialogue between business and public leaders and put specific emphasis on the technologies and applications in the nanoBioMed sector. We are indebted to the following Scientific Institutions and Government Agencies for their financial support: Institute for Bioengineering of Catalonia (IBEC), NanoSciences Grand Sud-Ouest (Câ&#x20AC;&#x2122;Nano GSO) and ICEX Spain Trade and Investment. We would also like to thank NanoMed Spain for their participation. In addition, thanks must be given to the staff of all the organising institutions whose hard work has helped planning this conference.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Antonio CORREIA President of the Phantoms Foundation (Spain) Dietmar PUM Deputy Head of the Biophysics Institute – BOKU (Austria) Josep SAMITIER Director of the Institute for Bioengineering of Catalonia – IBEC (Spain)

NanoBio&Med2015

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Speakers list: Alphabetical order Authors Ibane Abasolo (Vall d´Hebron Institute of Research (VHIR), Spain) Improved Pharmacokinetic Profile of Lipophilic Anti-Cancer Drugs Using Targeted Polyurethane-Polyurea Nanoparticles

Hugo Aguas (Universidade Nova de Lisboa, Portugal) Plasmonic Metal Nanoparticles: A tool for molecular and Bio detection

Lorenzo Albertazzi (Institute for Bioengineering of Catalonia (IBEC), Spain) Super Resolution Imaging of Nanomaterials: Looking at Nanomedicine with New Eyes

Adrian Bachtold (ICFO, Spain) Towards nanoMRI with mechanical resonators

Peter I Belobrov (Siberian Federal University, Institute of Biophysics SB RAS, Russia) Microfluidic bioassay based on bacterial luciferase and NADH: FMN-oxidoreductase

Ofra Benny (The Hebrew Univ of Jerusalem, Israel) Solidified Polymer Micelles for Drug Delivery in Cancer

Rafael Bernad (Bicosome S.L., Spain)

Session

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Oral

Parallel Session II

Keynote Plenary Session

Oral Parallel Session II

Keynote Plenary Session

Oral

Bicosome platform for Skin Drug delivery

Plenary Session

Miquel Bosch Pita (IBEC, Spain)

Invited

The molecular mechanisms of memory persistence: imaging how single synapses learn in real time

Remy Brossel (Cell Constraint & Cancer, France) Biomechanics: from in vitro to in vivo

David Caballero (IBEC, Spain) Bioengineered assays for cell migration studies

Rosalía Calleja (Mecwins S.A., Spain) Mecwins: Developing biomedicine applications using nanomechanical sensors

Alicia Calzado-Martín (Instituto de Microelectrónica de Madrid (CSIC), Spain) High Resolution Maps of Apparent Young’s Modulus of Breast Cancer Cells by Peak-Force Modulation Atomic Force Microscopy

Menglin Chen (Aarhus University, Denmark) Ultraporous interweaving electrospun nanofibers and their cellular response

Joël Chopineau (Institut Charles Gerhardt - CNRS/ENSCM/UM, France) Biomimetic bilayers designed for nanoparticles-membrane interactions and proteins transport studies

Etienne Dague (CNRS-LAAS, France) Biomedical applications of Atomic Force Microscopy

Ayelet David (Ben-Gurion University of the Negev, Israel) Biomedical polymers for treating primary and metastatic tumors

Alicia de Andrés (Instituto de Ciencia de Materiales de Madrid - CSIC, Spain) Graphene based platforms for Raman sensing

Yolanda Fernández (Vall d´Hebron Institute of Research (VHIR), Spain) In vitro and in vivo antimetastatic efficacy of a polymer-based paclitaxel conjugate for prostate cancer therapy

Xavier Fernàndez Busquets (University of Barcelona / IBEC, Spain) Nanomedicine: not for the developing world?"

Plenary Session

Oral Plenary Session

Keynote Plenary Session

Invited Plenary Session

Oral Parallel Session I

Oral Plenary Session

Keynote Plenary Session

Oral Plenary Session

Oral Parallel Session I

Keynote Plenary Session

Giancarlo Franzese (Universitat de Barcelona, Spain)

Oral

Contribution of Water to Pressure and Cold Denaturation of Proteins

Plenary Session

Petra Gener (CIBER-BBN, Spain) New fluorescent CSC models evidence that targeted nanomedicines improve treatment sensitivity of breast and colon cancer stem cells. Sensitivity of breast and colon cancer stem cells

David H. Gracias (The Johns Hopkins University, USA) 3D integrated bionanotechnology: From bionic devices to untethered surgical tools

Owen J Guy (Swansea University, UK) Graphene Biosensors for Point of Care Diagnostics

Oihane Ibarrola Moreno (Praxis Group, BioPraxis AIE, Spain) Nanomedicine, from PoC to reality: the importance of an industrial perspective and GMP scale up

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Oral Parallel Session I

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Authors Francesco Mallamace (Università degli Studi di Messina, Italy) The role of water in the protein activity: lysozyme from the glass to the unfolded state

Parallel Session II

Manuel Marqués (Universidad Autónoma de Madrid, Spain)

Invited

Analysis of the dielectric crossover in liquid water and the possible impact on biological and nanoscopic systems

Cécilia Ménard-Moyon (CNRS, France)

Plenary Session

Keynote

Biological Applications of carbon-based nanomaterials: from functionalisation to biodegradation

Plenary Session

Oscar Mendoza (Université de Bordeaux, France)

Keynote

Assembly of G-Quadruplex DNA nanostructures

Plenary Session

Karolina Mikulska-Rumińska (Institute of Physics Nicolaus Copernicus University , Poland) Nanomechanics of reelin autism related protein - AFM and SMD studies

Oral Parallel Session I

Angel Millán (Instituto de Ciencia de Materiales de Aragón, Spain)

Oral

Multifunctional Nanoplatform for hyperthermia. Heating and thermometry in a single nanoparticle

Parallel Session II

Boaz Mizrahi (Technion - Israel Inst. of Technology, Israel)

Keynote

Liquid Polymers for Drug Delivery and Regenerative Medicine

Plenary Session

Valery Pavlov (CIC BiomaGUNE, Spain) Biocatalytic modulation of semiconductor quantum dots: How to apply enzymatic growth and etching of CdS quantum dots to biosensing

Judit Perez Valero (Institut de Bioenginyeria de Catalunya, Spain)

Oral Plenary Session

Oral

Molecular Wires for the Improvement of DNA Electrochemical Sensors

Plenary Session

Danny Porath (The Hebrew University of Jerusalem, Israel)

Keynote

The Quest for Charge Transport in single Adsorbed Long DNA-Based Molecules

Plenary Session

Elisabet Prats-Alfonso (IMB-CNM (CSIC), Spain)

Keynote

Graphene in neural interface systems: perspectives and applications

Plenary Session

Anna Roig (Institut de Ciència de Materials de Barcelona (ICMAB-CSIC), Spain)

Oral

Bio-identity of Albumin-Iron Oxide Nanoparticles

Plenary Session

José Manuel Rojas (Centro Nacional de Biotecnología, CNB-CSIC, Spain)

Oral

Superparamagnetic iron oxide nanoparticle uptake alters M2 macrophage phenotype

Plenary Session

Samuel Sánchez (IBEC, Spain)

Keynote

Hybrid micro and nanoBots as future active drug carriers

Plenary Session

Gregory F. Schneider (Universiteit Leiden, The Netherlands)

Keynote

Single molecule graphene biosensors: chemistry matters

Plenary Session

Avi Schroeder (Technion - Israel Inst. of Technology, Israel)

Keynote

Personalized Cancer Nanomedicine

Plenary Session

Simó Schwartz (Vall d'Hebron Hospital - CIBBIM, Spain)

Keynote

Targeting cancer stem cells to increase sensitivity to chemotherapy

Plenary Session

Oded Shoseyov (The Hebrew University of Jerusalem, Israel)

Keynote

Nano crystalline cellulose-protein composites: Super performing biomaterials for tissue engineering and regenerative medicine

Plenary Session

Juliane Simmchen (MPI Intelligent Systems, Germany)

Oral

Light driven Micromotors for waste water remediation

Plenary Session

Lorena Simón Gracia (University of Tartu, Institute of Biomedicine, Estonia)

Oral

Affinity-targeted tumor penetrating polymersomes

Parallel Session I

Morgan M. Stanton (Max Planck Institute for Intelligent Systems, Germany)

Oral

Bio-hybrid Janus Motors Driven by Escherichia coli

Parallel Session I

Javier Tamayo de Miguel (IMM-CSIC, Spain)

Keynote

Nanomechanical and Optomechanical Systems for Cancer Research

Plenary Session

María Teresa Valero Griñán (Pfizer - Universidad de Granada - Junta de Andalucía Centre for Genomics and Oncological Research (GENYO), Spain) A nanotechnology approach to evaluate drug promiscuity in cancer

Oral

Graphene-based self-propelled micromotors as decontaminating micromachines of environmental pollutants

Sinéad Winters (Trinity College Dublin, Ireland)

Oral Parallel Session II

Diana Vilela Garcia (Max Planck Institute for Intelligent Systems, Germany)

Plenary Session

Keynote

Non-covalent functionalisation of graphene: Optimising molecular packing density and stability

Keynote Oral

Cork filter with silver nanoparticles for Lab-Scale water disinfection

Graphene for biomedical applications

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Plenary Session

Rosanna Margalef (UNESCO Chair on Sustainability - UPC, Spain)

Amaia Zurutuza (Graphenea, Spain)

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NanoBio&Med2015

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Plasmonic Metal Nanoparticles: A tool for molecular and Bio detection Hugo Aguas Universidade Nova de Lisboa, Portugal hma@fct.unl.pt Optical detection methods that take profit of the unique properties of metallic nanoparticles have become increasingly important in many fields of application, from biosensors to security and environmental monitoring, due their ease of application and comparative low cost. The interaction of light with metallic nanoparticles induces the formation of dipolar localized surface plasmon resonances (LSPR) – the resonant excitation of the delocalized free electrons within the nanoparticles. This LSPR can result in a strong absorption band in the visible frequency range, and strong electromagnetic fields at the particle surface that polarise the local volume around the nanoparticle inducing light scattering. The type, size and shape of the metallic nanoparticles determine the wavelength of the optical absorption and the scattering cross-section influencing their applicability. By fine tuning

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these parameters, Au nanoparticles have been successfully implemented for the detection of DNA, in a microfluidic setup, enabling significant discrimination between positive and negative assays using a target DNA concentration of 5 ng/µL, below the limit of detection of the conventionally used microplate reader (i.e., 15 ng/µL) with 10 times lower solution volume (i.e., 3µL). Also, Ag nanoparticles have been successfully applied in the fabrication of low cost plasmonic surface platforms for Surface Enhanced Raman Spectroscopy using physical self-assembled and chemical synthesis methods in rigid (glass and silicon) and flexible (paper) substrates. With these platforms we were able to detect rhodamine 6G (R6G) in the sub ppb range, opening the applicability of these substrates to the detection of chemical substances such as food toxins, pesticides and explosives.

november 18-20, 2015 - Barcelona (Spain)

Super Resolution Imaging of Nanomaterials: Looking at Nanomedicine with New Eyes Lorenzo Albertazzi, Nanoscopy for Nanomedicine Group Institute for Bioengineering of Catalonia (IBEC) Barcelona, Spain lalbertazzi@ibecbarcelona.eu The use of nanocarriers for intracellular delivery of therapeutic moieties is a great challenge for synthetic chemistry and nanotechnology. In this framework, supramolecular materials such as micells, liposomes self-assembled nanoparticles and nanofibers plays a pivotal role. A crucial factor limiting the design of effective materials is the lack of understanding about material-cell interactions that hampers the rational design of nanosized carriers. This is particularly relevant for supramolecular materials as their complex structure poses several unanswered questions. Here we discuss the use of super resolution microscopy to image materials in vitro and in mammalian cells. This novel technique, allowing to obtain a resolution down to 20nm, had a dramatic impact in the field of cell biology, however its use in the field of chemistry and nanotehcnology is poorly explored. Super resolution microscopy offers nanometric resolution and multicolor ability, therefore it is an ideal tool to study nano-sized

supramolecular assemblies of components in vitro and in cells.

multiple

We employed Stochastic Optical Reconstruction Microscopy (STORM) to image biomaterials in vitro, with special emphasis on supramolecular polymers and nanoparticles, unveiling novel information on materials structure and dynamics, a key issue of supramolecular materials. Moreover we propose a methodology to image nano-sized materials in cells, tracking them during their membrane targeting, cell uptake and intracellular targeting. We show how 2color STORM can be used to perform nanometric-accurate colocalization unveiling at the molecular level materials-cell interactions. This allow to look at nanomaterials in action with new eyes and use the information obtained for the â&#x20AC;&#x153;STORM-guidedâ&#x20AC;? design of novel nanomaterials for drug delivery and other targeted therapies.

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november 18-20, 2015 - Barcelona (Spain)

Towards nanoMRI with mechanical resonators Adrian Bachtold ICFO – The Institute of Photonic Sciences, Castelldefels (Barcelona), Spain adrian.bachtold@icfo.es Carbon nanotubes and graphene offer unique scientific and technological opportunities as nanoelectromechanical systems (NEMS). Namely, they allow the fabrication of mechanical resonators that can be operated as exceptional sensors of mass and force. The mass resolution can be as low as 1.7 yg, which is about the mass of 1 proton [1]. The force sensitivity can reach ~1 zN/Hz1/2 [2,3]. Such a force sensitivity may enable the detection of single nuclear spins by placing the resonator in a strong gradient of magnetic field. Here, I will review our efforts towards the detection of single nuclear spins and the realization of nano magnetic resonance imaging (nanoMRI).

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References [1] J. Chaste, A. Eichler, J. Moser, G. Ceballos, R. Rurali, A. Bachtold, Nature Nanotechnology 7, 301 (2012). [2] J. Moser, J. Güttinger, A. Eichler, M. J. Esplandiu, D. E. Liu, M. I. Dykman, A. Bachtold, Nature Nanotechnology 8, 493 (2013). [3] J. Moser, A. Eichler, J. Güttinger, M. I. Dykman, A. Bachtold, Nature Nanotechnology 9, 1007 (2014).

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Solidified Polymer Micelles for Drug Delivery in Cancer Ofra Benny The Hebrew Univ of Jerusalem, Israel OfraB@ekmd.huji.ac.il Nanomedicine is an emerging field in cancer therapy that has the potential of revolutionizing the way drugs are introduced to patients today. Many drugs have critical limitations such as low solubility, stability, and specificity â&#x20AC;&#x201C; leading to inefficient treatment and to adverse side effects. Among the different types of drug-delivery systems, polymer micelles represent an appealing technology for delivering drugs to tumors because of their relatively simple formulation and their small size, enabling efficient tumor extravasation from leaky tumor blood vessels. We found that self-assembled di-block polymers can be used successfully to deliver small molecule drugs by encapsulation or by chemical conjugation of the drug, and that

these nano-micelles can be further stabilized by a secondary solidification step. The formation of stable solidified nano-micelles enables efficient cellular internalization, and improve drugsâ&#x20AC;&#x2122; bioavailability half-time, enhances blood circulation time, increases tumor uptake, and reduces side effects as demonstrated in-vivo. Moreover, our studies also revealed that the endocytosis of stabilized particles occur via clathrin pathway, unlike the reported mechanism for non-solidified polymer micelles. Because of the high stability of this particles our data suggest that the nanoparticles can be transported by transcytosis and thereby they can also be efficient in tumors that are not vascularize.

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november 18-20, 2015 - Barcelona (Spain)

The molecular mechanisms of memory persistence: imaging how single synapses learn in real time Miquel Bosch Pita IBEC, Spain mbosch@ibecbarcelona.eu Memories are stored in our brain through the ability of synaptic connections to modify their structure and function in a long-lasting way. However, nobody has ever observed how these changes occur in a single synapse in real time. I will explain how we used a new combination of optical technologies to reveal the molecular remodeling that takes place inside a synapse during the creation of a memory. We used twophoton microscopy to stimulate individual

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synapses and to visualize protein trafficking in real time. We identified a unique protein that is rapidly and persistently captured in potentiated synapses, forming a new macromolecule that could serve as a memory tag. We developed a novel photo-marking technique that allowed us to localize the same synapses under both twophoton and electron microscopies. This way we observed how synaptic structures evolve asynchronously in different temporal phases during synaptic potentiation.

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Bioengineered assays for cell migration studies David Caballero1,2,3 1 Nanobioengineering group â&#x20AC;&#x201C; IBEC (Barcelona, Spain) 2 CIBER-BBN (Zaragoza, Spain) 3 Department of Electronics, University of Barcelona (Barcelona, Spain) dcaballero@ibecbarcelona.eu Many physiological and pathological processes involve directed cell motion. Directed cell migration is usually thought to depend on the presence of long-range gradients of either chemo-attractants or physical properties, such as stiffness or adhesion [1]. However, in vivo, chemical or mechanical gradients have not systematically been observed. In this talk, I will present recent in vitro experiments which show that other types of spatial guiding cues can bias cell motility [2]. Introducing local geometrical or mechanical anisotropy in the cell environment, such as adhesive [3, 4] or topographical microratchets [5, 6], show that local and periodic external cues can direct cell motion. I will show the importance of protrusion fluctuations in setting the direction of cell motion, and how their spatiotemporal distribution and dynamics determine the fluctuations and direction of cell motion. Under certain circumstances, mechanical constraints and chemical gradients can both contribute to the establishment of cell direction. We found that the nucleus dictates the direction of cell movement through mechanical guidance by its environment, and demonstrate that this direction can be tuned by combining the topographical ratchet with a biochemical gradient of adhesion. Interestingly, we found competition and cooperation between the two external cues. Together with modeling, these experiments suggest that cell motility is a stochastic phenomenon which can be biased by

various types of local cues, leading to directional migration. Finally, I will present our most recent results on cell migration using in-vitro native-like environments. Our aim is to study how cell migration is determined by both mechanical and biochemical factors during physiological and pathological processes.

References [1] D. Caballero and J. G. Goetz, Cell Adhesion & Migration 9, 325 (2015). [2] D. Caballero, J. Comelles, M. Piel, R. Voituriez, and D. Riveline, Trends in Cell Biology In Press (2015). [3] D. Caballero, R. Voituriez, and D. Riveline, Biophys J 107, 34 (2014). [4] D. Caballero, R. Voituriez, and D. Riveline, Cell Adhesion & Migration 9, 327 (2015). [5] J. Comelles, D. Caballero, R. Voituriez, V. HortigĂźela, V. Wollrab, A. Godeau, J. Samitier, E. Martinez, and D. Riveline, Biophys J 107, 1513 (2014). [6] R. Feynman, R. Leighton, and M. Sands, The Feynman Lectures on Physics Reading, MA, 1, 41-I, 1963), Reading, MA.

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Mecwins: developing biomedicine applications using nanomechanical sensors RosalĂa Calleja Mecwins S.A., Plaza de la Encina 10-11, Tres Cantos (Madrid), Spain rcalleja@mecwins.com Mecwins was founded in 2008 by Dr. Javier Tamayo and Dr. Montserrat Calleja, Bionanomechanics group leaders from the Institute of Microelectronics of CSIC (IMM - CSIC) [1]. Since then, we have been developing cutting edge technology for nanomechanical sensing. The technology, based on detecting variations in the deflection and resonance frequency of nanomechanical sensors, was the groundwork for the technical improvements that led us to our new ultrasensitive detection device, SCALA. SCALA (SCAnning Laser Analyzer) [2] is a commercial platform with high potential for the analysis of biomolecule interactions in human whole blood samples for biosensing applications. The technology uses cantilevers as trasducers for the detection of biomolecules. Cantilever arrays have been extensively explored as high-sensitive nanomechanical biosensors [3]. The molecular recognition on the surface of a biofunctionalized cantilever results in a nanomechanical response, that produces cantilever bending of a few nanometers (static mode) or changes in cantilever resonance frequency (dynamic mode) where the added mass due to selective molecular recognition decreases resonance frequency value. SCALA has been used for the successful detection of different protein biomarkers in clinical diagnosis, focusing our preliminary experiments on oncology, cardiac and infectious diseases biomarkers. SCALA combines mechanical detection technique (microcantilevers resonance frequency analysis)

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with a new optical detection technique that increases current levels of sensitivity in clinical practice a million times [4] in comparison with techniques currently used in hospitals and central laboratories and without increasing current cost per sample. The adoption of ultrasensitive detection equipment will enable screening for early detection of a wide range of diseases with established diagnostic biomarkers from a droplet of blood. The adoption of ultrasensitive detection equipment will enable screening for early detection of a wide range of diseases with established diagnostic biomarkers from a droplet of blood. Moreover, we have developed our technology to be portable, in order to be introduced in the market as a POC device (Figure 1) that allows the implementation of fast and efficient clinical procedures for diagnosis, monitoring and prognosis that will cut down costs in healthcare expenditure, insurance or laboratory work.

References [1] Spanish Research Council: http://www.immcnm.csic.es/bionano/es [2] http://mecwins.com/ [3] Arlett et al. Nat Nanotechnol 6(4),203-15 (2011) [4] Kosaka et al. Nat Nanotechnol. 9(12),1047-53 (2014)

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Figures

Figure 1. First prototype or version of the portable SCALA Platform

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Biomimetic bilayers designed for nanoparticles-membrane interactions and proteins transport studies JoĂŤl Chopineau Institut Charles Gerhardt - CNRS/ENSCM/UM, France joel.chopineau@unimes.fr Biological membranes carry out the essential function of a natural barrier that separates the cell from the outside environment. To study, in controlled conditions, the biological events occurring at the cell membrane interface, we recently develop biomimetic devices based on supported biomembranes. A solid supported biomimetic membrane was developed to study in vitro the translocation process of a bacterial toxin, the adenylate cyclase (CyaA) from Bordetella pertussis. The membrane was assembled over a calmodulin (CaM) layer and exhibits the fundamental characteristics of a biological membrane separating two cis and trans compartments. The activation of the catalytic activity of CyaA by the tethered CaM was used as a probe of its translocation across the bilayer. This work constitutes the first in vitro demonstration of protein translocation across a tethered lipid bilayer. Biomimetic assemblies are also important tools for the study of the interaction of mesoporous silica nanoparticles (MSN) with membranes. We have developed synthetic routes to achieve the production of gold loaded radial mesoporous silica nanoparticles (Au-MsNP) and MSN@Fe3O4 materials incorporating magnetic and fluorescent properties as multifunctional platforms. The size of particles was checked by dynamic light scattering while zeta potential measurements reflect their surface charge. The particles morphology was characterized by transmission and scanning electron

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microscopies. Their textural properties, specific surface area and pore size, were determined from N2 adsorption. The gold metallic nanoparticles embedded in the pore channels of Au-MsNP are responsible for a plasmonic activity. The coating with phospholipid bilayers of Au-MsNP particles provided a biofunctional device with plasmonic properties relevant for biosensing. For this purpose different model systems have been investigated, direct adsorption of bovine serum albumin or molecular recognition events between a biotin receptor, integrated in the supported lipid bilayer, and avidin molecules. The obtained results demonstrate the plasmonic sensitivity of the bare Au-MsNP particles or coated lipid bilayer Au-MsNP devices. We investigate MSN@Fe3O4 cell membrane interactions depending on nanoparticle surface coverage, to study MSN@Fe3O4 behavior in biological fluids. The dispersibility of MSN@Fe3O4 materials (pristine, lipid or polyethylene glycol coated) was largely dependent on medium composition and nanoparticle coating. A biomimetic membrane model was used to investigate MSN@Fe3O4 â&#x20AC;&#x201C; cell membrane interactions. The presence of a 80mV transmembrane potential applied in trans side seems to increase MSN@Fe3O4 interaction with the membrane. Dispersion media, MSN@Fe3O4 coating and transmembrane potential appeared as major factors influencing MSN cell membrane interactions.

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References [1] Rossi, C., Doumiati, S., C. Lazzarelli, D., Davi, M., Meddar, F., Ladant, D. and Chopineau, J. (2011) "A tethered bilayer assembled on top of immobilized calmodulin to mimic cellular compartmentalization" PLoS ONE 6(4): e19101. [2] Veneziano, R., Derrien, G., Tan, S., Brisson, A., Devoisselle, JM., Chopineau, J., and Charnay, C. (2012) "One step synthesis of gold loaded radial mesoporous silica nanospheres and supported lipid bilayers functionalization: towards bio-multifunctional sensors" Small. 8, 3674-3682. [3] Allouche, M., Pertuiset, C., Robert, J.L., Martel, C., Veneziano, R., Henry, C., Dein, O.S., Saint, N., Brenner, C., Chopineau, J. (2012) "ANTVDAC1 interaction is direct and depends on ANT isoform conformation in vitro" Biochem Biophys Res Commun. 429, 12-17.

[4] Subrini O, Sotomayor-PĂŠrez AC, Hessel A, Spiaczka-Karst J, Selwa E, Sapay N, Veneziano R, Pansieri J, Chopineau J, Ladant D, Chenal A. (2013) "Characterization of a membraneactive peptide from the Bordetella pertussis CyaA toxin"J. Biol. Chem. 288, 32585-98. [5] Veneziano R, Rossi C, Chenal A, Devoisselle JM, Ladant D, Chopineau J. (2013) "Bordetella pertussis adenylate cyclase toxin translocation across a tethered lipid bilayer" Proc Natl Acad Sci U S A. 110(51), 20473-8. [6] Damiati S1, Zayni S, Schrems A, Kiene E, Sleytr UB, Chopineau J, Schuster B, Sinner EK. (2015) "Inspired and stabilized by nature: ribosomal synthesis of the human voltage gated ion channel (VDAC) into 2D-proteintethered lipid interfaces". Biomater Sci.3 (10):1406-13.

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Biomedical Applications of Atomic Force Microscopy Etienne Dague1, 2, 3 1 CNRS, LAAS, 7 avenue du colonel Roche, F-31400 Toulouse, France 2 CNRS, ITAV-USR 3505, F-31100 Toulouse, France 3 Univ de Toulouse, LAAS, ITAV, F-31000 Toulouse, France edague@laas.fr Atomic Force Microscopy is now widely used to explore biological questions[1]. In this presentation, we will focus on 3 applications of AFM in life science and medicine. The first one is related to yeast cells. C. albicans is a human opportunistic pathogen responsible for benign to dreadful infections. Using AFM, we have explored its’ adhesive properties and discovered the formation of adhesive nanodomains made of aggregated proteins [2] (see left panel of figure 1). We also tested the effect of caspofungin (a last chance drug against C. albicans) on the cells nanomechanical and adhesive properties [3]. In the second application, Pseudomonas aeruginosa cells were treated with 2 major antibiotics: ticarcillin (figure 1 center panel) and tobramycin. We have demonstrated that treated cells present an altered shape, roughness and elasticity [4]. Moreover, we took advantage of force spectroscopy to study the cell wall of a multi resistant strain, and we unravelled the mechanism of action of an innovative molecule: CX1, efficient against this multi resistant strain [5]. Finally, I will deal with exciting results obtained on living cardiomyocytes (CM). The cells were extracted from mice heart, adhered to laminin coated petri dish and kept alive during the AFM experiments using the perfusing cell from Brucker (figure 1 right panel). Combining AFM and electron microscopy, we have demonstrated a dramatic morphological

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modification of the CM after heart failure that is correlated with the modification of the nanomechanical properties of the cells [6]. We have also studied the role of the protein ephrin B1 in CM elasticity and shape [7].

References [1] Pillet F., Chopinet L., Formosa C., Dague E., 2014 Atomic Force Microscopy and pharmacology; from microbiology to cancerology, Biochimica and Biopysica Acta General Subjects 1840 1028-1050 [2] Formosa C., Schiavone M., Boisramé A., Lavie Richard M., Duval R.E., Dague E., Revision submitted. Multiparametric imaging of adhesive nanodomains at the surface of Candida albicans by Atomic Force Microscopy Nanomedicine NBM [3] Formosa C., Schiavone M., Martin-Yken H., François J.M., Duval R. E., Dague E., 2013 Nanoscale effects of caspofungin against two yeast species; Saccharomyces cerevisiae and Candida albicans, Antimicrobial Agents and Chemotherapy 57. 3498-3506 [4] Formosa C., Grare M., Duval R.E., Dague E. 2012. Nanoscale effects of antibiotics on P. aeruginosa 8. 14-16 Nanomedecine NBM [5] Formosa C., Grare M., Coutable A., Jauvert E., Regnouf de Vains J.-B., Mourer M., Duval R.E., Dague E. 2012. Nanoscale analysis of the effects of antibiotics and Cx1 on Pseudomonas aeruginosa multidrugresistant, Scientific Reports (Nature Publishing Group) 2. 575

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[6] Dague E., Genet G., Lachaize V., Fauconnier J., Guilbeau-Frugier C., Payré B., Chopinet L., Alsteens D., Severac C., Thireau J., Heymes C., Honton B., Lacampagne A., Pathak A., Sénard J.M., Galés C. Revision Submitted AFM of living cardiomyocytes surface unveiled unexpected mitochondrial shifts in heart failure, In Press Journal of molecular and cellular cardiology

[7] Genet G., Guilbeau-Frugier C., Honton B., Dague E., Schneider M.D., Coatrieux C., Calise D., Cardin C., Nieto C., Payré B., Dubroca C., Marck P., Heymes C., Dubrac A., Avranitis C., Despas F., Altié M-F.; Seguelas M-H., Delisle MB., Davy A., Senard J-M., Pathak A., Gales C. 2012. Ephrin-B1 is a novel component of the lateral membrane of the cardiomyocyte and is essential for the stability of cardiac tissue architecture cohesion Circulation Research 110. 688-700

Figures

Figure 1. From left to right: AFM adhesion image obtained on Candida albicans, AFM deflection image of Pseudomonas aeruginosa treated by ticarcillin, AFM 3D image of a living cardiomyocyte.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Biomedical polymers for treating primary and metastatic tumors Ayelet David Department of Clinical Biochemistry and Pharmacology, Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, Israel ayeletda@bgu.ac.il In cancer therapy, several attempts have been made to target chemotherapeutic drugs and nanomedicines directly to different cell types of the tumor microenvironment. In this lecture I will describe the design of new polymer-drug conjugates that can actively target endothelial cells, tumor associated macrophages, and cancer cells of diverse tumor origin. The endothelial cell targeted polymer-drug conjugates significantly inhibit primary tumor growth, prevented the development of pulmonary metastases, and could further control the establishment of metastasis. Our research highlights the endothelial cells targeted systems as effective nanomedicines to treat primary and metastatic tumors.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Nanomedicine: not for the developing world? 1

Xavier FernĂ ndez-Busquets Nanomalaria Joint Unit, Institute for Bioengineering of Catalonia (IBEC), Barcelona, Spain 2 Nanomalaria Joint Unit, Barcelona Institute for Global Health (ISGlobal), Spain 3 Nanoscience and Nanotechnology Institute of the University of Barcelona (IN2UB), Spain xfernandez@ibecbarcelona.eu

Because malaria pathophysiology is so complex and the disease is so widespread, it is generally accepted that to achieve eradication a combination of tools targeting the parasite and/or mosquito will be needed [1]. These include the improvement of existing approaches and the development of new ones [2], with drug therapy remaining the mainstay of treatment and prevention to target the parasite reservoir [3], and nanotechnology being able to provide innovative useful strategies [4]. Encapsulation of drugs in targeted nanovectors is a rapidly growing area with a clear applicability to infectious disease treatment [5], and pharmaceutical nanotechnology has been identified as a potentially essential tool in the future fight against malaria [6, 7]. The application of nanotechnology to malaria has been traditionally neglected; the reasons for this gap in nanomedical research are surely varied, but among them are the lack of interest of a profit-seeking industry and the timid support of public administrations to small groups working off the main path of developed world diseases. However, the implementation of novel delivery approaches is less expensive than finding new antimalarial drugs and may optimize the rate of release of current and novel compounds [8]. An essential aspect for the successful development of antimalarial nanomedicines resides on the choice of encapsulating and targeting elements, which need to be tailored and optimized for their biocompatibility, cell specificity, binding affinity, ease of modification and conjugation to the drugs, production cost, scalability, amenability to oral administration formulation, and stability in mass production. The three

elements that constitute a targeted therapeutic nanovector (nanocapsule, targeting molecule and the drug itself) can be exchanged, as if they were LEGO parts, to obtain new structures better suited to each particular situation. Targeting agents for future malaria medicines can consist of cost-efficient heparin-like molecules, or may even be substituted altogether by self-targeting polymeric structures. These, after delivering their active cargo to target cells, can have an up to recently unsuspected second life as vaccination adjuvants. Drug delivery does not necessarily have to be to parasitized red blood cells, but can be engineered to prefill the more easily druggable uninfected erythrocyte. Finally, the design of antimalarial nanomedicines directly administered to mosquitoes and targeted at malaria parasite stages exclusive to the insect might spectacularly reduce costs and bench-totreatment time because in this way clinical trials could be significantly simplified.

Acknowledgements This research was supported by grants BIO201452872-R (Ministerio de EconomĂa y Competitividad, Spain), which included FEDER funds, 2013-0584 (Fondazione Cariplo, Italy), and 2014-SGR-938 (Generalitat de Catalunya, Spain).

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november 18-20, 2015 - Barcelona (Spain)

References [1] R. G. Feachem, A. A. Phillips, G. A. Targett, R. W. Snow, Lancet 376, 1517 (2010). [2] P. L. Alonso, Int. Microbiol. 9, 83 (2006). [3] J. P. Daily, J. Clin. Pharmacol. 46, 1487 (2006). [4] R. Duncan, R. Gaspar, Mol. Pharmaceutics 8, 2101 (2011). [5] P. Urbán et al., Curr. Drug Targets 13, 1158 (2012).

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[6] N. Kuntworbe, N. Martini, J. Shaw, R. AlKassas, Drug Dev. Res. 73, 167 (2012). [7] P. Urbán, X. Fernàndez-Busquets, Curr. Med. Chem. 21, 605 (2014). [8] P. Murambiwa, B. Masola, T. Govender, S. Mukaratirwa, et al., Acta Tropica 118, 71 (2011).

november 18-20, 2015 - Barcelona (Spain)

3D integrated bionanotechnology: From bionic devices to untethered surgical tools David H. Gracias Department of Chemical and Biomolecular Engineering, Johns Hopkins University, USA dgracias@jhu.edu An important challenge of nanotechnology is to integrate physical processes and human engineered devices with biological cells and organisms. Human engineered devices can facilitate communication, logic and memory processing while biological devices are selfpropelled, adaptive and environmentally responsive. Hence, combining these attractive features and facilitating mechanisms to bridge the divide between the two worlds could significantly augment the capabilities of human engineering and also provide new strategies for medical diagnostics and therapeutics. However, bridging the divide between the engineered and biological worlds is very challenging. Firstly, there is an inherent mismatch in the processes used to create electronic devices and biological systems. Most processes such as thin film deposition in electronic fabrication are vacuum based processes and many of the devices are incompatible with aqueous media. Further, physical devices are often mechanically rigid while biological devices are often soft and squishy. Finally, many functional micro and nanoscale devices are produced by inherently planar lithographic or serial approaches while biological cells and organisms are truly 3D and self-organize in large numbers. In an attempt to tackle these challenges, I will highlight by way of examples, two strategies that merge concepts, processes and/or functionalities from the engineering and biological worlds at small size scales.

Firstly, I will discuss how functionalized nanoparticles can be attached to the surfaces of microorganisms such as bacteria to alter the behavior of these microorganisms. Importantly, nanoparticles can be grown or self-assembled in large numbers so that bacterial hybrids can be formed using solution based antibodylinking schemes [1]. I will further illustrate how such bionic nano-particle bacteria hybrids can be used to ferry cargo and transmit signals between the external physical and biological worlds. In addition, I will also discuss the creation of bionic organs via 3D printing of nanoparticles and cells to form ear-like devices with embedded antennas [2]. Secondly, I will discuss the creation of large numbers of sub-mm and even single cell sized devices for medicine. These include biodegradable surgical microdevices and mechanical traps for single cell capture and analysis [3]. Additionally, I will also discuss the design and operation of dust-sized biopsy forceps to perform the first ever in-vivo biopsies [4] illustrating that the development and safe deployment of mass-producible untethered surgical tools could make surgery more effective and less invasive.

References [1] R. Fernandes et al, Small 7, 5, 588-592 (2011). [2] M. S. Mannoor, et al, Nano Letters, 13, 6, 2634â&#x20AC;&#x201C; 2639 (2013). [3] K. Malachowski et al, Nano Letters 14, 7, 41644170 (2014). [4] E. Gultepe, et al, Advanced Materials 25, 4, 514-519 (2013).

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november 18-20, 2015 - Barcelona (Spain)

Graphene Biosensors for Point of Care Diagnostics Owen J Guy, Z. Tehrani, R. Forsyth, R. Bigham, G.Burwell, K-A. D. Walker Centre for Nanohealth, College of Engineering, Swansea University, Singleton Park, Swansea, UK o.j.guy@swanse.ac.uk Graphene devices and sensors promise to be a disruptive technology in next generation electronics and healthcare diagnostics - due to grapheneâ&#x20AC;&#x2122;s exceptional electronic properties. This presentation outlines the development of novel graphene sensor technology for healthcare diagnostics based on a chemically functionalised graphene microchannel. Graphene biosensors have been fabricated using graphene grown on both the silicon and carbon faces of on-axis 4HSiC substrates [1-3], screen printed graphene sensors [4] and on CVD graphene. The presentation will review chemical functionalisation methods and sensing applications of graphene. Following a brief review of different sensing techniques, the presentation will focus on electrochemical and CHEMFET biosensors. The suitability of different types of graphene for sensing applications will be discussed. A range of different chemistries will be presented including methods used for exfoliated and solution based graphene as well as CVD and epitaxial graphene. Direct and indirect (using a modification of an adsorbed layer or polymer film on top of the graphene) functionalisation techniques including diazotisation, aminosilane chemistry and plasma functionalisation methods will be reviewed. An emphasis on production friendly techniques suitable for fabricating devices on a large scale (full wafers or large area) will be presented. Single layer, bilayer and few-layer graphene have all been functionalised using an aryl diazonium coupling reaction to achieve aniline terminated graphene. This chemistry has been used to modify

NanoBio&Med2015

graphene in order to attach "bioreceptor" molecules, capable of specific and selective detection of target biomarkers will be reviewed. The aniline molecule has been used to graft antibodies - targeted against the cancer risk biomarker 8-hydroxydeoxyguanosine (8-OHdG) onto the graphene surface. Antibody attachment to graphene has been verified using Fluoresence Microscopy to detect quantum-dot labelled antibodies bound to the graphene surface. Changes in the current-voltage characteristics of the graphene sensors have been used to detect 8OHdG at nanoMolar concentrations. The effect of functionalisation on electrical transport properties including carrier type and mobility has been demonstrated using direct current and electrochemical platforms. Issues including characterisation of sensor operation, reproducibility and reliability will all be addressed. There are several advantages of graphene sensors over alternative sensor platforms such as carbon nanotubes (CNTs) or silicon nanowires (SiNWs) [5]. The main benefits of graphene for sensing applications will be highlighted in a comparison with other materials. Finally, the issues of integrating graphene sensors into packaging and microfluidics will be considered and the range of potential applications from DNA sensors to immunoassays to detection of food toxins will be reviewed. Results from antibody based â&#x20AC;&#x153;immunesensorsâ&#x20AC;? and nucleic acid detection devices will be reported.

november 18-20, 2015 - Barcelona (Spain)

References [1] "Generic Epitaxial Graphene Biosensors for Ultrasensitive Detection of Cancer Risk Biomarker" by Tehrani, Zari; Burwell, Gergory; Mohd Azmi, Mohd Azraie; Castaing, Ambroise; Rickman, Robbert; Almarashi, Jamal; Dunstan, Peter; Miran Beigi, Ali Akbar; Doak, Shareen; and Guy, Owen, 2D Materials. 1 (2014) 025004 [2] “Effects of a modular two-step ozone-water and annealing process on silicon carbide graphene”, Webb, Matthew J. and Polley, Craig and Dirscherl, Kai and Burwell, Gregory and Palmgren, Pål and Niu, Yuran and Lundstedt, Anna and Zakharov, Alexei A. and Guy, Owen J. and Balasubramanian, Thiagarajan and Yakimova, Rositsa and Grennberg, Helena, Applied Physics Letters, 105 (2014) 081602

[3] Teixeira, S. Burwell, G. Castaing, A. Gonzalez, D. Conlan, R. & Guy, O. (2014). Epitaxial graphene immunosensor for human chorionic gonadotropin. Sensors and Actuators B: Chemical, 190 (2014) 723. [4] “Label-free human chorionic gonadotropin detection at picogram levels using oriented antibodies bound to graphene screen printed electrodes”, Teixeira, S. Conlan, S. Guy, O. & Sales, M.. Journal of Materials Chemistry B, 2014, 2, 1753-1754 [5] “Highly sensitive covalently functionalised integrated silicon nanowire biosensor devices for detection of cancer risk biomarker”, M.A. Mohd Azmi, Z. Tehrani, R.P. Lewis, K.-A.D. Walker, D.R. Jones, D.R. Daniels, S.H. Doak, and O.J. Guy. Elsevier Biosensors and Bioelectronics, 52 (2014) 216.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

The role of water in the protein activity: lysozyme from the glass to the unfolded state (Nuclear Magnetic Resonance and Neutron Spectroscopy studies from the protein dynamical crossover to the irreversible unfolding) Francesco Mallamace1,2,3, Piero Baglioni4, Carmelo Corsaro1, Sow-Hsin Chen2, Domenico Mallamace5, Cirino Vasi6, Sebastiano Vasi1 and H. Eugene Stanley3 1 Departiment MIFT , Section Physics, Universita di Messina and CNR-IPCF, Italy 2 Department of Nuclear Science and Engineering, MIT, Cambridge, USA 3 Center for Polymer Studies and Department of Physics,Boston University, Boston, USA 4 Dipartimento di Chimica and CSGI, UniversitĂ di Firenze, Italy 5 Dipartimento SASTAS, UniversitĂ di Messina, Messina, Italy 6 CNR-IPCF, Messina, Italy Francesco.Mallamace@unime.it The effect of water on proteins is studied in a very large temperature range from 180 to 370 K. By using in a comparative way the Nuclear Magnetic Resonance and the Neutron scattering we explore this protein system at different hydration level h (h=0.3, 0.37, 0.42 and 0.61). The hydration level h=0.3 is equivalent to a single monolayer of water around the globular protein. Our interest is focused to study the water role in the protein dynamical transition (glass transition or the transition from a harmonic solid like behavior to an anharmonic and liquid like motion) and the irreversible unfolding. We demonstrate also by considering neutron scattering experiments that the protein dynamical transition belongs to the universal class of dynamical crossover characterizing supercooled liquids and materials. The thermal evolution of the spectral features allows identifying that the dynamical crossover observed for water coincides with that of the protein dynamical transition. We stress that we are able to demonstrate at a molecular level the interaction of water with the protein peptides and how via the HB it drives the protein activity. In addition on considering water thermodynamics we identify a special temperature T* that marks the crossover of water, by increasing T, from the state of a complex anomalous liquid to that of a simple

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conventional one. Furthermore, the combination of Scattering and NMR data allows us to clarify the role of T* in the protein properties, in particular T* is the limit of the protein native state. At the same time we are able to clarify at microscopic level the underlying mechanisms that govern the reversibility of the folding-unfolding and irreversible denaturation processes of the protein. New NMR observations at the temperature above and below the protein irreversible unfolding (TD) show that folding-unfolding process takes place as a function of the temperature; we observe that T acts as a control parameter of the measured nuclear magnetization M(T). Whereas far from this singular temperature, in the protein native state, the M(T) behavior is Arrhenius, approaching TD (in a large T-interval) the system changes dramatically it energetic configurations by means a power law behavior. Hence, by following the thermal behavior of different protein-peptide metabolites we are able to explore the funneled energy landscape. On these bases, by taking advantage of the polymer physics we propose this complex process (protein folding/unfolding) as a sort of sol-gel transition driven by water as the cross-linker between different protein peptides, an with TD as the percolation threshold temperature.

november 18-20, 2015 - Barcelona (Spain)

Analysis of the dielectric crossover in liquid water and the possible impact on biological and nanoscopic systems M. I. Marqués1,2, L. M. Maestro1, E. Camarillo3, D. Jaque1, J. García Solé1, J. A. Gonzalo1,4, F. Jaque1, F. Mallamace5, H. E. Stanley6, Juan C. del Valle7, Carmen Aragó1, Karla Santacruz-Gómez8, Roberto C. Carrillo-Torres8 1 Departamento de Física de Materiales, Universidad Autónoma de Madrid, Madrid, Spain 2 Condensed Matter Physics Center (IFIMAC) and Instituto Nicolás Cabrera, UAM, Madrid, Spain 3 Institute of Physics, UNAM, Mexico DF, México 4 Escuela Politécnica, Universidad San Pablo-CEU, Madrid, Spain 5 Dipartimento di Fisica, Università di Messina and CNR-IPCF, Messina, Italy 6 Center for Polymer Studies, and Department of Physics, Boston University, Boston, MA, USA 7 Departamento de Química Física Aplicada, Universidad Autónoma de Madrid, Madrid, Spain 8 Departamento de Física, Universidad de Sonora, Hermosillo, México manuel-marques@uam.es Several properties of liquid water, including the dielectric constant show a bilinear behavior defining a crossover in the temperature range 50 ± 10ºC between two possible states in water (see Figure 1). The existence of these two states in liquid water plays an important role in nanometric and biological systems. For example, the optical properties of metallic (gold and silver) nanoparticles dispersed in water [1], used as nanoprobes, and the emission properties of CdTe quantum dots (QDs), used for fluorescence bioimaging and tumor targeting, show a singular behavior in this temperature range (see Figure 2). In addition, the structural changes in liquid water may be associated with the behavior of biological macromolecules in aqueous solutions and in particular with protein denaturation.

Figures

Figure 1. Temperature dependence of the dielectric constant of water at 0.1 MPa

References [1] Juan C. del Valle, Enrique Camarillo, Laura Martinez Maestro, Julio A. Gonzalo, Carmen Aragó, Manuel I. Marqués, Daniel Jaque, Ginés Lifante, José García Solé, Karla Santacruz- Gómez, Roberto C. CarrilloTorres and Francisco Jaque (2015): Dielectric anomalous response of water at 60 °C, Philosophical Magazine, DOI: 10.1080/14786435.2014.1000419

Figure 2. Temperature dependence of the emission peak wavelength in CdTe quantum dots with an average size of 1.2 nm

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Biological Applications of Carbon-based Nanomaterials: From Functionalisation to Biodegradation Cécilia Menard-Moyon CNRS, Institut de Biologie Moléculaire et Cellulaire, Laboratoire d’Immunopathologie et Chimie Thérapeutique, Strasbourg, France c.menard@ibmc-cnrs.unistra.fr Carbon based-nanomaterials (CNMs), including carbon nanotubes (CNTs) and graphene, are promising tools owing to their outstanding properties and large surface area, offering a variety of opportunities for applications in nanomedicine, such as diagnosis, disease treatment, imaging, and tissue engineering.[1,2] But, the low solubility of CNTs and graphenefamily nanomaterials in most organic solvents and in water hampers their manipulation and limits the full exploitation of their properties. Hence, surface functionalization is crucial to increase the biocompatibility of CNMs and impart multiple functionalities. The oxidized form of graphene, graphene oxide (GO), is often used as starting material for the preparation of graphene derivatives for biomedical applications as the presence of polar oxygencontaining species makes it more hydrophilic than pristine graphene. This is fundamental for further functionalization and processability. Health impact and biopersistence, along with environmental accumulation are key issues for the development of CNMs in the biomedical field and other related areas. It is essential to evaluate their systematic toxicological effects before their use in different domains. In this talk, I will show whole body imaging and pharmacokinetic data following intravenous administration of functionalized GO in mice.[3] Biodistribution studies have important implications in the design of graphene-based nanomaterials for therapy, imaging, and

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diagnosis, as well as for the determination of their safety profile. Understanding human health risk associated with the rapidly emerging graphene-family nanomaterials represents a great challenge because of the diversity of applications and the wide range of possible ways of exposure to this type of materials. It is mandatory to elucidate the key aspects associated with biodegradability of CNMs for their real translation into possible clinical innovations as well as for their safe disposal in the environment. In this context, I will report our study on the biodegradation of GO by myeloperoxidase derived from human neutrophils.[4] The degradation capability of the enzyme on three different GO samples displaying a variable dispersibility in aqueous media has been compared, revealing that MPO failed in degrading the most aggregated GO, but succeeded to completely metabolize highly dispersed GO samples. I will also present our work on the covalent functionalization of CNTs with specific functional molecules such as potential reducing substrates (coumarin derivatives) and redox mediators (catechol) to enhance the catalytic activity of horseradish peroxidase (HRP), leading to accelerated degradation of the nanotubes by the enzyme, in comparison to simply oxidized CNTs.[5] The results demonstrate the crucial importance of the type of surface functionalities onto CNTs as a strategy to modulate their

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enzymatic biodegradability. Our finding will certainly help to guide development of future biomedical applications using CNTs and GO by designing biodegradable carriers for drug delivery.

References [1] C. Ménard-Moyon et al. Expert Opin. Drug Discov. 2010;5:691. [2] K.V. Krishna et al. Nanomedicine (Lond) 2013;8:1669. [3] D.A. Jasim,† C. Ménard-Moyon,† et al. Chem. Sci. 2014;6:3952. [4] R. Kurapati et al. Small 2015;11:3985. [5] A.R. Sureshbabu et al. Biomaterials 2015;72:20.

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november 18-20, 2015 - Barcelona (Spain)

Assembly of G-Quadruplex DNA nanostructure Oscar Mendoza1, 2 and Jean-Louis Mergny1,3 1 Univ. Bordeaux, 33600 Bordeaux, France 2 CBMN, CNRS UMR-5248, F-33600 Pessac, France 3 INSERM, ARNA Laboratory, U869, IECB, F-33600 Pessac, France oscar.mendoza@u-bordeaux.fr DNA’s remarkable molecular recognition and its programmable self-assembly properties have brought this molecule to the materials field [1]. The particular recognition of the base-pare formation allows the construction of supramolecular structures in a nanoscale precision with a large number of potential applications [2]. Appling the well-known W-C base-pair formation is certainly the most common technique for the construction of DNAnanostructures. However, the construction of nanostructures based on non-canonical DNA structures is gaining importance and nanodevices based on motifs such as quadruplex or triplexes have been described [3]. G-quadruplexes are a family of nucleic acid structures based on the formation of two or more G-quartets, in which four co-planar guanines establish a cyclic array of H-bonds, further stabilized by the presence of positively charged cations located in the central channel. In comparison to duplex DNA, G-quadruplex motifs are highly polymorphic: they can be formed by one or several DNA strands (intra or intermolecular); DNA strands may show different polarities (parallel, antiparallel or "3+1"); nature of the central cation, etc.

References [1] Seeman, N. C. Annu. Rev. Biochem. 2010, 79, 65. [2] Yang, D.; Campolongo, M. J.; Nhi Tran, T. N.; Ruiz, R. C. H.; Kahn, J. S.; Luo, D. Wiley Interdiscip. Rev. Nanomedicine Nanobiotechnology 2010, 2, 648. [3] Yatsunyk, L. A.; Mendoza, O.; Mergny, J.-L. Acc. Chem. Res. 2014, 47, 1836. [4] Mendoza, O.; Porrini, M.; Salgado, G. F.; Gabelica, V.; Mergny, J.-L. Chem. - A Eur. J. 2015, 21, 6732. [5] Yatsunyk, L. A.; Piétrement, O.; Albrecht, D.; Tran, P. L. T.; Renčiuk, D.; Sugiyama, H.; Arbona, J. M.; Aimé, J. P.; Mergny, J. L. ACS Nano 2013, 7, 5701.

Figures

In this report we investigated how duplex DNA and G-quadruplex DNA can be combined to create new supramolecular DNA nanostructures. The programmable selfassembly of WC duplexes was employed to create and orientate G-quadruplex structures in a specific conformation. This induced the synapsable formation of quadruplex structures via duplex-formation [4,5].

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Liquid Polymers for Drug Delivery and Regenerative Medicine Boaz Mizrahi Faculty of Biotechnology and Food Engineering, Technion-Israel Institute of Technology, Israel bmizrahi@technion.ac.il

References [1] Neat Mucoadhesive Polymers. Regina Kelmansky and Boaz Mizrahi. Aus. J. Biotech & Bioeng. 1,1 2014. [2] Long-lasting Antifouling Coating from MultiArmed Polymer. Boaz Mizrahi, Xiaojuan Khoo, Homer Chang, Katalina Sher, Jung-Jae Lee, Silvia Irusta and Daniel S. Kohane. Langmuir, 29, 32, 2013. [3] A Stiff Injectable Biodegradable Elastomer. Boaz Mizrahi, Kathryn A. Whitehead, Brian P. Timko, Mike Lawlor, Robert Langer, Daniel G. Anderson, and Daniel S. Kohane., Advanced Functional Materials, 23, 12, 2013.

Liquid Systems Figures

The Structure:

O C O CO O C O

O O

H2O C

Injectable materials often have shortcomings in mechanical and drug-eluting properties that are attributable to their high water contents. For example, tissue adhesives such as cross-linked hydrogels adhere very weakly to tissues while polymersomes are often impermeable to many small organic molecules and as a result show a limited or slow release rate. These drawbacks are related in part to the high molecular weight of currently used biomaterials, that are solid in their basic form and therefore require a high solvent content in order to be administrated. In this presentation I will discuss new strategies for designing neat (without solvent) biomaterials for medicine and biotechnology as well as new concepts in drug delivery and tissue reconstruction. These biomaterials possess the following advantages: (a) they are liquid at room temperature and, therefore, can be applied without the need of solvent; (b) although they have low viscosity at room temperature, they can rapidly harden when crosslinked; (c) they possess a higher number of potentially reactive end groups per molecule compared to high molecular weight polymers of similar molecular weight; and (d) they have low immunogenicity and toxicity.

O C O2 7 C2 H

a 15

a OH

ly mb sse lf-A Se

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november 18-20, 2015 - Barcelona (Spain)

The Quest for Charge Transport in single Adsorbed Long DNA-Based Molecules Danny Porath Institute of Chemistry and Center for Nanoscience and Nanotechnology, The Hebrew University of Jerusalem, Israel danny.porath@mail.huji.ac.il DNA and DNA-based polymers have been at the focus of molecular electronics owing to their programmable structural versatility. The variability in the measured molecules and experimental setups, caused largely by the contact problem, has produced a wide range of partial or seemingly contradictory results, highlighting the challenge to transport significant current through individual DNAbased molecules. A well-controlled experiment that would provide clear insight into the charge transport mechanism through a single long molecule deposited on a hard substrate has never been accomplished. In this lecture I will report on detailed and reproducible charge transport in G4-DNA, adsorbed on a mica substrate. Using a novel benchmark process for testing molecular conductance in single polymer wires, we observed currents of tens to over 100 pA in many G4-DNA molecules over distances ranging from tens to over 100 nm, compatible with a long-range thermal hopping between multi-tetrad segments. With this report, we answer a long-standing question about the ability of individual polymers to transport significant current over long distances when adsorbed a hard substrate, and its mechanism. These results may re-ignite the interest in DNA-based wires and devices towards a practical implementation of these wires in programmable circuits. Keywords: Molecular electronics, Single molecules characterization, DNA-based Nanotechnology

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References [1] "Direct measurement of electrical transport through DNA molecules", Danny Porath, Alexey Bezryadin,Simon de Vries and Cees Dekker, Nature 403, 635 (2000). Cited 1281 times [2] "Charge Transport in DNA-based Devices", Danny Porath, Rosa Di Felice and Gianaurelio Cuniberti, Topics in Current Chemistry Vol. 237, pp. 183-228 Ed. Gary Shuster. Springer Verlag, 2004. Cited 191 times [3] “Direct Measurement of Electrical Transport Through Single DNA Molecules of Complex Sequence”, Hezy Cohen, Claude Nogues, Ron Naaman and Danny Porath, PNAS 102, 11589 (2005). Cited 190 times [4] “Long Monomolecular G4-DNA Nanowires”, Alexander Kotlyar, Nataly Borovok, Tatiana Molotsky, Hezy Cohen, Errez Shapir and Danny Porath, Advanced Materials 17, 1901 (2005). Cited 69 times [5] “Electrical characterization of self-assembled single- and double-stranded DNA monolayers using conductive AFM”, Hezy Cohen et al., Faraday Discussions 131, 367 (2006). Cited 42 times [6] “High-Resolution STM Imaging of Novel Poly(G)-Poly(C)DNA Molecules”, Errez Shapir, Hezy Cohen, Natalia Borovok, Alexander B. Kotlyar and Danny Porath, J. Phys. Chem. B 110, 4430 (2006). Cited 22 times [7] "Polarizability of G4-DNA Observed by Electrostatic Force Microscopy Measurements", Hezy Cohen et al., Nano Letters 7(4), 981 (2007). Cited 55 times [8] “Electronic structure of single DNA molecules resolved by transverse scanning tunneling

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spectroscopy”, Errez Shapir et al., Nature Materials 7, 68 (2008). Cited 84 times [9] “A DNA sequence scanned”, Danny Porath, Nature Nanotechnology 4, 476 (2009). [10] “The Electronic Structure of G4-DNA by Scanning Tunneling Spectroscopy”, Errez Shapir, et.al., J. Phys. Chem. C 114, 22079 (2010). [11] “Energy gap reduction in DNA by complexation with metal ions”, Errez Shapir, G. Brancolini, Tatiana Molotsky, Alexander B. Kotlyar, Rosa Di Felice, and Danny Porath, Advanced Maerials 23, 4290 (2011). [12] "Quasi 3D imaging of DNA-gold nanoparticle tetrahedral structures", Avigail Stern, Dvir Rotem, Inna Popov and Danny Porath, J. Phys. Cond. Mat. 24, 164203 (2012). [13] "Comparative electrostatic force microscopy of tetra- and intra-molecular G4-DNA", Gideon I. Livshits, Jamal Ghabboun, Natalia Borovok, Alexander B. Kotlyar, Danny Porath, Advanced materials 26, 4981 (2014). [14] "Long-range charge transport in single G4-DNA molecules", Gideon I. Livshits et. al., Nature Nanotechnology 9, 1040 (2014).

Figures

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november 18-20, 2015 - Barcelona (Spain)

Graphene in neural interface systems: perspectives and applications Elisabet Prats-Alfonso Institut de MicroelectrĂ˛nica de Barcelona, IMB-CNM (CSIC), Campus UAB, 08193. Barcelona, Spain Centro de Investigacion Biomedica en Red, Biomateriales y Nanomedicina (CIBER-BBN), Madrid, Spain elisabet.prats@csic.es One of the scientific challenges of the coming years is to advance in the study and understanding of the brain function. Recent advances in micro and nanotechnologies have generated a wide interest in applying them to build neural prostheses. These systems provide the neurophysiologists with tools for recording with multiple electrodes the neural activity in in vivo and also in vitro conditions. However, despite the advances in this technology, there are still some drawbacks to be tackled [1]. An ideal neural interface should create seamless integration [2] into the tissue to allow its reliability for long periods of time. Additionally, it should register the brain activity with enough accuracy to obtain relevant information from neural signals. To accomplish all these requirements the interface material should include many physical and chemical properties [3]. The latest advances in new materials [4] are being used to build novel neural prostheses that provide improved signal/noise ratio and greater biocompatibility. In this regard, due to its properties, graphene seems to be one of the most promising materials to provide an improved biologically-artificial interface. In particular, the flexibility, biocompatibility and chemical stability among other properties, place the graphene in a privileged position for being the ideal neural interface [5].

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In our group we have started to develop electrodes and Solution Gated Field Effect Transistors (SGFETs) based on CVD graphene to assess the potential of this new material for recording neural signals. Interestingly, its use for building SGFETs would allow the reduction of the sensing area and thus can increase the density of active points. Here, we present a general overview of the latest advances in neural interfaces systems, and specifically to the graphene based ones which have been carried out by our group.

References [1] C. Riggio, G. Ciofani, V. Raffa, S. Bossi, S. Micera, A. Cuschieri, Polymeric thin film technology for neural interfaces: Review and perspectives, www.intechopen.com, (2010). [2] P. Fattahi, G. Yang, G. Kim, M. R. Abidian, Adv. Mater. (2014), 26, 1846-1885. [3] X. Navarro, T. B. Krueger, N. Lago, S. Micera, T. Stieglitz, P. Dario, Journal of the Peripheral Nervous System (2005), 10, 229-258. [4] S. Unarunotai, Y. Murata, C. E. Chialvo, H.-s. Kim, S. MacLaren, N. Mason, I. Petrov, J. A. Rogers, Appl. Phys. Lett. (2009), 95, 202101-202101202103. [5] L. H. Hess, M. Jansen, V. Maybeck, M. V. Hauf, M. Seifert, M. Stutzmann, I. D. Sharp, A. OffenhĂ¤usser, J. A. Garrido, Adv. Mater. (2011), 23, 5045-5049.

november 18-20, 2015 - Barcelona (Spain)

Hybrid micro and nanoBots as future active drug carriers Samuel Sánchez Institute for Bioengineering of Catalonia (IBEC), Barcelona, Spain Institut Català de Recerca i Estudis Avancats (ICREA), Barcelona, Spain Max Planck for Intelligent Systems, Stuttgart, Germany ssanchez@ibecbarcelona Most of the current drug delivery systems (DDS) rely on passive transport into the fluid where they swim, which is not ideal for delivering payloads to specific locations with high efficiency. It was envisioned decades ago that scientists would engineer tiny nano-bots that actively and directly transport payloads to specific locations. Ever since then, great advances have been made in the field of nanorobotics, however it has been only until very recently when biocompatible, metal-free motors using biocompatible fuels have been reported. Mimicking biomotors, scientists used catalytic reaction to power artificial nano-bots. [1]. Nanomotors demonstrated the transport of drugs [2] micro-objects [3] and cells [4] with wireless magnetic guidance [5], temperature [6], and light control [7]. Furthermore, they can act collectively reacting to external stimuli like chemotactic behaviour [8] and are capable of cleaning polluted water [9]. Here, I will present our recent developments in this fascinating field. We fabricate nano-bots from mesoporous silica nanoparticles, microspheres and rolled-up thin films into microtubular jets. Very recently, we have found that hybrid Micro-bio-bots combine the best from the two worlds, biology and nanomaterials providing very promising bio-related applications.

Keywords: nanomotors, nanotechnology, drug delivery, active matter, self-propulsion, bots.

References [1] S. Sanchez, Ll. Soler and J. Katuri. Angew.Chem.Int.Edit. 54,1414-1444 (2015) [2] X. Ma, K. Hahn and S. Sanchez. J. Am. Chem.Soc. 137 (15), 4976–4979 (2015) [3] A. A. Solovev et al, Adv. Funct. Mater., 20, 2430 (2010); Solovev A. A. et al., ACS Nano, 6, 1751. (2012) [4] S. Sanchez et al, Chem. Commun., 47, 698. (2011) [5] Khalil, App. Phys. Lett. 103, 172404 (2013) [6] S. Sanchez et al, J. Am. Chem. Soc., 133, 4860 (2011); Soler et al. LabChip 13, 4299 (2013) [7] Solovev A.A. et al, Angew. Chem. Int. Ed., 50, 10875. (2011); [8] Baraban, L. et al., Angew. Chem. Int. Ed., 52, 5552. (2013) [9] Soler, Ll. et al., ACS Nano, 7, 9611 (2013); Gao, W., Wang, J., ACS Nano, 8, 3170 (2014); Soler, L. Sanchez, S. Nanoscale, 6, 7175 (2014)

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november 18-20, 2015 - Barcelona (Spain)

Single molecule graphene biosensors: chemistry matters Gregory F. Schneider Universiteit Leiden, The Netherlands g.f.schneider@chem.leidenuniv.nl Exploiting the full potential offered by graphene in sensing applications requires extensive fundamental studies of the behaviour of the surface and of the edges of graphene upon their interaction with biological systems (lipids, proteins, enzymes, DNA, RNA and ultimately biological cells), as well as a quantification of the measurable electronic response of the graphene surface (and edge respectively) caused by a biological stimuli such as the presence and the passage of a biomolecule. The surface and the edges of graphene operate as sensors in two fundamentally different ways: in a typical solution-gated graphene field-effect transistor, the surface is sensitive to charge transfer conferred by a molecule in the vicinity of graphene and therefore could potentially detect a single molecule as a whole, while edges can be used as atomically flat electrodes that could transversally sense the precise structure and chemical composition of a biomolecule passing close to the edges. In both cases, biomolecules are being sensed, but the level of output information is different: surfaces can trap, detect and sense while edges can provide sequence information. This holds the potential

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that one can combine both and use the surface to selectively trap and identify, guide electrophoretically the trapped molecule towards the edge, and obtain molecular information; for example, using a transverse electrochemical current generated between two edges separated by a physical gap on the order of the lateral dimension of the biomolecule.

In my research group, we conduct interdisciplinary research on graphene in the field of bionanotechnology. We particularly investigate the chemical properties of graphene from the perspective of using this material, for example, as a sensor by exploiting its unique surface and edge reactivity. To these ends, graphene has three fantastic properties: it conducts electricity outstandingly well, its edge is only a single carbon atom thin, and the fact that all the atoms are located on the surface makes graphene very sensitive to nearby environmental changes.

november 18-20, 2015 - Barcelona (Spain)

Personalized Cancer Nanomedicine Avi Schroeder Assistant Professor of Chemical Engineering, Technion â&#x20AC;&#x201C; Israel Institute of Technology, Israel avids@technion.ac.il The field of medicine is taking its first steps towards patient-specific care. Our research is aimed at tailoring treatments to address each personâ&#x20AC;&#x2122;s individualized needs and unique disease presentation. Specifically, we are developing nanoparticles that target disease sites, where they perform a programmed therapeutic task. These systems utilize molecular-machines and cellular recognition to improve efficacy and reduce side effects. Two examples will be described: the first involves a nanoscale theranostic system for predicting the therapeutic potency of drugs against metastatic cancer. The system provides patient-specific drug activity data with singlecell resolution. The system makes use of barcoded nanoparticles to predict the therapeutic effect different drugs will have on the tumor microenvironment.

The second system makes use of enzymes, loaded into a biodegradable chip, to perform a programed therapeutic task â&#x20AC;&#x201C; surgery with molecular precision. Collagenase is an enzyme that cleaves collagen, but not other tissues. This enzyme was loaded into the biodegradable chip and placed in the periodontal pocket. Once the collagenase releases from the chip, collagen fibers that connect between the teeth and the underlying bone are relaxed, thereby enabling enhanced orthodontic corrective motion and reducing pain. This new field is termed BioSurgery. The clinical implications of these approaches will be discussed.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Targeting cancer stem cells to increase sensitivity to chemotherapy Sim贸 Schwartz Vall d'Hebron Hospital - CIBBIM, Spain simo.schwartz@vhir.org Drug delivery by nanoparticles may well circumvent the resistance machinery of cancer stem cells (CSC). To be able to study efficacy of nanomedicines in population of CSC, we first developed an in vitro model in which CSC are tagged by a fluorescent reporter gene under the control of a CSC specific promoter. Using this system, we demonstrated that while bulk cancer cells die, CSC population augments after paclitaxel (PTX) treatment. We then investigated the prospects of different targeted and nontargeted delivery systems loaded with PTX and functionalized with specific antibodies against cancer stem cell populations in regular breast cancer cell lines, as well as in our CSC models. Our data shows that reducing tumor resistance of cancer stem cells might be related to specific active targeting of DDS and not attributed to a general mechanism of action of nanomedicines

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Nano crystalline cellulose-protein composites: Super performing biomaterials for tissue engineering and regenerative medicine Oded Shoseyov The Robert H Smith Institute of Plant Science and genetics. The Faculty of Agriculture, The Hebrew University of Jerusalem, Israel oded.shoseyov@mail.huji.ac.il A platform technology that brings together the toughness of cellulose nano-fibers from the plant kingdom, the remarkable elasticity and resilience of resilin that enables flees to jump as high as 400 times their height from the insect kingdom, and the adhesion power of DOPA, the functional molecule of mussels that enable it to bind tightly under water to organic and inorganic matter from the marine kingdom and all that combined with Human Recombinant Type I collagen produced in tobacco plants; SUPERPERFORMING BIOMATERIALS. Resilin is a polymeric rubber-like protein secreted by insects to specialized cuticle regions, in areas where high resilience and low stiffness are required. Resilin binds to the cuticle polysaccharide chitin via a chitin binding domain and is further polymerized through oxidation of the tyrosine residues resulting in the formation of dityrosine bridges and assembly of a high-performance proteincarbohydrate composite material. Plant cell walls also present durable composite structures made of cellulose, other polysaccharides, and structural proteins. Plant cell wall composite exhibit extraordinary strength exemplified by their ability to carry the huge mass of some forest trees. Inspired by the remarkable mechanical properties of insect cuticle and plant cell walls we have developed novel composite materials of resilin and NanoCrystalline Cellulose (resiline-NCC) that display remarkable mechanical properties combining

strength and elasticity. We produced a novel resilin protein with affinity to cellulose by genetically engineering a cellulose binding domain into the resilin. This CBD-Resilin enable, interfacing at the nano-level between the resilin; the elastic component of the composite, to the cellulose, the stiff component. Furthermore, chemical and enzymatic modifications of the composite are developed to produce DOPAResiline-NCC which confers adhesive and sealant properties to the composite. As a central element of the extracellular matrix, collagen is intimately involved in tissue development, remodeling, and repair and confers high tensile strength to tissues. Numerous medical applications, particularly, wound healing, cell therapy, and bone reconstruction, rely on its supportive and healing qualities. Its synthesis and assembly require a multitude of genes and posttranslational modifications. Historically, collagen was always extracted from animal and human cadaver sources, but bare risk of contamination and allergenicity and was subjected to harsh purification conditions resulting in irreversible modifications impeding its biofunctionality. In parallel, the highly complex and stringent post-translational processing of collagen, prerequisite of its viability and proper functioning, sets significant limitations on recombinant expression systems. A tobacco plant expression platform has been recruited to effectively express human collagen,

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

along with three modifying enzymes, critical to collagen maturation. The plant extracted recombinant human collagen type I forms thermally stable helical structures, fibrillates, and demonstrates bioactivity resembling that of native collagen. Combining collagen at the

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nano-scale with resilin to produce fibers resulted in super-performing fibers with mechanical properties which exceed that of natural fibers.

november 18-20, 2015 - Barcelona (Spain)

Nanomechanical and Optomechanical Systems for Cancer Research Javier Tamayo Bionanomechanics lab, Institute of Microelectronics of Madrid, CSIC, Madrid, Spain jtamayo@imm.cnm.csic.es The advances in micro- and nanofabrication technologies are enabling increasingly smaller mechanical transducers capable of detecting the forces, motion, mechanical properties and masses that emerge in biomolecular interactions and fundamental biological processes. Thus, biosensors based on nanomechanical systems have gained considerable relevance in the last decade[1]. This talk will provide insight into the mechanical phenomena that occur in suspended mechanical structures when either biological adsorption or interactions take place on their surface. In addition, I will show how coupling nanomechanics and nanooptics allows to achieve sensing devices with higher performance and novel transduction paradigms. I will describe then some relevant experiments running in our laboratory that harness nanomechanical and optomechanical systems for cancer research in three battlefronts: 1) ultrasensitive detection of cancer biomarkers in blood [2], ii) cancer cell nanomechanics [3], and iii) nanomechanical spectrometry [4-5].

References [1] Tamayo, J., Kosaka, P. M., Ruz, J. J., San Paulo, Ă . & Calleja, M. Biosensors based on nanomechanical systems. Chemical Society Reviews 42, 1287-1311 (2013). [2] Kosaka, P.; Pini, V.; Ruz, J.; da Silva, R.; GonzĂĄlez, M.; Ramos, D.; Calleja, M.; Tamayo, J., Detection of cancer biomarkers in serum using a hybrid mechanical and optoplasmonic nanosensor. Nature Nanotechnology 9, 1047-1053 (2014). [3] Encinar, Calzado et al, in preparation [4] Gil-Santos, E. et al. Nanomechanical mass sensing and stiffness spectrometry based on two-dimensional vibrations of resonant nanowires. Nature Nanotechnology 5, 641-645 (2010). [5] Ramos, D. et al. Optomechanics with silicon nanowires by harnessing confined electromagnetic modes. Nano letters 12, 932937 (2012).

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Non-covalent functionalisation of graphene: Optimising molecular packing density and stability Sinéad Winters, Nina C. Berner and Georg S. Duesberg Trinity College Dublin, College Green, Dublin 2, Ireland swinters@tcd.ie The application of graphene in gas and biological sensors is currently of great interest due to the high sensitivity of graphene to external influences. However, in order to produce selective sensors the graphene surface must be modified to respond to a specific analyte [1]. Non-covalent functionalization of graphene via π-π stacking of aromatic molecules is an attractive strategy as it allows surface modification without disturbing the graphene lattice [2]. Implementing this technique faces certain challenges, such as characterising the molecular layers, particularly in the presence of contamination on the graphene surface. We investigated the non-covalent functionalization of chemical vapour deposited (CVD) graphene with aromatic molecules via wet chemical adsorption. A method of producing high-packing density molecular films on graphene has been developed by addition of a functionalization step to the standard CVD graphene transfer process. Raman spectroscopy and scanning tunneling microscopy give insight into the arrangement and packing density of the molecules on the graphene surface.

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The influence of packing density on the stability of the molecules upon further derivatisation was also studied, which is important for additional of selective sensing markers onto graphene. X-ray photoelectron spectroscopy (XPS) is used to characterise the surface modification. These measurements provide insight into the behaviour of molecules on pristine graphene, an important step towards implementation in selective sensing applications.

References [1] Allen, M. J.; Tung, V. C.; Kaner, R. B., Chem. Rev. 110 (2010) 132–145. [2] Mann, J.; Dichtel, W., J. Phys. Chem. Lett. 4 (2013) 2649–2657.

november 18-20, 2015 - Barcelona (Spain)

Graphene for Biomedical Applications Amaia Zurutuza Graphenea S.A., Tolosa Hiribidea 76, Donostia - San Sebastian, Spain a.zurutuza@graphenea.com Graphene has emerged as a very interesting nanomaterial for biomedical applications. Graphene based materials could be used in sensing and diagnostic platforms as well as for diverse medical treatments. During this talk I will cover the use of CVD graphene in biosensors [1] and functionalized graphene oxide for the targeted treatment of bacteria [2]. CVD graphene in combination with gold nanoparticles has shown attomolar sensitivity to detect DNA hybridization processes.1 Furthermore, this could be extended to the use of graphene plasmons in sensing applications [3,4]. On the other hand, nanopores in CVD graphene could in the future provide a platform for the sequencing of DNA [5]. In addition, the future industrialization of graphene in some of these applications will require large scale, reliable and relatively fast fabrication methods, therefore, I will also cover some relevant aspects for the processing of graphene devices.

References [1] O. Zagorodko, J. Spadavecchia, A. Yanguas Serrano, I. Larroulet, A. Pesquera, A. Zurutuza, R. Boukherroub, and S. Szunerits, Anal. Chem., 86, (2014), 11211. [2] K. Turcheniuk, C.-H. Hage, J. Spadavecchia, A. Yaguas Serrano, I. Larroulet, A. Pesquera, A. Zurutuza, M. Gonzalez Pisfil, L. Héliot, J. Boukaert, R. Boukeherroub, and S. Szunerits, J. Mater. Chem. B, 3, (2015), 375. [3] J. Chen, M. Badioli, P. Alonso-González, S. Thongrattanasiri, F. Huth, J. Osmond, M. Spasenović, A. Centeno, A. Pesquera, P. Godignon, A. Zurutuza Elorza, N. Camara, F. J. García de Abajo, R. Hillenbrand, and F.H.L. Koppens, Nature, 487, (2012), 77. [4] P. Alonso-González, A.Y. Nikitin, F. Golmar, A. Centeno, A. Pesquera, S. Vélez, J. Chen, G. Navickaite, F. Koppens, A. Zurutuza, F. Casanova, L. E. Hueso, and R. Hillenbrand, Science, 344, (2014), 6190. [5] W.L. Wang, E.J.G. Santos, B. Jiang, E.D. Cubuk C. Ophus, A. Centeno, A. Pesquera, A. Zurutuza, J. Ciston, R. Westervelt, and E. Kaxiras, Nano Lett., 14, (2014), 450. [6] D.M.A. Mackenzie, J.D. Buron, P.R. Whelan, B.S. Jessen, A. Silajdzic, A. Pesquera, A. Centeno, A. Zurutuza, P. Bøggild and D.H. Petersen, 2D Mater., 2, (2015), 045003. [7] K. Asadi, E.C. Timmering, T.C.T. Geuns, A. Pesquera, A. Centeno, A. Zurutuza, J.H. Klootwijk, P.W.M. Blom and D.M. de Leeuw, ACS Appl. Mater. Inter., 7, (2015), 9429. [8] A.A. Sagade, D. Neumaier, D. Schall, M. Otto, A. Pesquera, A. Centeno, A. Zurutuza Elorza and H. Kurz, Nanoscale, 7, (2015), 3558.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Improved Pharmacokinetic Profile of Lipophilic Anti-Cancer Drugs Using Targeted Polyurethane-Polyurea Nanoparticles Yolanda Fernández1,2, Pau Rocas3, Natalia García-Aranda1,2, Laia Foradada1,2, Pilar Calvo4, Pablo Avilés4, María José Guillén4, Simó Schwartz Jr.1,2, Josep Rocas5, Fernando Albericio3,6, Ibane Abasolo1,2 Vall d'Hebron University Hospital-UAB, Functional Validation & Preclinical Studies. Barcelona, Spain Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Spain 3 Institute for Research in Biomedicine of Barcelona, Barcelona, Spain 4 PharmaMar S.A., Colmenar Viejo, Madrid, Spain 5 Ecopol Tech S.L., L'Arboç,Tarragona, Spain 6 Department of Organic Chemistry, University of Barcelona, Barcelona, Spain 1

ibane.abasolo@vhir.org Meticulously designed nanosystems conveying multiple functionalities and complex multiwalled nanostructures will likely enhance pharmacokinetics and biodistribution of lipophilic chemotherapeutic agents. In the present study, polyurethane-polyurea nanoparticles (PUUa NP) with a disulfide-rich multiwalled structure have been synthesized, characterized and biologically tested. These novel nanoparticles were prepared using recently described procedures [1], with slight changes in size, shell structure, loading capacities and z-potential to improve the in vivo behaviour of final nanoparticles. Specifically, glutathione degradable PUUa NP were synthetized with sizes around 100 nm, a neutral zeta potential and tremendous encapsulation efficiency (above 90%) for a very lipophilic anticancer drug, called plitidepsin [3]. Moreover, PUUa NP were functionalized with a cyclic RGD peptide [2] binding αVβ3 integrins, for further improving their biodistribution and cell internalization. In vitro cell toxicity assays revealed that PUUa NP release their cargo, rendering half maximal inhibitory concentration (IC50) values similar to those of the free plitidepsin. In vivo toxicity studies indicated that the maximum tolerated

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dose (MTD) of plitidpesin could be reduced from 3 mg/kg to 0.9 mg/kg when the drug was delivered in PUUa NP. Moreover, pharmacokinetic assays performed in athymic nude mice showed that pitidepsin in PUUa NP had higher values of maximum concentration (Cmax), area under curve (AUC) and plasma half-life times as compared to the free drug. Biodistribution assays were also performed in athyimic nude mice by fluorescent imaging (FLI) of fluorescently (DiR) labelled PUUa NP. Interestingly, in vivo FLI showed that RGD-decorated PUUa NP tended to accumulate less in the liver. Ex vivo analysis of FLI and plitidepsin content confirmed that integrin targeted nanoparticles had lower accumulation rates in liver and spleen, suggesting that targeted nanoparticles avoid sequestration by macrophages from the reticuloendothelial system. Overall our results indicate that the polyurethane-polyurea nanoparticles represent a very valuable nanoplatform for the delivery of lipophilic drugs, such plitidpesin. Without affecting its cytotoxic activity, encapsulation of plitidepsin improves the pharmacokinetic and toxicological profile of the drug, and in addition, the functionalization of the PUUa NP surface with targeting moieties seems to improve the stealh properties of the delivery system.

november 18-20, 2015 - Barcelona (Spain)

References [1] Rocas P, Fernández Y, Schwartz S Jr, Abasolo I, Rocas J, Albericio F. J Mat Chem B (2015). [2] Dai X, Su Z, Liu JO. Tetraedron Lett 41 (2000) 6295-6298. [3] Muñoz-Alonso, González-Santiago L, Martínez T, Losada A, Galmarini CM, Muñoz A. Curr Opin Investig Drugs 10 (2009) 536-42.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Microfluidic bioassay based on bacterial luciferase and NADH:FMN-oxidoreductase PI Belobrov1,2, IA Denisov1, КA Lukyanenko1, AS Yakimov1, EN Esimbekova1,2, KI Belousov3, AS Bukatin1, IV Kukhtevich1, VV Sorokin1, AA Evstrapov3,4 Siberian Federal University, 79 Svobodny Pr., 660041, Krasnoyarsk, Russia Institute of Biophysics SB RAS, 50/50 Aсademgorodok St., 660036, Krasnoyarsk, Russia 3 ITMO University, 49 Kronverksky Pr., 197101, St. Petersburg, Russia 4 Institute for Analytical Instrumentation of RAS, 31 Ivana Chernykh, 198095, St. Petersburg, Russia 1

peter.belobrov@gmail.com The disposable microfluidic chips [1] for measurement of water pollution can be made with biomodule based on enzymes of luminous bacteria: luciferase and NADH:FMNoxidoreductase [2, 3]. Interaction of organic and inorganic pollutants with these enzymes from the bacterial luminescence system leads to quenching of light emission and changing of shapes of measured kinetic curves. Here we show the automation of this biomodule for ecological bioassay with microfluidic techniques. Luminescence chemical reactions in biomodule are described by the equations: NADH:FMN-oxidoreductase NADH + FMN + H⁺ ———————→ FMNH₂ + NAD⁺ luciferase FMNH₂ + RCHO + O₂——→ FMN + RCOOH + H₂O + hν The components of the reaction: luciferase (EC 1.14.14.3) from Photobacterium leiognathi, NADH:FMN-oxidoreductase (EC 1.6.99.3) from Vibrio fischeri, NADH and tetradecanal were immobilized in gel made from potato starch and placed in reactor chamber. FMN for reaction activation was deposited separately in chip by the process of droplet drying. Channelized surface of poly(methyl methacrylate) microfluidic chips was formed by direct cutting with the milling machine MDX-20 (Roland, Japan). Sealing of microfluidic chips was carried out by spraying of acetone or

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dichloroethane on flat half of chip followed by pressing of channelized half with pressure ~ 4 N/mm². The influence of acetone and 1,2dichloroethane on enzymes was estimated. The kinetics of light emission during bioluminescent reaction was recorded with the single tube luminometer GloMax 20/20 (Promega, USA). The bioluminescent reaction starts when FMN mixed with components around gel film solving in reaction chamber. As FMN activates the reaction of light emission it is important to have a uniform concentration of FMN in all parts of the reaction chamber. To improve mixing and achieve reproducible results different topologies of microfluidic chip (Fig. 1) were studied. Dynamics of FMN dissolution and mixing was studied using numerical simulations. It was shown that FMN dissolution takes 0.7–0.8 s depending on the flow rate but the passive mixing efficiency is not enough to achieve uniform concentration profiles in the reaction chamber. Because of that we suppose that active mixing strategies should be implemented in the microfluidic chip for increasing results reproducibility. In topology (c) sample proceeded through the input channel into a serpentine mixer with dried FMN, where it dissolved FMN, and then stirred with it. Dissolved FMN with a sample then

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entered the reaction chamber. In (b) and (c) passive mixing was used and in (d) and (e) active mixing. In topology (d) FMN was dried next to the immobilized enzyme system in reaction chamber. When the flow got to the chamber it stopped at the output hole, which was located in the end of the reaction chamber. Then the active mixing started for a few seconds followed by the measurement of the kinetics. Topology (e) differs from (d) in a way that the flow came to the reaction chamber from two different directions and stopped at the “stopline”. Active mixing join two droplets through this “stop-line“ and mixing them, then the measurement of kinetics is starting.

References [1] Sackmann, E., Fulton, A., Beebe, D., Nature, 507 (2014) 181-189. [2] Esimbekova E., Kondik A., Kratasyuk V., Environmental monitoring and assessment, 7 (2013) 5909-5916. [3] Esimbekova E., Kratasyuk V., Shimomura O., Adv Biochem Eng Biotechnol, 144 (2014) 67-109.

Figures

The influence of surface modification of microfluidic chip to the properties of immobilized luminescent system components was studied in topology (a) with 4 different surfaces (Fig. 2). The experiments were carried out with clean water without any pollutant. The immobilization with gelatin (3) was shown to be the best option. We assume, that gelatine protects enzymes during drying process, so they provide better repeatability of measurements and the highest intensity. Observed decreasing of luminescence in sealed chips is due to the lack of O₂ in the reaction chamber. Active mixing provides more uniform concentration of FMN in reaction chamber and therefore is more preferable. Dean vortices significantly influence the speed of FMN dissolution. Starch gel film on top of gelatin film provides maximum reproducibility of the results. Acknowledgement The research was supported by the grant of the Russian Science Foundation (project no.15-1910041).

Figure 1. Evaluated microfluidic chip topologies. Inlets at the bottom. Dried gel droplets are shown by nets. Dried FMN droplets are shown by dots.

Figure 2. Maximal intensity of bioluminescent reaction for unsealed (black) and sealed (grey) microfluidic chips with different types of surface treatment before compounds immobilization: (1) the droplet with reactants was dried in a reactor chamber (2) the PMMA was treated with 1M NaOH solution for 18 hours, then washed and a droplet of gel was dried in a reaction chamber (3) surface of reaction chamber was covered with gelatin, then the droplet of gel with reactants was dried on top of it (4) the surface of reaction chamber was modified with amylose and amylopectin molecules covalently bonded by ethylenediamine dihydrochloride to it. Percent demonstrates the decrease in luminescence intensity in sealed chip compared to unsealed chips.

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november 18-20, 2015 - Barcelona (Spain)

Bicosome platform for Skin Drug delivery Rafael Bernad, Lucyanna Barbosa-Barros, Gelen Rodríguez, Mercedes Cócera, Olga López Bicosome S.L. c/ Jordi Girona 18, 08034 Barcelona, Spain

Rafael.Bernad@bicosome.com The main function of our skin is to act as a barrier protecting our body from external aggression. This is regulated mainly by the outermost layer of the skin, the stratum corneum (SC). Skin tissue provides a strong barrier function which impedes effective treatments. Skin penetration is therefore one of the biggest challenges for formulators of skin care products. Most topical ingredients available on the market do not really penetrate the SC, acting only on the skin’s surface and disappearing after a single wash. Other ingredients include chemical enhancers and/or other aggressive compounds that penetrate disrupting the skin barrier and causing damage. In both cases, efficacy and safety are compromised. Thus, there is a necessity of products able to penetrate gently the skin, remain there, and deliver their content in the targeted layers. Bicosome has developed a platform that offers a solution to these challenges. Structurally, Bicosomes are made up of internal smart biocompatible structures enclosed in a lipid vesicle that protects and boosts their effects. A number of active molecules can be allocated in the different compartments of this platform. The smart structures of the bicosomes are small enough to penetrate the skin and self-aggregate into the tissue and grow being retained in specific layers. This induces a reinforcement of the skin structures and a targeted delivery. This delivery strategy allows a prolonged effect of actives because bicosome components retained in specific target layers remain there

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until this layer is lost following the natural desquamation process of the skin. These systems open a new and disruptive strategy, in which actives are addressed specifically into the target layers and remain there exerting their action for days.

References [1] A.N. Ellepola, G.J. Panagoda, L.P. Samaranayake, Microbiol. Immunol., 1999, 14:358-363. [2] C. Surber, P. Elsner, M. Farage, Curr. Probl. Dermatol., 2011, 40. [3] C.S. Boon, et al. Crit. Rev. Food Sci. Nutr., 2010, 50, 215-532. [4] E. Fernández, et al., RCS Advances, 2014, 4, 53109-53121. [5] Elias, P.M. J. Control. Release, 1991, 15: 199208. [6] G. Rodríguez, G. Soria, E. Coll, L. Rubio, L. Barbosa-Barros, C. López-Iglesias, A. M. Planas, J. Estelrich, A. de la Maza, O. López, Biophys. J., 2010, 99,480. [7] G. Rodríguez, L. Barbosa-Barros, L. Rubio, M. Cócera, C. López-Iglesias, A. de la Maza, O. López, Colloids and Surfaces B: Biointerfaces, 2011, 84, 390. [8] J. W. Wiechers, Science and Applications of Skin Delivery Systems, Allured Publishing Corporation, 2008. [9] L. Barbosa-Barros, G. Rodríguez, C. Barba, M. Cócera, L. Rubio, J. Estelrich, C. LópezIglesias, A. de la Maza, O. López, Small, 2012, 8, No 6, 807. [10] L. Rubio, G. Rodríguez, L. Barbosa-Barros, C. Alonso, M. Cócera, A. de la Maza, J.L. Parra,

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O. L贸pez, Colloids and Surfaces B: Biointerfaces, 2012, 92, 322. [11] M. C贸cera, et al. Soft Matter, 2011, 7, 86058611. [12] M.E. Darwin, et al., J Dermatol. Sci., 2011, 64, 53-58. [13] A. Holzer, M. Athar, C. Elmets, J. Invest. Dermatol., 2010, 130(6), 1496-1499. [14] C. Calles et al., J. Invest. Dermatol., 2010, 130: 1424-36. [15] T. Herrling, et al., Spectrochim Acta A Mol Biomol Spectrosc., 2006, 63(4), 840-845.

Figures

Figure 1. Illustration of Bicosome structure and penetration mechanism.

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november 18-20, 2015 - Barcelona (Spain)

Biomechanics : from in vitro to in vivo Remy Brossel Cell Constraint & Cancer, France

Malignant tissue has been “normalized” in vitro using mechanical signals (MJ Paszek, 2005; F Montel, 2011, M Olcum, 2014). We apply the same principles of mechanobiology in vivo to show the effect of a ‘constraint field’, here a pressure, on tumor growth. Human breast cancer cell line MDA MB 231 admixed with ferric nanoparticles was grafted subcutaneously into mice. The nanoparticles rapidly surrounded the tumor. We applied a constraint field, and by consequence biomechanical signals, accross the transplanted tumor in vivo using two permanent magnets located on either side of the tumor which was surrounded by ferric nanoparticles.

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This biomechanical treatment was applied 2 hours/day during 21 days. There was a significant difference between the volume of treated tumors and untreated controls. In addition to significant difference in volume in vivo, there were morphologic – surface - differences on ex vivo histologic examination of the excised tumor masses. This first evidence of the action of stress on tumor growth in vivo means that biomechanical intervention may have a high translational potential as a therapeutic tool specially in locally advanced tumors like pancreatic cancer or primary hepatic carcinoma.

november 18-20, 2015 - Barcelona (Spain)

High Resolution Maps of Apparent Young’s Modulus of Breast Cancer Cells by PeakForce Modulation Atomic Force Microscopy Alicia Calzado–Martín, Mario Encinar, Javier Tamayo, Montserrat Calleja, Álvaro San Paulo Instituto de Microelectrónica de Madrid (CSIC), Isaac Newton 8, 28760, Tres Cantos, Madrid. Spain

alicia.calzado@csic.es Metastasis is the leading cause of death from breast cancer [1]. The physical changes taking place in metastatic cells remain unclear, due to the absence of suitable instrumentation and methodologies. To date, AFM has provided information mostly about the apparent Young's modulus of different cell types by the use of local F–z curves [2–4]. Namely, Peak-Force modulation (PFM) mode uses the maximum repulsive force registered during each actuation cycle of the probe as the feedback parameter, enabling quantitative measurements of mechanical properties at high speed and high resolution [5]. In this work we use PFM mode for comparative elasticity imaging of three human epithelial breast cell lines with different degree of malignancy; MCF-10A (healthy), MCF-7 (tumorigenic, non-invasive) and MDA-MB-231 (tumorigenic, invasive). We established the optimum specific image acquisition and analysis parameters for simultaneous high resolution topography and apparent Young’s modulus imaging of living cells in liquid media. Furthermore, we describe systematic differences found in contrast patterns from healthy and tumor cells (Fig. 1). These differences are resolved as a consequence of the unique high-resolution capability of PFM mode, and they provide crucial information for understanding differences in stiffness between normal and cancerous cells. The origin of these differences is investigated by complementary immunofluorescence assays, and a correlation

is found between the features detected in the apparent Young’s modulus images and the cytoskeleton arrangement (Fig. 2). The complementary information obtained from both techniques allows evaluating the relative contributions to cell stiffness of filamentous actin and microtubules in healthy cells, as compared to tumorigenic ones. Our results show that high resolution PFM mode AFM imaging of live cells provides otherwise inaccessible information about local cytoskeletal conformations that determine the disparate mechanical properties of cancerous and normal cells. In the particular case of the three epithelial breast cell lines assessed in this work, the combination of AFM and fluorescence microscopy imaging allows to establish a relationship between the contributions of Factin and microtubule cytoskeleton and the cell stiffness: F-actin forms structures around one order of magnitude stiffer than microtubules as mapped in high resolution apparent Young’s modulus images of healthy cells. Such F-actin structures are observed mainly in healthy cells, while their reduced presence in both tumorigenic cell lines explains the different global stiffness of the studied cell lines.

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References [1] Lu J, Steeg PS, Price JE, Krishnamurthy S, Mani SA, Reuben J, et al. Cancer Res. 2009:15;69(12):4951-3. [2] Li QS, Lee GY, Ong CN, Lim CT. Biochem Biophys Res Commun. 2008;374(4):609-13.

[3] Faria EC, Ma N, Gazi E, Gardner P, Brown M, Clarke NW, Snook RD. Analyst. 2008;133(11):1498-500. [4] Ketene AN, Schmelz EM, Roberts PC, Agah M. Nanomedicine.2012;8(1):93-102. [5] Pittenger, B., Erina, N., Su, C., 2010. Bruker Application Note #128.

Figures

Figure 1. Comparative Imaging results for the three cell lines considered. First column images (a,b) and cross-section profiles (g,j) correspond to MCF-10A cells. Second column images (b,e) and cross-section profiles (h,k) correspond to MCF-7 cells. Third column images (c,f) and cross-section profiles (i,l) correspond to MDAMB-231 cells. First row images (a,b,c) show topography contrast, while second row images (d,e,f) show apparent Youngâ&#x20AC;&#x2122;s modulus contrast. Image size is 60x60um in all cases. Topography range is 2 um for MCF-10A and 6 um in all other cases. Apparent Youngâ&#x20AC;&#x2122;s modulus images are displayed in a logarithmic color scale, from 2.5 to 250 kPa.

Figure 2. Confocal fluorescence microscopy images of cells from the three lines considered. First column images (a,b) and orthogonal projections (g) correspond to MCF-10A cells. Second column images (b,e) and orthogonal projections (h) correspond to MCF-7 cells. Third column images (c,f) and orthogonal projections (i) correspond to MDA-MB-231cells. First row images (a,b,c) show red-stained F-actin filaments, while second row images (d,e,f) show green-stained microtubules. Image size is 80x80 um in all cases. Both contrast signals are mixed in the orthogonal projections.

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Ultraporous interweaving electrospun microfibers and their cellular response Menglin Chen Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus DK-8000, Denmark

Menglin@inano.au.dk In the field of tissue engineering, integration of micro-porosity, nano-topogaphical features and weattability into one three-dimensional (3D) scaffold remains a challenge.Here we report that a nanoscale immiscible polymer blend solution electrojet can assemble into ultraporous interweaving microfibers. The hierarchical porosity influenced cell infiltration, proliferation and differentiation significantly. Methods The polymer solutions were prepared by dissolving PCL (Mw 70 000â&#x20AC;&#x201C;90 000, Aldrich) and PEO (Mw 900 000, Aldrich) in DCM/DMF (3:2) at room temperature and the homogeneous solutions were used for electrospinning under the following conditions: applied voltage 18 kV, feeding rate 1mL h-1, and distance between the tip of the needle and collector 12 cm. The experiments were carried out at room temperature, and the relative humidity was between 30% and 60%. The morphology, crystallinity, surface chemistry and wettability of these fibers were studied by scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM), X-ray diffraction (XRD), Xray photoelectron spectrometry (XPS), and contact angle (CA) measurements. The NIH3T3 cell viability and proliferation was measured by LDH and MTS assay, cell attachment and infiltration were assessed by confocal microscopy.

between each component reached a threshold and where the electrospinning parameters were delicate controlled. The morphology, crystallinity, surface chemistry and wettabilities were characterized to understand the mechanism of formation. The interplay of the two semicrystalline polymers and the pair of solvents/non-solvents with the electrospinning processing parameters was found to be critical for the formation of the unique structure (Fig 1) [1]. The hydrophilic, hierarchically porous fibers were appilied in culturing fibroblasts and studied the cell infiltration and colonization. Compared to the tight-packed, hydrophobic PCL scaffold, the hydrophilic, micro-porous fibers enhanced the cell infiltration and colonziation significantly. Moreover, the unique nano-topographical environment that may stimulate cells in a drastically different manner from that of traditional solid, smooth electrospun fibers, which holds great potential in reconstructing tissues that require strong contractile forces (Fig 2) [2]. Acknowledgments This work was supported by the Danish Council for Strategic Research, Aarhus University Research Foundation and Carlsberg foundation.

Results Multi-lamellar cylindrical structure was originated from a blend of PCL and PEO in DCM/DMF mixed solution when the ratio

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References [1] Li Y; Rubert, M; Yu Y; Dong, M; Besenbacher, F; Chen, M*. Nanoscale, 2014, 6, 3392-402, highlighted in Global Medical Discovery June 23, 2014, https://globalmedicaldiscovery.com/keyna notechnology-articles/ultraporousinterweaving-electrospun-microfibersfrom-pcl-peo-binary-blendsand-theirinflammatory-responses/ [2] Li, Y; Gregersen, H; Nygaard, J; Cheng, W; Huang, Y; Dong, M; Besenbacher, F; Chen, M*. Nanoscale, 2015, 7, 14989 â&#x20AC;&#x201C; 14995

Figures

Figure 1.

Figure 2.

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Graphene based platforms for Raman sensing Alicia de Andrés1, Félix Jimenez-Villacorta1, Rafael Ramírez-Jíménez2, 1, Leo Álvarez-Fraga1, Esteban Climent-Pascual1, Carlos Prieto1 1

Instituto de Ciencia de Materiales de Madrid, Consejo Superior de Investigaciones Científicas, Madrid, Spain 2 Departamento de Física, Escuela Politécnica Superior, Universidad Carlos III de Madrid, Madrid, Spain

ada@icmm.csic.es The detection of protein biomarkers secreted by tumors at early stages for preventive cancer medicine is of vital relevance but their extremely low concentration in blood and the presence of mixtures of proteins makes difficult their detection. Raman spectroscopy meets the required specificity criterion since the vibrational spectrum of every component of a biological specimen is a specific signature that can be used for its identification. Moreover, compared to magnetic resonance imaging, Raman micro-spectroscopy has the ability of visualizing morphological details in cells and tissues on a much higher spatial resolution. Also, no external markers are required, it has a sub-micron resolution and quite good penetration depth. Nevertheless, Raman spectroscopy is strongly limited by its sensitivity. For the last years, a great effort is underway to increase Raman intensity mainly by enhancing the Raman signal by localized surface plasmon resonances from metal nanoparticles (NPs) [1], but these SERS platforms deals with problems such as the NPs stability, their interaction with the analyte and the adsorption, distribution and arrangement of the probed molecules on the substrate. Another enhancement process is the use of excitation wavelengths in resonant conditions for the sensed molecule, in general using ultra-violet lasers. Recently, the Raman signal of graphene has been shown to increase significantly due to the constructive interference processes in graphene/SiO2/Si [2]. Graphene has several roles to play in optical sensing since it provides a biocompatible surface adequate for many organic and bio materials and also a protection

of the metal NPs increasing the stability of the system [3]. Our aim is the fabrication of different architectures of SERS substrates that combine different enhancement processes of Raman signal in one multilayered hybrid system based on graphene. We will present our approaches to the different Raman enhancement processes. In particular we will show the formation of graphene protected ruthenium ultrathin films with controlled size and shape of the particles with different characteristics of the Ru plasmon resonance adequate for UV-SERS. We have explored the limit of ultrasmall Ag nanoparticles (4 nm) to study their interaction with graphene and the SERS amplification capabilities [4]. We will also demonstrate that the spontaneous formation of Cu/air/graphene bubbles in Cu foils where graphene was deposited by chemical vapor deposition produces an enhancement of the graphene Raman signal up to 70. Moreover, the transfer matrix method used to simulate the multireflection processes predicts amplification factors up to 11000 for graphene/air/aluminum systems [5]. The results of these approaches pave the way to the design and fabrication of extremely sensitive graphene based bio-compatible platforms by combining the different Raman enhancement processes.

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References [1] Xie, W.; Schlücker, S. Rationally Designed Multifunctional Plasmonic Nanostructures for Surface-Enhanced Raman Spectroscopy: A Review. Rep. Prog. Phys. 77 (2014), 116502. [2] Wang, Y. Y.; Ni, Z. H.; Shen, Z. X.; Wang, H. M.; Wu, Y. H.; Wang, Y. Y.; Ni, Z. H.; Shen, Z. X. Interference Enhancement of Raman Signal of Graphene. Appl. Phys. Lett. 92 (2008), 043121. [3] Wang, P.; Liang, O.; Zhang, W.; Schroeder, T.; Xie, Y.-H. Ultra-Sensitive GraphenePlasmonic Hybrid Platform for Label-Free Detection. Adv. Mater. 25(2013), 4918.

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[4] F. Jimenez-Villacorta, E. Climent-Pascual, R. Ramirez-Jimenez, J. Sanchez-Marcos, C. Prieto and A. de Andrés. Graphene – ultrasmall silver nanoparticle interactions and their effect on transport properties and Raman enhancement. (Submitted) [5] R. Ramírez-Jíménez, L. Álvarez-Fraga, F. Jimenez-Villacorta, E. Climent-Pascual, C. Prieto and A. de Andrés. Interference Enhanced Raman Effect in Graphene Bubbles. (Submitted)

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In vitro and in vivo antimetastatic efficacy of a polymer-based paclitaxel conjugate for prostate cancer therapy Yolanda Fernández1, Julie Movellan2, Laia Foradada1, Vanessa Giménez2, Natalia García-Aranda1, Sandra Mancilla1, Ana Armiñán2, Richard M. England2, María J. Vicent2, Simó Schwartz Jr.1, Ibane Abasolo1 Vall d'Hebron University Hospital-UAB and Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN). CIBBIM-Nanomedicine - Functional Validation & Preclinical Studies, Barcelona, Spain 2 Centro de Investigación Príncipe Felipe. Polymer Therapeutics Laboratory, Valencia, Spain 1

yolanda.fernandez.amurgo@vhir.org The design of improved biodegradable polymeric carriers which can enhance drug’s efficacy by controlling the concentration and release of the active principle in the tumors is an ongoing challenge. Paclitaxel (PTX) is a clinically well-established and highly-effective antineoplastic drug used for the treatment of many carcinomas, including prostate. However, the clinical use of PTX is limited by the side effects caused by a poor biodistribtuion of the drug and by the solvent (Cremophor) in wich PTX is dissolved for delivery. Thus, it is hypothesized that the conjugation of PTX to a polymeric carrier would offer an enhancement of PTX clinical benefit in prostate cancer patients by allowing a low-dose clinical regime, a controlled release of the drug and a complete absence of solvent-related toxicities. Looking for high Mw, biodegradable and pHresponsive polymeric carriers [1,2], PTX was conjugated to the side-chains of a pHsusceptible biodegradable Poly(ethylene glycol) polymer yielding: tert-PTXpolyacetal. In vitro efficacy of the synthesised conjugate was tested in prostate cancer cells looking at cell viability to determine the half maximal inhibitory concentration (IC50) values. Further, in vivo tolerability and therapeutic antitumor and antimetastatic efficacy of the conjugate was evaluated in prostate ortothopic tumors using

in vivo bioluminescence optical imaging and histopatology. In vitro, tert-PTX-polyacetal conjugate reduced cell viability of LNCaP prostate tumor cells at the same order of magnitude that the free PTX. In vivo, tert-PTX-polyacetal conjugate was effective in inducing orthotopic LNCaP prostate tumor growth inhibition. Relevantly, tert-PTXpolyacetal also inhibited the disease propagation by significantly reducing the lymphatic, hematologic and coelomic dissemination as confirmed by in vivo and ex vivo BLI imaging and histopathological analysis. Moreover, tert-PTXpolyacetal conjugate significantly reduced the systemic toxicity associated with free PTX. These results indicate that the tert-PTXpolyacetal could be used as a robust drug delivery system for antitumoral and antimetastatic treatments based on PTX.

References [1] Duro-Castano A, Movellan J, Vicent MJ. Biomater Sci. 3(10) (2015)1321-34. [2] Giménez V, James C, Armiñán A, Schweins R, Paul A, Vicent MJ. J Control Release. 2159(2) (2012) 290-301.

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Contribution of Water to Pressure and Cold Denaturation of Proteins G. Franzese Departament de Fisica Fonamentalo, Universitat de Barcelona, Marti i Franques 1, Barcelona, Spain

gfranzese@ub.edu The mechanisms of cold and pressure denaturation of proteins are matter of debate and are commonly understood as due to watermediated interactions. Here, we study several cases of proteins, with or without a unique native state, with or without hydrophilic residues, by means of a coarse-grain protein model in explicit solvent. We show, using Monte Carlo simulations, that taking into account how water at the protein interface changes its hydrogen bond properties and its density fluctuations is enough to predict protein stability regions with elliptic shapes in the temperature-pressure plane, consistent with previous theories. Our results clearly identify the different mechanisms with which water participates to denaturation and open the perspective to develop advanced computational design tools for protein engineering [1-6].

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References [1] M. G. Mazza, K. Stokely, S. E. Pagnotta, F. Bruni, H. E. Stanley, G. Franzese, Proc. Natl. Acad. Sci. 108, 19873 (2011). [2] V. Bianco, S. Iskrov, and G. Franzese, J. Biol. Phys. 38, 27 (2012). [3] G. Franzese and V. Bianco, Food Biophys. 8, 153 (2013). [4] P. Vilaseca, K.A. Dawson, G. Franzese, Soft Matter 9, 6978 (2013). [5] V. Bianco and G. Franzese, Sci. Rep. 4, 4440 (2014). [6] V. Bianco and G. Franzese, Phys. Rev. Lett. 115, 108101 (2015).

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New fluorescent CSC models evidence that targeted nanomedicines improve treatment sensitivity of breast and colon cancer stem cells.Sensitivity of breast and colon cancer stem cells Petra Gener, Guillem Romero Sabat, Diana Fernandes de Sousa Rafael, Núria Bergadà Fort, Yolanda Fernández, Rafael Miñana Prieto, Ibane Abasolo, Mafalda Videira, Simo Schwartz Jr CIBER-bbn, Spain

To be able to study the efficacy of targeted nanomedicines in marginal population of highly aggressive cancer stem cells (CSC), we have developed a novel in vitro fluorescent CSC model that allow us to visualize these cells in heterogeneous population and to monitor CSC biological performance after therapy. In this model tdTomato reporter gene is driven by CSC specific (ALDH1A1) promoter and contrary to other similar models, CSC differentiation and un-differentiation processes are not restrained and longitudinal studies are feasible. We used this model for preclinical validation of poly[(D,Llactide-co-glycolide)-co-PEG] (PLGA-co-PEG) micelles loaded with paclitaxel. Further, active targeting against CD44 and EGFR receptors was validated, in breast and colon cancer cell lines. Accordingly, specific active targeting towards surface receptors enhances the performance of nanomedicines and sensitizes CSC to paclitaxel based chemotherapy.

This work was supported by grants from EuroNanoMed 2009 (NANOSTEM project), PI14/02079 from ISCIII, Spanish Ministry of Economy and Competitiveness. Authors declare no conflict of interest.

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Nanomedicine, from PoC to reality: the importance of an industrial perspective and GMP scale up O. Ibarrola, A. del Pozo, E. Gainza BioPraxis AIE, Hermanos Lumière 5, 01510 Miñano, Spain

oibarrola@praxisph.com Nanotechnology and specially its application to improvement of Human health (Nanomedicine) is expected to be one of the pillars of novel products and processes in the next future, in this case to get innovative therapies. Nanoenabled therapies, together with cell and gene therapies and personalized medicine will pave the way for a revolution in the treatment of several diseases which currently have no treatment or low efficacy ones. There is a common consensus on the potential of nanomedicine to contribute to this global improvement, but this will only be possible if all the nano medicine community is able to foster the translation of the promising results obtained at laboratory scale to real products applicable at clinical settings. This full deployment of nanomedicine must be based on the identification of the existent gaps for translation, at different levels, and on the industrialization of nanomedicines production, taking into account Good Manufacture Practices (GMP) and regulatory aspects. From Biopraxis, the Research and Innovation Unit of the pharmaceutical Group Praxis, we have developed an intensive work to identify and propose solutions for many of those gaps. In the current moment Biopraxis holds the Chairmanship of the Nanotherapeutics Working Group in the European Technology Platform for Nanomedicine, making us a privileged stakeholder to receive inputs from the nanomedicine community and to highlight our

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contributions to nanomedicines.

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translation

In this communication, we aim to share all these lessons learned when trying to move nanomedicine to the next steps. To do this, we have identified and proposed solutions to the different challenges at different levels, and, due to our industrial commitment, with a special focus in GMP up scale of nanomedicines production. Main challenges are the following:  Society: There is a need of higher acknowledgment and support for nanomedicine, avoiding a potential “nanofobia”  Risks: We identify risks at two main levels: safety and environment impact, and propose preventive and corrective actions  Regulation: medicines market is a highly regulated market, and there is a need for the definition of the regulatory requirements for nanomedicines. We set the problem at three different levels: Preclinical development, clinical research and market access  Technology implementation: There is a huge amount of technology offers, which has to be discerned by industry. An open innovation scheme is proposed as a potential solution to this challenge  Industrialization: Biopraxis is specialized scaling up the manufacturing of diferent nanoformulations from milligram-scale laboratory (Fig 1) synthesis up to multigramscale production to generate sufficient

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material for clinical and regulatory assays. We standardise the up-scale production of nanoparticles under GMP (Fig 2) considering the main bottleneck aspects: reproducibility, stability, and non-immunogenicity (sterility and non-pyrogenicity). At the same time, we consider critical aspects of the GMPs design such as: continuous quality control, risk assessment for manufacturing process, specifications for excipients, intermediates and finished products; rooms classification, equipment, supplies (water, heat, stirring, gasesâ&#x20AC;Ś) ď&#x201A;&#x2014; Business models: New paradigms need also innovative approaches to business models. Nanomedicines still present some weakness on this sense, i.e. detailed cost evaluations. As it has been shown, current landscape for nanomedicines is full of potential, but presents a series of challenges, which need to be solved, and in many cases, only the commitment and driving action of the industry can bridge this potential valley of death. At Biopraxis we are managing several projects for the development of nanomedicines, and we want to share our experiences and potential answers to contribute to nano-enabled therapies in the next future, in a context with the view from the Industry is sometimes difficult to find.

Figures

Figure 1.

Figure 2.

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Cork filter with silver nanoparticles for Lab-Scale water disinfection Lina Garcia1, Ángel Gallegos2, Rosanna Margalef2, Lorena Aguilar2, Marc Vives2, Antonio Francesko3, Petya Petkova3, Tzanko Tzanov3, Juan Casas1, Jordi Morató2 1

Grupo de Investigación en Ciencia e Ingeniería en Sistemas Ambientales (GCISA), Civil Engineering Faculty, Universidad del Cauca, Popayán, Colombia 2 Laboratori de Microbiologia Sanitaria i Mediambiental (MSMLab), UPC-BarcelonaTech, Spain 3 Grup de Biotecnologia Molecular i Industrial (GBMI), Department of Chemical Engineering, UPC-BarcelonaTech, Spain

Nowadays there is a major concern in the public health field: waterborne diseases, specifically those caused by the presence of infectious agents in tap water. According to the World Health Organization (WHO) one tenth of the global disease burden could be prevented by improving the water supply, sanitation, hygiene and management of water resources [1], so taking measures to improve the quality of the consumption water is essential to guarantee the harmlessness of water for human beings. One of the most recurrent diseases caused by waterborne pathogens is diarrhoea. It occurs world-wide and causes 4% of all deaths and 5% of health loss to disability. It is most commonly caused by gastrointestinal infections which kill around 2.2 million people globally each year, mostly children under five years old living in developing countries [2]. Given this fact, the United Nations (UN) has established amongst its purposes on sustainable development to meet before 2030, the Global Water Goal, which aims to reduce by half the population that have no access to potable water in their homes [3]. Focusing on the Global Water Goal, it is widely known that to obtain potable water it is necessary to perform a series of treatments, including a disinfection step. It is performed both in surface water and groundwater, which are exposed to faecal contamination in order to eliminate the possible presence of pathogenic microorganisms. This operation is frequently accomplished by adding chemicals like

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chlorine, but has numerous disadvantages as the limited efficiency against certain microorganisms, specifically protozoa and viruses or the generation of by-products because of the chemical reactions between the chlorine and the organic material present in the water that has to be treated [4]. Despite the chlorine addition is much more used than other ways of treatment because it is a cheap and an easy way which provides high efficiencies in attaining safe water conditions, it is necessary to investigate alternative approaches that would permit to achieve a good disinfection while avoiding the mentioned drawbacks. Furthermore, during the last years and due to the climate change, the concerns about waterborne diseases have worsened. An increase both in the temperatures and in rainfall events that may lead to floods can be translated to an excessive microbial proliferation in surface waters and groundwater, so it is crucial to find potential corrective strategies to solve any possible emergency situation [5]. Given the fact that the use of silver in many fields has been demonstrated to possess antimicrobial properties and it has promising applications [6], the MSMLab at the Universitat Politècnica de Catalunya-BarcelonaTech (UPC) has been investigating the possibility of using cork with silver nanoparticles attached to its surface as a filter media for water disinfection.

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Currently there is little information about the use of cork as a filtration media, so we are facing an innovative technology that fulfils the circular economy principle regarding that cork is a major waste product in Catalonia (wine producing region). The adsorption of the silver nanoparticles on the cork surface is performed following the methodology described by Francesko et al. (2015). The procedure consists on an enzymatically catalysed process between a silver nanoparticles solution and the cork. The laboratory scale filter cartridges filled with treated cork were used to test their antimicrobial efficiency against Escherichia coli (Figure 1). The experiments were carried out in two phases: the first one using a continuous flow and the second one operating under a batch mode. In both cases the removal efficiency exceeded 99,9%. The use of cork filter media with attached silver nanoparticles, is an innovative and appropriate technology for water disinfection that present many advantages, like easy operation and adaptability with other treatments and different environmental contexts, as well as an easy application and potential use in small communities.

[3] United Nations (UN). 2014. A Post-2015 Global Goal for Water: Synthesis of key findings and recommendations from UNWater. Website of the United Nations [consulted on August 26, 2015] http://www.unwater.org/fileadmin/user_up load/unwater_new/docs/Topics/UNWater_paper_on_a_Post2015_Global_Goal_for_Water.pdf [4] World Health Organization (WHO). 2006. Guidelines for Drinking Water Quality. Website of the World Health Organization [consulted on August 26, 2015]. http://www.who.int/es/ [5] HUNTER, P.R. 2003. Climate change and waterborne and vector-borne disease. Journal of Applied Microbiology, 94, 37–46. [6] FRANCESKO, A. et al. 2015. Enzymatic functionalization of cork surface with antimicrobial hybrid biopolymer/silver nanoparticles. ACS Applied Material Interfaces, 18, 9792-9.

Figures

References [1] Prüss-Üstün A et Al. 2008 Safer water, better health: costs, benefits and sustainability of interventions to protect and promote health. World Health Organization, Geneva, 2008. [2] World Health Organization (WHO). 2014. Water Sanitation Health, Water-related Diseases, Diarrhoea. Website of the World Health Organization [consulted on October 26, 2015]. http://www.who.int/water_sanitation_heal th/diseases/diarrhoea/en/.

Figure 1. The Inoculated Water with Escherichia coli is pumped by a peristaltic pump (Pp) to the Filtering Cartridge (FC), fill it with cork treated with silver nanoparticles.

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Nanomechanics of reelin autism related protein - AFM and SMD studies K. Mikulska-Rumińska, J. Strzelecki, W. Nowak Institute of Physics, Faculty of Physics, Astronomy and Informatics, Nicolaus Copernicus University, Torun, Poland

karolamik@fizyka.umk.pl Molecules in the nervous system are the most crucial elements in every living organism. They control life functions performing a lot of complex operations. Defects of these molecules may result in many diseases such as autism or schizophrenia [1]. Among proteins with links to diseases are contactin, neuroligin and reelin [2]. Reelin is a large modular protein (Fig. 1) containing 8 characteristic BNR-EGF-BNR fragments. It is suspected to be associated with autism, schizophrenia, bipolar disorder depression and Alzheimer's disease. In the adult brain this protein is involved in proper plasticity, learning and memory [3]. In order to investigate the nanomechanical properties of reelin and other autism related proteins single molecule force spectroscopy (SMFS AFM) and steered molecular dynamics simulations (SMD) were used. SMFS measurements were performed using non-commercial AFM in a constant speed mode. As an auxiliary tool in understanding the molecular processes induced by mechanical stretch in SMFS measurements we applied all-

atom and coarse-grained molecular dynamics simulations. All simulations were performed using the NAMD and CHARMM codes. New data on reelin mechanical unfolding and their interpretation will be presented. We believe that combination of this two complementary approaches will help to elucidate a role of residues critical for proteins’ stability and possible abnormalities induced by mechanical manipulations at atomic level. Supported by National Science Centre (Poland) 951-F (2012/05/N/ST3/03178).

References [1] S. M., Klauck, Eur. J. Hum. Gen. 14 (2006) 714-720. [2] J.L. Silverman, M. Yang, C. Lord, J.N. Crawley Nat. Rev. Neurosci. (2010) 11 490– 502. [3] S. Fatemi Mol. Psychiatry 10 (2004) 251-257.

Figures

Figure 1. Cristal structure of reelin protein showing two BNR-EGF-BNR repeat fragments.

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november 18-20, 2015 - Barcelona (Spain)

Multifunctional Nanoplatform for hyperthermia. Heating and thermometry in a single nanoparticle Angel Millán1, Rafael Piñol1, Carlos D. S. Brites2, Rodney Bustamante1, Abelardo Martínez3, Nuno J. O. Silva2, José L. Murillo1, Rafael Cases1, Julian Carrey4, Carlos Estepa1, Cecilia Sosa5, Gustavo Lou1, Diego De Miguel Samaniego6, Lilianne Beola7, Amalia Ruiz7, Jesús M de La Fuente1, Luis Martinez-Lostao6, Fernando Palacio1, Belinda Sánchez8, Luis D. Carlos2 ICMA, CSIC–Universidad de Zaragoza, 50009 Zaragoza (Spain) Departamento de Física and CICECO, Universidade de Aveiro, 3810–193 Aveiro (Portugal) 3 Departamento de Electrónica de Potencia. I3A. Universidad de Zaragoza 50018 Zaragoza (Spain) 4 Université de Toulouse, INSA, UPS, CNRS (UMR 5215), F-31077, Toulouse (France) 5 Departamento de Toxicología, Universidad de Zaragoza, 50013 Zaragoza (Spain) 6 Departamento de Bioquímica, Biología Molecular y Celular, Universidad de Zaragoza, Zaragoza (Spain) 7 Centro de Estudios Avanzados de Cuba, La Habana (Cuba) 8 Centro de Inmunología Molecular 1

amillan@unizar.es Nanoparticles can be made large enough to carry several active molecules with different functionalities but still small enough to cross biological barriers. We can call them then multifunctional nanoplatforms. There are several chemical strategies to build such platforms including encapsulation in liposomes, porous nanogeles, binding to a nanoparticle surface or attached to side chains of branched polymers. We describe an alternative strategy based on a platform with anchoring sites and a set of interchangeable pieces that fit on them (Fig. 1). Several tools have been incorporated to this nanoplatform, so far, including: imaging tags (fluorescent, magnetic resonance or radioactive), therapeutical drugs, magnetic nanoparticles, and biological vectors (antibodies and polypeptides). Another interesting tool that has also been incorporated is a molecular thermometer. Magnetic induction heating of nanoparticles can be a powerful non-invasive technique for hyperthermia therapy of cancer and other

diseases. To make it effective at local level it requires an adequate monitoring of the nanoheaters local temperature. Here we present a single nanoparticle integrating magnetic heating [1] and luminescent thermometry [2]. The temperature readout is optical and the thermometric probes are dualemissive Eu3+/Tb3+ lanthanide complexes. These complexes are encapsulated in the hydrophobic inner part of the nanoparticle copolymer coating, around the iron oxide magnetic core. The resulting heater/thermometer nanoplatform shows an outstanding performance in terms of: low thermometer heat capacitance (0.021·K−1) and heater/thermometer resistance (1 K·W−1); high temperature sensitivity (5.8%⋅K−1 at 296 K) and uncertainty (0.5 K), physiological working temperature range (295−315 K), readout reproducibility (>99.5%), and fast time response (0.250 s). Experiments of time-resolved thermometry under an AC magnetic field reveal the existence of an unexpected temperature gradient between nanoheaters and surrounding media for relatively long time intervals (t > 10 s) and relatively low heat powers (10-16 W/heater). A proof of concept of temperature mapping has

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been realized on cells that were incubated with the nanoparticles [3]. The fluorescence microscopy in two different wavelengths simultaneously after beam splitting permits the mapping of the intracellular local temperature using the pixel-by-pixel ratio of the Eu3+/Tb3+ intensities (Figs. 2&3). The heater/thermometer single nanoplatform reported here shows great potential for impact on the design of hyperthermia therapies based on localized manipulation of heat flows and short application times. In this way, local energy supply which is not immediately dissipated at the surrounding media could be enough to induce irreversible intracellular damage in tumor cells within a short time period, while maintaining the neighboring tissue temperature unchanged [4] Together with an adequate vectorization of the particles, unprecedented specificity would be achieved. The use of the

system presented here can help to settle these questions and to give a fair account of the real potential of local hyperthermia. In a more extended vision, accurately controlled local heating and precise temperature determination in the cellular media will enable thermal conductivity studies in cellular organelles and across membranes, as well as detailed studies in cell physiology related to thermal processes.

References [1] C. D. S. Brites, et al. Adv. Mater, 22 (2010) 4499. [2] R. Bustamante et al. Phys. Rev. B 88 (2013) 184406. [3] R. Pinol et al. ACS Nano 9 (2015) 3134. [4] A. Schroeder, et al. Nat. Rev. Cancer 12 (2012) 39.

Figures

Figure 1. Scheme of the Multifunctional Nanoplatform and electron microscopy of the thermometric-heatingnanoplatform.

Figure 2. Imaging of Tb3+ (A&C) and Eu3+ (B&D) emissions from cell-internalized thermometricheating-nanoplatform. Scale bars are 40 Îźm.

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Figure 3. Pseudocolour maps of spot 1 in Fig 2C&2D illustrating the co-localization of the Tb3+ (A) and Eu3+ (B) emissions, temperature map (C) computed from this emissions at every pixel, and (D) histogram of the temperature distribution near the cell nucleus. Scale bars correspond to 10 Îźm.

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november 18-20, 2015 - Barcelona (Spain)

Biocatalytic modulation of semiconductor quantum dots: How to apply enzymatic growth and etching of CdS quantum dots to biosensing Valery Pavlov, Ruta Grinyte, Javier Barroso, Laura Saa CIC BiomaGUNE, Paseo Miramon 182, San Sebastian, Spain

vpavlov@cicbiomagune.es The traditional fluorogenic enzymatic assays broadly employed in bioanalysis are based on the biocatalytic cleavage of bonds between presynthesized organic fluorescent molecules or fluorescent semiconductor nanoparticles (SNPs), so called quantum dots (QDs) and quenching moieties [1]. Usually, they suffer from insufficient quenching of fluorophores by quenchers and nonspecific adsorption on surfaces resulting in high background signals [2]. We pioneered enzymatic assays in which formation of CdS QDs in situ is modulated by biocatalytic processes. The first group of assays employs enzymatic production of S2- ions leading to formation of CdS QDs in the presence of Cd2+ ions (Cd2+ + S2- = CdS) [3]. The second group of QDs-generating fluorogenic enzymatic assays developed by us relies on modulating the growth of CdS QDs with the products of biocatalytic transformation [4]. Enzymatically generated CdS QDs show homogeneous size distribution with the medium diameter of 2 nm [3,4]. The size of the resulting SNPs is controlled by the nature of capping agents such as citrate, orthophosphate, L-cystein, glutathione, etc. The advantages of biocatalytic modulation of QDs over employment of traditional organic chromogenic and fluorogenic enzymatic substrates, include lower background signals, higher quantum yield, reduced photo-bleaching and lower costs.

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We demonstrated the use of the peroxidasemimicking DNAzyme (peroxidase-DNAzyme) as general and inexpensive platform for development of fluorogenic assays that do not require organic fluorophores [5]. The system is based on the affinity interaction between the peroxidase-DNAzyme bearing molecular beacon and the analyte (DNA or low-molecular weight molecule), which changes the folding of the hairpin structure and consequently the activity of peroxidase-DNAzyme. Hence, in the presence of the analyte the peroxidase-DNAzyme structure is disrupted and does not catalyze the aerobic oxidation of L-cysteine to cystine. Thus, L-cystein is not removed from the system and the fluorescence of the assay increases due to the in situ formation of fluorescent CdS QDs. The capability of the system as a platform for fluorogenic assays was demonstrated through designing model geno- and aptasensor for the detection of a tumor marker DNA (Figure 1) and a low-molecular weight analyte, adenosine 5Â´triphosphate (ATP), respectively. We developed an innovative photoelectrochemical process (PEC) based on graphite electrode modified with electroactive polyvinylpyridine bearing osmium complex (Osâ&#x20AC;&#x201C; PVP). The system relies on the in situ enzymatic generation of CdS QDs. Alkaline phosphatase (ALP) catalyzes the hydrolysis of sodium thiophosphate (TP) to hydrogen sulfide (H2S), which in the presence of Cd2+ ions yields CdS

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SNPs. Irradiation of SNPs with the standard laboratory UV-illuminator (wavelength of 365 nm) results in photooxidation of 1-thioglycerol (TG) mediated by Osâ&#x20AC;&#x201C;PVP complex on the surface of graphite electrode at applied potential of 0.31 V vs.Ag/AgCl. (Figure 2) A novel immunoassay based on specific enzyme linked immunosorbent assay (ELISA) combined with the PEC methodology was developed. Having selected the affinity interaction between bovine serum albumin (BSA) with anti-BSA antibody (AB) as amodel system, we built the PEC immunoassay for AB. The new assay displays a linear range upto 20 ng mL-1 and a detection limit of 2 ng mL-1 (S/N = 3) which is lower 5 times that of the traditional chromogenic ELISA test employing p-nitro-phenylphosphate. We observed for the first time enzymatic etching of CdS QDs. Fluorescence of semiconductor CdS QDs is modulated irreversibly by the enzymatic reaction catalyzed by horseradish peroxidase (HRP). We detected blue-shifts of corresponding fluorescence peak for CdS QDs and decrease in the intensity of the fluorescence signal. During the study of this phenomenon it was found out that CdS QDs are enzymatically oxidized by hydrogen peroxide resulting in formation of sulfate ions and etching of the initial SNPs (confirmed by electron microscopy) according to Figure 3. Formation of sulfate ions was confirmed by two independent analytical methods. This oxidation reaction occurs also when CdS QDs are adsorbed on the surface of polyvinyl chloride microspheres. This study indicates that CdS QDs act as a substrate for HRP. In order to characterize etching of QDs different techniques were employed e.g. fluorescence technique, transmission electron microscopy and wide field fluorescence microscopy. In order to validate our assay we

applied it to detection of hydrogen peroxide in tap and rain water. It should be noted that the novelty of the reported sensing strategy lies on the use of inexpensive compounds for the development of fluorimetric bioanalytical systems. In comparison with other reported fluorogenic assays based on pre-synthesized QDs modified with recognition elements, our assays require neither any synthetic procedures for chemical modification of QDs nor any organic fluorogenic enzymatic substrates.

References [1] I. L. Medintz, T. Pons, S. A. Trammell, A. F. Grimes, D. S. English, J. B. Blanco-Canosa, P. E. Dawson, Hedi Mattoussi, J. Am. Chem. Soc., 130, (2008), 16745; K. Boeneman, B. C. Mei, A. M. Dennis, G. Bao, J. R. Deschamps, H. Mattoussi, I. L. Medintz, J. Am. Chem. Soc., 131, (2009), 3828. [2] R. Freeman, I. Willner , Nano Lett., 9, (2009), 322. [3] L. Saa and V. Pavlov, Small, 8, (2012) 3449; L. Saa, J. Mato V. Pavlov, Anal. Chem., 84, (2012), 8961. [4] G. Garai-Ibabe, M. MoĚ&#x2C6;ller, V. Pavlov, Anal. Chem., 84, (2012), 8033; N. Malashikhina, G. Garai-Ibabe, V. Pavlov, Anal. Chem., 85, (2013), 6866. [5] G. Garai-Ibabe, L. Saa, V. Pavlov, Anal. Chem., 86, (2014), 10059. [6] J. Barroso, L. Saa, R. Grinyte, V. Pavlov, Biosens. Bioelectron., 77, (2016), 323. [7] R. Grinyte, L. Saa, G. Garai-Ibabe, V. Pavlov, Chem. Commun., (2015) DOI: 10.1039/C5CC05613F

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Figures

Figure 1. DNA detection through peroxidase-DNAzyme modulated growth of CdS QDs in situ.

Figure 2. Photoelectrochemical immunosensors based on enzymatic formation of CdS QDs by alkaline phosphatase (ALP) and detection of photocurrent.

Figure 3. Biocatalytic etching of CdS NPs by horseradish peroxidase (HRP).

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Molecular Wires for the Improvement of DNA Electrochemical Sensors Judit P. Valero1, Sam Bacena Dulay1, Wilmer A. Pardo1, Mònica Mir1,2, Josep Samitier1,2,3 Nanobioengineering Laboratory, Institute for Bioengineering of Catalonia (IBEC), Barcelona, Spain Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Zaragoza, Spain 3 Department of Electronics, Barcelona University (UB), Barcelona, Spain 1

juditp.valero@gmail.com In recent years, there have been intensive research efforts in the field of DNA electrochemical biosensors seeking designs to provide better analytical characteristics in terms of sensitivity, selectivity, reliability, ease of fabrication and use, and lower limits of detection. The performance of DNA biosensors is dependent on the overall efficiency on the principles of hybridisation between surface immobilised nucleic acids and target complementary sequences and it is well established that the surface chemistry is a critical factor [1] including electrochemical detection of electroactive species into electrode’s surfaces making of central interest, especially in the field of biosensing for forensic or clinical analysis from detecting, diagnosing and treatment of infectious and genetic diseases [2]. Conventionally, there are two main methods applied for DNA detection: a) Optical and/or b) electrical methods. One of the optical methods that are frequently used is the fluorescence, which has been widely used due to its well established sensitivity, but its optical transduction is costly [3]. Another technique that uses optical methods is the surface plasmon resonance spectroscopy (SPR) that can provide excellent signal-to-noise ratios and at the same time offer a dynamic range of detection. Additionally, surface coverage area for any immobilised molecules can be calculated with this technique providing

information about monolayer formation, adsorption/desorption kinetics and interaction events [4]. Albeit this technique is highly sensitive and provides much information, it is costly and needs skilled personnel to run the instrument. Electrochemical DNA biosensors have more advantages over other transduction methods obtaining a direct electronic detection of the signal, with no further transduction required, reducing the cost and the size of the devices [5]. Although, the redox enzyme labels are the ones that provide higher signal amplification, the enzymes are unstable under certain temperature and cannot be directly used in PCR primers and their use requires the addition of the enzyme substrate and a mediator. Ferrocene based molecules have been widely used as redox molecule reporters in commercial sensors [6, 7] due to its well-established chemistry and being attractive as reagentless electrochemical detection. In addition, ferrocene is a neutral compounds stable in the presence of water and air making it very interesting for biological applications. It has high reversible redox systems that can be switched from ferrocene to ferrocinium cation at low potentials [8]. However, low current signal has been reported due to the lone electron obtained from ferrocene during redox reaction that has to be transferred to the electrode surface in the most efficient way to allow detection.

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In this paper, the improvement of the platform with molecular wires has been considered in order to enhance the transduction of the ferrocene detection. In this case, the electrode surface for some electrodes was modified with 6-Mercaptohexanol as a blocking agent and S[4-[2-[4-(2-Phenylethynyl)phenyl]ethynyl]phenyl] thiol acting as the molecular wire helps in the transport of ferrocene electron to the electrode by means of its π orbitals in the aromatics. Different molecules have been studied as bridges to favour the electron transfer being the most studied ones oligo(phenyleneethynylenes) (OPEs) and oligo(phenylenevinylenes) (OPVs) [7,9]. However, their application in DNA biosensors is still in its early stages and fewer examples are reported [5,6]. Therefore, the other aim of this work was to improve the electron transfer rate constant by means of the molecular wire from the ferrocene-labelled complementary DNA strands hybridized with the co-immobilized complementary DNA sequences on the working electrode.

References [1] Gooding, J.J., Lai, L. M. H., and Goon, I. Y. Nanostructured electrodes with unique properties for biological and other applications. Chem. Modif. electrodes 11, 1– 56 (2009). [2] Wang, J., Liu, G. & Merkoi, A. Electrochemical Coding Technology for Simultaneous Detection of Multiple DNA Targets Electrochemical Coding

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[3]

[4]

[5]

[6]

[7]

[8]

[9]

Technology for Simultaneous Detection of Multiple DNA Targets. Society 3214–3215 (2003). doi:10.1021/ja029668z Swager, T. M. The Molecular Wire Approach to Sensory Signal Amplification. Acc. Chem. Res. 31, 201–207 (1998). Boozer, C., Kim, G., Cong, S., Guan, H. & Londergan, T. Looking towards label-free biomolecular interaction analysis in a highthroughput format: a review of new surface plasmon resonance technologies. Curr. Opin. Biotechnol. 17, 400–5 (2006). Drummond, T. G., Hill, M. G. & Barton, J. K. Electrochemical DNA sensors. Nat. Biotechnol. 21, 1192–9 (2003). Umek, R. M. et al. Electronic detection of nucleic acids: a versatile platform for molecular diagnostics. J. Mol. Diagn. (2001). doi:10.1016/S1525-1578(10)60655-1 Eckermann, A. L., Feld, D. J., Shaw, J. A. & Meade, T. J. Electrochemistry of redoxactive self-assembled monolayers. Coordination Chemistry Reviews (2010). doi:10.1016/j.ccr.2009.12.023 Seiwert, B. & Karst, U. Ferrocene-based derivatization in analytical chemistry. Analytical and Bioanalytical Chemistry (2008). doi:10.1007/s00216-007-1639-7 Ulgut, B. & Abruña, H. D. Electron transfer through molecules and assemblies at electrode surfaces. Chemical Reviews (2008). doi:10.1021/cr068060w

november 18-20, 2015 - Barcelona (Spain)

Bio-identity of Albumin-Iron Oxide Nanoparticles Anna Roig, Siming Yu, Laura Gonzalez-Moragas, Maria Milla, Jordi Faraudo, Anna Laromaine Nanoparticles and Nanocomposites Group, Institut de CiĂ¨ncia de Materials de Barcelona, (ICMAB-CSIC), Spain

roig@icmab.es Superparamagnetic iron oxide nanoparticles (SPIONs) are already demonstrating their huge potential in nanomedicine as MRI contrast agents, in hyperthermia treatments, for drug delivery and targeting therapies or in biosensing. However, the stability of SPIONs in complex biological environments remains a challenge. Simple biocompatible surface coatings to stabilize them and enhance their therapeutic effect are currently being investigated. Albumin is the most abundant protein in serum with key physiological functions and can be also found in the formulation of some nanomedical products such as AbraxaneÂŽ (paclitaxel-albumin nanoparticles). I will show that bovine serum albumin (BSA) largely improves the colloidal stability of SPIONs in biological media [1]. BSA forms a well-defined protein corona on the nanoparticle modifying its initial surface chemistry and providing a new but distinct bio-identity to the nanoparticle when exposed to biological media, cells and organisms. Binding affinity, conformation changes of the protein and its impact on cytotoxicity, cellular uptake, distribution and fate of nanoparticles will be reported. Experimental evidences will be complemented with molecular simulations. Finally, the albumin-iron oxide biocomposite assessment in a simple animal model, C. elegans, will be presented [2] (Figure 1). In particular, based on its transparency, short life cycle, differentiated anatomy and ease of cultivation, C. elegans is a suited model organism for in vivo NP evaluation within the synthetic laboratory [3]. Interestingly, results confirm in vitro observations regarding enhanced stability of BSA-coated SPIONs in biological environment

and reduced interaction with cells. Furthermore, all findings indicate the protective effects of the protein both to the nanoparticles and to the worms especially at high concentrations.

References [1] Enhanced stability of superparamagnetic iron oxide nanoparticles in biological media using a pH adjusted and BSA adsorption protocol, S. Yu, A. Laromaine, A. Roig, Journal of Nanoparticle Research 16 (2014) 2484 DOI: 10.1007/s11051-014-2484-1. [2] Protective effects of Bovine Serum Albumin on superparamagnetic iron oxide nanoparticles evaluated in the nematode Caenorhabditis elegans, L. Gonzalez-Moragas, S.Yu, E. Carenza, A. Laromaine, A. Roig, ACS Biomaterials Science and Engineering DOI10.1021/acsbiomaterials.5b00253. [3] C-elegans as an in vivo toolkit for nanoparticles assessment, L. Gonzalez-Moragas, A. Roig and A. Laromaine, Advances in Colloid and Interface Science 219 (2015) 10-26,DOI10.1016/j.cis.2015.02.001.

Figures

Figure 1. Prussian Blue stained C. elegans, SPIONs appear blue in the intestinal track.

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Superparamagnetic iron oxide nanoparticle uptake alters M2 macrophage phenotype José M Rojas1, Laura Sanz-Ortega1, Vladimir Mulens-Arias1, Lucía Gutiérrez2, Sonia Perez-Yagüe1, Domingo F Barber1 1

Department of Immunology and Oncology and Nanobiomedicine Initiative, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Cientificas (CNB-CSIC), Madrid, Spain 2 Department of Biomaterials and Bioinspired Materials, Instituto de Ciencias Materiales de Madrid/CSIC (ICMM-CSIC), Madrid, Spain

jmrojas@cnb.csic.es Superparamagnetic iron oxide nanoparticles (SPIONs) have shown promise as contrast agent and nanocarriers for drug delivery [1]. Their administration is well-tolerated and long-term in vivo biodistribution studies have shown that they can be transformed to non-superparamagnetic iron forms and eliminated with no signs of toxicity [2]. SPION inoculation will nonetheless result in nanoparticle exposition to cells of the mononuclear phagocyte system like macrophages. SPION treatment has been linked to a pro-inflammatory switch in macrophages [3]. Although this could be advantageous in some applications, it could also prove detrimental to M2 macrophages involved in inflammation resolution, immune modulation and angiogenesis. Since SPION impact on M2 macrophages has not been well studied, we synthesised and characterised SPIONs coated with dimercaptosuccinic acid, aminopropyl silane or aminodextran, which offered a range of charge and hydrodynamic diameter for comparison. We evaluated the effects of these nanoparticles in two M2 macrophage models: murine primary IL4-activated bone marrow-derived macrophages and human M2-like differentiated THP-1 cells. All SPIONs were internalised and no cell toxicity was observed. SPION treatment produced reactive oxygen species and activated the extracellular signal-regulated kinase and AKT pathways. After 24 hour SPION incubation, M2 macrophages switched their iron metabolism towards an ironreplete status, as shown by decreased expression

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of iron-importing genes and reduced surface expression of transferrin receptor. SPION treatment in both M2 macrophage models promoted IL-10 production, but no specific IL-12 production was detected, indicating that these nanoparticles did not induce an M2 to M1 activation switch. Surface expression of activation markers showed little change after treatment. SPION impacted on macrophage migration; chemotactic migration was impaired in transwell assays whereas protease-dependent invasion was promoted. These data confirm SPION safety, although they also suggest that exposition to these nanosystems could bias some macrophages responses (Figure 1). These results highlight the need to evaluate the interactions between SPIONs and cells to take full advantage of the intrinsic properties of these nanoparticles in biological systems. Modification of SPION surface chemistry could prove a useful tool in immunotherapy to offer adjuvancy or immunomodulation during treatment.

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References [1] Mejias, R., Perez-Yague, S., Gutierrez, L., Cabrera, L. I., Spada, R., Acedo, P., Serna, C. J., Lazaro, F. J., Villanueva, A., Morales Mdel, P., Barber, D. F. (2011) Dimercaptosuccinic acid-coated magnetite nanoparticles for magnetically guided in vivo delivery of interferon gamma for cancer immunotherapy. Biomaterials 32, 2938-52. [2] Mejias, R., Gutierrez, L., Salas, G., PerezYague, S., Zotes, T. M., Lazaro, F. J., Morales, M. P., Barber, D. F. (2013) Long term biotransformation and toxicity of dimercaptosuccinic acid-coated magnetic nanoparticles support their use in biomedical applications. J Control Release 171, 225-33.

[3] Mulens-Arias, V., Rojas, J. M., Perez-Yague, S., Morales, M. P., Barber, D. F. (2015) Polyethylenimine-coated SPIONs trigger macrophage activation through TLR-4 signaling and ROS production and modulate podosome dynamics. Biomaterials 52, 494-506.

Figures

Figure 1. SPION effects on M2 macrophages

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Light driven Micromotors for waste water remediation Juliane Simmchen1,2, Alejandro Baeza3,4, Albert Miguel-Lopez1, Morgan Stanton1, Maria Vallet-Regi3,4, Daniel Ruiz2, and Samuel Sanchez1,5,6 Max-Planck-Institut fĂźr Intelligente Systeme, Heisenbergstr. 3, D-70569 Stuttgart, Germany CSIC-Consejo Superior de Investigaciones Cientificas, ICN2 Building Campus UAB, 08193 Bellaterra (Barcelona), Spain 3 Dep. de QuĂmica InorgĂĄnica y BioinorgĂĄnica, UCM, Plaza RamĂłn y Cajal s/n, Madrid, Spain 4 Centro de InvestigaciĂłn BiomĂŠdica en Red de BioingenierĂa, Biomateriales y Nanomedicina (CIBER-BBN), Spain 5 InstituciĂł Catalana de Recerca i Estudis Avancats (ICREA), Pg. LluĂs Companys 23, 08010, Barcelona, Spain 6 Institut de Bioenginyeria de Catalunya (IBEC), Baldiri I Reixac 10-12, 08028 Barcelona, Spain 1

simmchen@is.mpg.de In the field of micromotors many efforts are taken to find a substitute for peroxide as fuel. Most approaches turn towards other high energy chemicals as hydrazine etc. but here we want to introduce an energy source that is widely used in nature: light [1]. Light is an ideal source of energy and some materials as AgCl are able to harvest this energy directly by undergoing chemical changes. In case of silver chloride one observed process after light exposure is surface modification. Here we present endeavours to use those processes to create motion at the microscale and apply them for waste water remediation. The underlying mechanism is supposedly the gradient creation by decomposition of silver chloride in light đ?&#x2018;&#x2C6;đ?&#x2018;&#x2030;â&#x2C6;&#x2019;đ?&#x2018;&#x2122;đ?&#x2018;&#x2013;đ?&#x2018;&#x201D;â&#x201E;&#x17D;đ?&#x2018;Ą

4đ??´đ?&#x2018;&#x201D;đ??śđ?&#x2018;&#x2122; + 2đ??ť2 đ?&#x2018;&#x201A; â&#x2020;&#x2019;

Here we describe the synthesis of new micromotors based on novel shapes of AgCl particles which present interesting properties beyond the well-known tube-shaped or spherical motors. Their motion is fuelled by the light triggered degradation of AgCl, so that no toxic chemicals are required. The fabricated microstar shaped AgCl micromotors exhibit three different motion modes in the presence of UV such as translational, rocking and rotational motion, which were quantified. We also envision future applications based on the intrinsic properties of AgCl, such as catalytic behavior for organic molecules which can be used to degrade organic pollutants in waste water. An additional benefit of having AgCl as base material are the bacteriostatic effects due to ion release.

4đ??´đ?&#x2018;&#x201D; + 4đ??ť + + 4đ??śđ?&#x2018;&#x2122; â&#x2C6;&#x2019; + đ?&#x2018;&#x201A;2

decomposes silver chloride

In 2009 Ibele et al. found that AgCl particles with asymmetric shape move autonomously in deionized (DI) water when exposed to UV light. The authors attribute this movement to asymmetric photodecomposition of the particles that creates a localized electrolyte gradient around the particle that results in selfdiffusiophoresis [2] and study their UV light induced collective motion [3-5].

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References [1] Guix, M.; Mayorga-Martinez, C. C.; Merkoci, A. Nano/Micromotors in (Bio)chemical Science Applications. Chem. Rev. 2014, 114, 6285-6322. [2] Ibele, M.; Mallouk, T. E.; Sen, A. Schooling Behavior of Light-Powered Autonomous Micromotors in Water. Angewandte Chemie International Edition 2009, 48, 3308-3312. [3] Ibele, M. E.; Lammert, P. E.; Crespi, V. H.; Sen, A. Emergent, Collective Oscillations of Self-Mobile Particles and Patterned Surfaces under Redox Conditions. ACS Nano 2010, 4, 4845-4851. [4] Duan, W.; Ibele, M.; Liu, R.; Sen, A. Motion analysis of light-powered autonomous silver chloride nanomotors. European Physical Journal E 2012, 35. [5] Ibele, M.; Mallouk, T. E.; Sen, A. Schooling Behavior of Light-Powered Autonomous Micromotors in Water. Angew. Chem., Int. Ed. 2009, 48, 3308-3312.

Figures

Figure 1. AgCl microstars

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Affinity-targeted tumor penetrating polymersomes Lorena Sim贸n Gracia1, Hedi Hunt1, Pablo Scodeller1, Gary B. Braun2, Anne-Mari A. Willmore1, Jens Gaitzsch3, Ramana Kotamraju2, Giuseppe Battaglia4, Erkki Ruoslahti2,5, Tambet Teesalu1,2 Laboratory of Cancer Biology, Institute of Biomedicine, Centre of Excellence for Translational Medicine, University of Tartu, Tartu, Estonia 2 Cancer Research Center, Sanford-Burnham-Prebys Medical Discovery Institute, California, USA 3 Department of Chemistry, University of Basel, Basel, Switzerland 4 Department of Chemistry, University College London, Gower St. WC1E 6BT London, UK 5 Center for Nanomedicine and Department of Cell, Molecular and Developmental Biology, University of California, USA 1

lorenasimongracia@gmail.com Tumor-homing peptides can be used to deliver drugs, imaging agents, and nanoparticles into tumors. Tissue-penetrating CendR peptides specifically recognize the endothelium of tumor vessels, extravasate, and penetrate deep into the tumor parenchyma [1]. The cell and tissue-penetration of these peptides is mediated by the C-end Rule (CendR): the motif R/KXXR/K must be exposed at the C-terminus of the peptide [2]. CendR motif binds to neuropilin receptor (NRP1/2), a cell surface receptor with essential roles in angiogenesis and regulation of vascular permeability that is overexpressed in a variety of tumors. iRGD peptide is a tumor-specific CendR peptide that uses a multistep mechanism for tumor homing and penetration. It contains the RGD motif that binds to 伪v-integrins on angiogenic tumor endothelium. Once recruited to tumor blood vessels, the iRGD peptide is proteolytically cleaved to expose the CendR motif, resulting in the extravasation, tumor penetration and cell entry of the peptide and payload [3,4]. Polymersomes are nanoscale vesicles formed by self-assembly of amphiphilic block-copolymers in aqueous solutions. Polymersomes can be used to encapsulate water-insoluble compounds in the hydrophobic membrane and watersoluble compounds in the vesicular lumen [5]. The surface of polymersomes can be functionalized with affinity ligands and dyes for

NanoBio&Med2015

targeted delivery and for tracking [6]. The high molecular weight of block copolymers enables the formation of highly entangled membranes, that confers them unique mechanical properties and elasticity [7]. Whereas the pH-sensitive polymersomes are stable at neutral pH, they disintegrate to release their cargo after cellular uptake and exposure to the low pH in the endolysosomal compartment [8,9]. Here, we used tumor penetrating peptides conjugated to polymersomes to deliver the drug-loaded nanoparticles to peritoneal and subcutaneous tumors in vivo. We studied the effect of iRGD functionalization on homing and therapeutic efficacy of paclitaxel-loaded pHsensitive polymersomes assembled from diblock co-polymer poly(oligoethylene glycol methacrylate)-poly(2-(diisopropylamino)ethyl methacrylate) (POEGMA-PDPA). Our study showed that intravenously or intraperitoneallyadministered polymersomes targeted with iRGD home to malignant lesions in different cancer models in mice (gastric, colon and prostatic cancer. After intraperitoneal administration, iRGD-polymersomes penetrated and accumulated in peritoneal tumors by a combination of direct penetration and systemic exposure. The improved tumor homing is translated to improved antitumor activity of intraperitoneal iRGD-functionalized paclitaxelloaded polymersomes compared to the free

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paclitaxel and Abraxane, a nanoformulation clinically used for cancer treatment. Our studies show that tumor penetrating peptides can be used to increase the selectivity of drugloaded polymersomes to achieve increased therapeutic efficacy and lower side effects.

References [1] Teesalu T, Sugahara KN, Ruoslahti E, Front Oncol, 3 (2013) 326. [2] Teesalu T, Sugahara KN, Kotamraju VR, et al. Proc Natl Acad Sci USA, 106 (2009)16157. [3] Sugahara KN, Teesalu T, Karmali PP, Kotamraju VR, Agemy L, Greenwald DR, Ruoslahti E. Science, 328 (2010) 1031. [4] Sugahara KN, Teesalu T, Karmali PP, et al. Cancer Cell, 16 (2009) 510.

[5] Pegoraro C, Cecchin D, Simon-Gracia L, Warren N, Madsen J, Armes SP, Lewis A, MacNeil S, Battaglia G. Cancer Lett 334 (2013) 328. [6] Tian X, Nyberg S, Sharp PS, Madsen J, Daneshpour N, Armes SP, Berwick J, Azzouz M, Shaw P, Abbott NJ, Battaglia, G. Sci Rep 5 (2015) 11990. [7] Pegoraro C, Cecchin D, Madsen J, Warren N, Armes SP, MacNeil S, Lewis A, Battaglia G. Biomater Sci 2 (2014) 680. [8] Du J, Tang Y, Lewis AL, Armes SP. J Am Chem Soc 127 (2005) 17982. [9] Simón-Gracia L, Hunt H, Scodeller P, Gaitzsch J, Braun GB, Willmore AA, Ruoslahti E, Battaglia G, Tambet Teesalu T. Molecular Cancer Therapeutics. Submitted.

Figures

Figure 1. Overview of the polymersome formation and tumor targeting mechanism. POEGMA-PDPA co-polymer form stable vesicles in aqueous medium at pH ˃ 6.5. iRGD peptide attached to the polymersome surface binds to αv integrins on angiogenic tumor endothelium, where the peptide is proteolytically processed to expose the CRGDK CendR motif at the Cterminus. The activated CendR motif acquires the ability to bind to cell- and tissue-penetration receptor NRP-1 (C). The drug-loaded polymersomes, extravasate, spread over the tumour stroma and enter into tumour cells. Inside the cell, polymersomes dissolve in acidic endolysosomal compartment releasing the drug to the cytosol due to the “proton sponge” effect.

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Bio-hybrid Janus Motors Driven by Escherichia coli Morgan M. Stanton1, Juliane Simmchen1, Xing Ma1, Albert Miguel-López1, Samuel Sánchez1,2,3 Max Planck Institute for Intelligent Systems, Heisenberg Str. 3, Stuttgart, Germany Institute for Bioengineering of Catalonia (IBEC), Baldiri I Reixac 10-12, Barcelona, Spain 3 Institució Catalana de Recerca i Estudis Avançats (ICREA), Pg. Lluís Companys 23, Barcelona, Spain 1

stanton@is.mpg.de A bio-hybrid microswimmer combines motile cells with artificial materials for a functional system capable of locomotion [1]. There has been significant interest in the development of microswimmers for medical drug and cargo delivery, but the majority of current micromotors rely on toxic fuel sources and materials in their design making them irrelevant for biomedical applications. Biologically driven motors offer the opportunity to efficiently convert chemical energy into mechanical work for propulsion without the toxic fuel sources that are used for catalytic micromotors [2]. Here, for a bio-hybrid system, a non-pathogenic form of Escherichia coli (E. coli), was integrated onto metal capped Janus micro and nano particles. The ability of the swimming E. coli to propel and interact with multiple material surfaces was investigated. Fabrication of the bio-hybrid was rapid and simple for a microswimmer capable of magnetic guidance and ferrying an anti-cancer agent. Cell adhesion was regulated as E. coli adhered only to the particle’s metal caps allowing the PS surface to be utilized for drug attachment, creating a multifunctional system. The dual capability of the bio-hybrid to have guided cell adhesion and localized drug attachment allows the swimmer to have multiple applications for biomedical microswimmers, future bacteria-interface systems, and micro-biorobots.

NanoBio&Med2015

References [1] Carlsen, R. W.; Sitti, M. “Bio-Hybrid CellBased Actuators for Microsystems,” Small. 2014, 10, 3831. [2] Stanton, M. M.; Trichet-Paredes, C.; Sánchez, S. "Applications of threedimensional (3D) printing for microswimmers and bio-hybrid robotics", Lab Chip, 2015, 15, 1634.

Figures

november 18-20, 2015 - Barcelona (Spain)

A nanotechnology approach to evaluate drug promiscuity in cancer Teresa Valero1,2, Victoria Cano-Cortés2, Asier Unciti-Broceta1 and Rosario Sánchez-Martín2 1

Edinburgh Cancer Research UK Centre (MRC Institute of Genetics and Molecular Medicine), University of Edinburgh, UK 2 Pfizer - Universidad de Granada - Junta de Andalucía Centre for Genomics and Oncological Research (GENYO), Parque Tecnológico de Ciencias de la Salud (PTS), Spain

teresa.valero@genyo.es Cancer remains an important public health problem in Europe. Among currently available treatments for cancer, the use of kinase inhibitors has gained unprecedented popularity during the last decade, in part motivated by the success of Imatinib in the treatment of Chronic Myeloid Leukemia [1]. However, the appearance of resistance episodes led to the development of other tyrosine kinase inhibitors (TKI) with a broader target spectrum such as Dasatinib [2-3]. This promiscuous kinase inhibitor targets BcrAbl, Src family, receptor tyrosine kinases and TEC family kinases [4]. Therefore, it is difficult to elucidate which target/s of Dasatinib is/are responsible for the phenotypic effect observed in each cancer type. Inhibitor’s promiscuity may be advantageous or a major drawback depending on the result of the inhibition, if it circumvents resistance mechanisms or if it leads to severe side effects. Therefore it is of crucial importance to discover all the targeted kinases and which of the kinases produce each effect in each cancer type, in order to develop personalized highly selective therapies [5]. Our research team has developed the synthesis, multifunctionalization and a variety of biological applications of polymeric micro/nanospheres as cellular delivery and tracking devices capable of introducing a range biological and chemical modalities into a vast majority of both primary and cell lines, including embryonic stem cells [6-9]. Importantly, and due to their cross-link nature, these synthetic nanoparticles allows multistep solid-phase chemistry, allowing covalent

binding of small molecules and macromolecules onto them, as well as the possibility to label them with different trackers, such as fluorophores and metals, thus creating multifunctional micro/nanospheres, an unique feature for this type of nanodevices. These nanodevices have been also fully proven to be biocompatible and have been used to sort out cells for subsequent downstream analysis. Upto-date, cargoes delivered into eukaryotic cells include metals (palladium nanoparticles) [10], proteins (such as Enhanced Green Fluorescent Protein (EGFP) and β-Galactosidase) [11], oligonucleotides (DNA, siRNA) [12-13]. Of special importance has been the development of sensors for measuring intracellular pH and calcium ions [14-15], and fluorogenic peptides to assess in situ caspase activity [16]. All of these applications require the internalization of these nanodevices inside cells. Recently, we have developed a quick method to quantify the number of nanoparticles per sample using a standard spectrophometry approach [17]. Based on this expertise, the central objective of this project is to develop an in situ and costefficient method based on nanotechnology to detect protein kinases which are targeted by promiscuous kinase inhibitors and to prove its functionality on living systems. This nanotechnology approach has been developed by conjugation of kinases inhibitors to fluorescently labelled nanoparticles. Dasatinib conjugation was used as proof of concept. Our recent results in the development of this nanotechnology will be presented.

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References [1] Z. a Knight, H. Lin, and K. M. Shokat, “Targeting the cancer kinome through polypharmacology.,” Nat. Rev. Cancer, vol. 10, no. 2, pp. 130–137, 2010. [2] A. Hochhaus and H. Kantarjian, “The development of dasatinib as a treatment for chronic myeloid leukemia (CML): From initial studies to application in newly diagnosed patients,” J. Cancer Res. Clin. Oncol., vol. 139, no. Cml, pp. 1971–1984, 2013. [3] J. Das, P. Chen, D. Norris, R. Padmanabha, J. Lin, R. V Moquin, Z. Shen, L. S. Cook, A. M. Doweyko, S. Pitt, S. Pang, D. R. Shen, Q. Fang, H. F. De Fex, K. W. McIntyre, D. J. Shuster, K. M. Gillooly, K. Behnia, G. L. Schieven, J. Wityak, and J. C. Barrish, “2-Aminothiazole as a novel kinase inhibitor template. Structure-activity relationship studies toward the discovery of N(2-chloro-6-methylphenyl)-2-[[6[4-(2hydroxyethyl)-1-piperazinyl]-2-methyl-4pyrimidinyl]amino]-1, 3-thiazole-5-carboxamide (Dasatini,” J. Med. Chem., vol. 49, no. 23, pp. 6819–6832, 2006. [4] J. Araujo and C. Logothetis, “Dasatinib: A potent SRC inhibitor in clinical development for the treatment of solid tumors,” Cancer Treat. Rev., vol. 36, no. 6, pp. 492–500, 2010. [5] G. C. Terstappen, C. Schlüpen, R. Raggiaschi, and G. Gaviraghi, “Target deconvolution strategies in drug discovery,” Nat. Rev. Drug Discov., vol. 6, no. 11, pp. 891–903, 2007. [6] R. M. Sanchez-Martin, M. Muzerelle, N. Chitkul, S. E. How, S. Mittoo, and M. Bradley, “Bead-based cellular analysis, sorting and multiplexing.,” Chembiochem, vol. 6, no. 8, pp. 1341–5, 2005. [7] A. Tsakiridis, L. M. Alexander, N. Gennet, R. M. Sanchez-Martin, A. Livigni, M. Li, M. Bradley, and J. M. Brickman, “Microsphere-based tracing and molecular delivery in embryonic stem cells.,” Biomaterials, vol. 30, no. 29, pp. 5853–61, 2009. [8] N. Gennet, L. M. Alexander, R. M. SánchezMartín, J. M. Behrendt, A. J. Sutherland, J. M. Brickman, M. Bradley, and M. Li, “Microspheres as a vehicle for biomolecule delivery to neural stem cells.,” N. Biotechnol., vol. 25, no. 6, pp. 442–9, 2009. [9] K. Turksen, Stem Cell Nanotechnology, vol. 1058. 2013.

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[10] A. Unciti-Broceta, E. M. V Johansson, R. M. Yusop, R. M. Sánchez-Martín, and M. Bradley, “Synthesis of polystyrene microspheres and functionalization with Pd0 nanoparticles to perform bioorthogonal organometallic chemistry in living cells,” Nat. Protoc., vol. 7, no. 6, pp. 1207–1218, 2012. [11] R. M. Sanchez-Martin, L. Alexander, M. Muzerelle, J. M. Cardenas-Maestre, A. Tsakiridis, J. M. Brickman, and M. Bradley, “Microspheremediated protein delivery into cells,” Chembiochem, vol. 10, pp. 1453–1456, 2009. [12] J. G. Borger, J. M. Cardenas-Maestre, R. Zamoyska, and R. M. Sanchez-Martin, “Novel strategy for microsphere-mediated DNA transfection,” Bioconjug. Chem., vol. 22, pp. 1904–1908, 2011. [13] L. M. Alexander, R. M. Sanchez-Martín, and M. Bradley, “Knocking (anti)-sense into cells: The microsphere Approach to Gene Silencing,” Bioconjug. Chem., vol. 20, no. Scheme 1, pp. 422–426, 2009. [14] M. C. S. M. M. B. Rosario M. Sachez-Marti, “Microsphere-Based Real-Time Calcium Sensing,” Angew. Chemie-International Ed., vol. 45, no. 33, pp. 5472–5474, 2006. [15] M. Bradley, L. Alexander, K. Duncan, M. Chennaoui, A. C. Jones, and R. M. SánchezMartín, “pH sensing in living cells using fluorescent microspheres.,” Bioorg. Med. Chem. Lett., vol. 18, no. 1, pp. 313–7, 2008. [16] J. M. Cárdenas-Maestre, A. M. Pérez-Lõpez, M. Bradley, and R. M. Sánchez-Martín, “Microsphere-based intracellular sensing of caspase-3/7 in apoptotic living cells,” Macromol. Biosci., vol. 14, pp. 923–928, 2014. [17] J. D. Unciti-Broceta, V. Cano-Cortés, P. AlteaManzano, S. Pernagallo, J. J. Díaz-Mochón, and R. M. Sánchez-Martín, “Number of Nanoparticles per Cell through a Spectrophotometric Method - A key parameter to Assess Nanoparticle-based Cellular Assays,” Sci. Rep., vol. 5, no. May, p. 10091, 2015.

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Graphene-based self-propelled micromotors as decontaminating micromachines of environmental pollutants D. Vilela1, J. Parmar2 and S. Sánchez1,2,3 Max-Planck Institute for Intelligent Systems, Heisenbergstr. 3, 70569 Stuttgart, Germany Institute for Bioengineering of Catalonia (IBEC), Baldiri I Reixac 10-12, 08028 Barcelona, Spain 3 Institució Catalana de Recerca i Estudis Avançats (ICREA), Psg. Lluís Companys, 23, 08010 Barcelona, Spain 1

vilela@is.mpg.de Pollution of water by contaminants and chemical threats is a prevalent topic in scientific, economic, political and, consequently, in the public media. These contaminants cause problems that can range from contamination of drinking water leading to human health problems including endocrine effects and bacterial resistance. It has motivated the use and development of "smart" materials such as catalytically powered micro- and nanomotors for efficient environmental monitoring and water remediation [1]. Graphene is a two-dimensional (2-D) sheet of carbon atoms connected by sp2 bonds. The graphene structure shows extraordinary properties, such as a high surface area, excellent thermal and electrical conductivities, optical transparency and a high mechanical strength and elasticity [2]. In addition, graphene-based nanomaterials have been also reported as adsorbents of various types of contaminant in air and water systems [3]. Here, novel graphene-modified self-propelled micromotors containing polymeric materials for the decontamination of environmental

pollutants have been developed. These motors were added in an aqueous polluted solution in the presence of H2O2 to propel the motors catalytically. After solution decontamination was complete the motors were extracted magnetically to preserve solution purification. This powerful decontamination is derived from the convection of self-propelled motors combined with the synergic effect of graphene as an adsorbent.

References [1] Ll. Soler and S. Sanchez, Nanoscale 6 (2014) 7175-7182; W. Gao and J. Wang, ACS Nano 8 (2014) 3170-3180; S. Sanchez, Ll. Soler, J. Katuri, Angew. Chem. 54 (2015) 1414-1444. [2] Y. Zhu, S. Murali, W. Cai, X. Li, J. W. Suk, J. R. Potts, R. S. Ruoff, Adv. Mater. 22 (2010) 12231234. [3] S. Wang, H. Sun, H. M. Ang, M. O. Tade, Chemical Engineering Journal 226 (2013) 336-347; Z. Gan, X. Wu, M. Meng, X. Zhu, L. Yang, P. K. Chu, ACS Nano 9 (2014) 93049310.

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Posters list: Alphabetical order Authors

Country Poster title

Aguilera Velázquez, José Raúl V. Venegas, J. M. Oliva, M. J. Sayagués, M. de Miguel, J. A. Sánchez Alcázar, M. ArévaloRodríguez, S. Calero and A. P. Zaderenko

Alea Reyes, Maria Elisa Sara Duran, Asensio Gonzalez, José A. Plaza and Lluïsa Pérez-García

Amirthalingam, Ezhil J. Soriano, S. Durán, J. A. Plaza, I. Mora-Espí, A. González-Campo and L. Pérez-García

Spain

EGFR-Targeted Tannic Acid Nanoparticles.

Spain

Pyridinium–mediated incorporation of porphyrin derivatives into micro and nanoparticles for photodynamic therapy.

Spain

Intracellular Reactive Oxygen Species (ROS) Sensing using Bi-Functional Microparticles for Cancer Theranostics.

Poland

Synthesis, characterization and application of ZnO/ZnS nanolayers in biosensor design.

Spain

Simple synthetic routes for the functionalization of nanodiamonds as biocompatible carbon nanoallotropes for future sensing and (bio)targeting applications.

Spain

Nanoscale Electric Permittivity of Single Bacterial Cells at GHz frequency by Scanning Microwave Microscopy.

United Kingdom

Nanomedicines for the Treatment of Tuberculosis.

Italy

Doxorubicin loaded – PAS masked human ferritin nano-cages for pancreatic tumor targeting.

Austria

Effect of Nanoparticle-Enzyme Interaction on Cellulase Activity.

Poland

Changes of endothelium nano-mechanics in response to the vascular dysfunction.

Germany

Studying synthetic micro swimmers near surfaces and using them to build self-assembled machines.

Baranowska-Korczyc, Anna Małgorzata Jasiurkowska-Delaporte, Anna Reszka, Tomasz Wojciechowski and Krzysztof Fronc

Bastos Arrieta, Julio Zhao Jingjing and Cristina Palet

Biagi, Maria Chiara Rene Fabregas, Georg Gramse, Marc Van Der Hofstadt, Antonio Juárez, Ferry Kienberger, Laura Fumagalli and Gabriel Gomila

Donnellan, Samantha MK Karen Stevenson, Helinor Johnston, Andrew Owen and Vicki Stone

Fracasso, Giulio Giamaica Conti, Cristina Anselmi, Elisabetta Falvo, Elisa Tremante, Alessandro Arcovito, Massimiliano Papi, Marcella Pinto, Nadav Elad, Alberto Boffi, Veronica Morea, Patrizio Giacomini and Pierpaolo Ceci

Hafner, Manuel Christian Becker

Jaglarz, Magdalena Marta Targosz-Korecka, Katarzyna MalekZietek, Elżbieta Buczek, Aleksandra Gregorius, Barbara Sitek, Stefan Chlopicki and Marek Szymonski

Katuri, Jaideep Juliane Simmchen, Laurent Helden, Clemens Bechinger and Samuel Sanchez

NanoBio&Med2015

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Authors Kumar, Ravi Michael Hirtz and Harald Fuchs

Lado Touriño, Isabel Arisbel Cerpa Naranjo, Viviana Negri, Sebastián Cerdán and Paloma Ballesteros

Country Poster title Germany

Multiplex Polymer pen lithography and its application in Biology.

Spain

Coarse-grained molecular dynamics simulation of water diffusion in the presence of carbon nanotubes.

Spain

Gold nanoparticles functionalized with serine protease inhibitors as a novel approach to Rosacea’s therapy.

Poland

The role of glycocalyx in cellular interactions between lung carcinoma cells and the endothelium.

Spain

New bacteriorhodopsin mutants with highly stable M intermediate as candidates for BR-based optical devices.

Spain

Imidazolium-based nanostructured supramolecular gels for drug delivery.

Spain

Pirimidine analog-capped gold nanoparticles for gene delivery.

Spain

Design of a Bio-Multi-Functional device for viscosity measurements.

Spain

Synthesis of Nanomaterials with multiple applications in Biomedicine.

Spain

Cross-fertilization of Key Enabling Technologies. Insights from Nano-enabled Medical Devices.

Spain

Multifunctional, reusable, self-powered nano-bots for cleaning polluted water.

Spain

An Instantaneous Portable Anemia Detection Device.

United Kingdom

Dithranol-loaded PLGA microspheres in conjugation with hydrogel patch: Potential application for psoriasis.

Israel

SP1, a Protein from Aspen Trees, as a Nanopore.

Spain

Catalytic/Bio-catalytic Janus Mesoporous Silica Nano-motors for Active Drug Delivery.

Limón, David María José Fábregas, Josefa Badia, Laura Baldomà, Ana C. Calpena and Lluïsa PérezGarcía

Malek-Zietek, Katarzyna Marta Targosz-Korecka and Marek Szymonski

Marco Garcia, Guillem Esteve Padrós Morell and Tzvetana Lazarova

Nunes Rodrigues, Ana Mafalda David Limón, Ezhil Amirthalingam, Gowtham Sathyanarayanan, Romen Rodriguez-Trujillo, Anna Calpena, David B. Amabilino and Lluïsa Pérez-García

Oliva Montero, José María L. Velarde, J. A. Sánchez-Alcázar, M. J. Sayagués and A. P. Zaderenko

Ortega Tañá, Laura Joan Cid Vidal, Ángeles Ivón Rodríguez Villarreal, Jordi Colomer Farrarons and Pere Miribel Català

Ortiz de Solórzano García, Isabel Laura Usón and Jesús Santamaría

Paez-Aviles, Cristina Esteve Juanola-Feliu and Josep Samitier

Parmar, Jemish Diana Vilel, Lluís Soler and Sámuel Sánchez

Punter-Villagrasa, Jaime Joan Cid Vidal, Ángeles Ivón Rodríguez Villarreal, Jordi Colomer Farrarons and Pere Miribel Català

Rahbari, Raha

Rotem, Dvir M. Akerman, N. Attias, L. Nesiel, K. Liu, A. Karmi, Y. Nevo, O. Shoseyov and D. Porath

Sanchez, Samuel Xing Ma, Anita Jannasch, Urban-Raphael Albrecht, Kersten Hahn, Albert Miguel-López and Erik Schäffer

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

Authors Sanuy, Andreu Pablo Loza-Alvarez, Jordi Andilla and David Gascón

Serrà, Albert N. Gimeno, E. Gómez, M. Mora, M. Ll. Sagristá and E. Vallés

Country Poster title Spain

Low-noise Wideband pre-amplifier for fluorescence microscopy.

Spain

Mesoporous Nanowires as Carriers for Drug Delivery.

Spain

Controlled linear movement of nanoparticles in suspension by homogeneous magnetic field gradients.

Poland

Methodology and evaluation of nanomechanical changes of endothelium in model of hyperglycemia: from in vitro to ex vivo experiments.

Spain

Caco-2 cells as an in vitro model to determine detrimental effects on the intestinal barrier. Studies with SiO2- and ZnO-NPs at sub-toxic doses.

Spain

Versatility of polymeric nanocapsules for the encapsulation of topical active ingredients of different physico-chemical nature.

Germany

Nobel TiO2/Au fuel-free nanomotors based on active Brownian motion under visible light.

Serrano Olmedo, Jose Javier Carlos David Amaya Jaramillo, Milagros Ramos Gómez and Francisco del Pozo Guerrero

Targosz-Korecka, Marta Katarzyna Malek-Zietek, Magdalena Jaglarz and Marek Szymonski

Vila Vecilla, Laura Alba García, Ricard Marcos and Alba Hernández

Vilar Palos, Gemma Jessica Romero, Lorena García-Fernández, Elisabet Fernández-Rosas, Jaume Oliva, Ignacio Umbert and Socorro VázquezCampos

Wang, Xu Varun Sridhar and Samuel Sánchez

NanoBio&Med2015

november 18-20, 2015 - Barcelona (Spain)

EGFR-Targeted Tannic Acid Nanoparticles a

J. R. Aguilera, V. Venegas, J. M. Oliva, M. J. Sayagués, M. de Miguel, J. A. Sáncheze f a a Alcázar, M. Arévalo-Rodríguez, S. Calero and A. P. Zaderenko a

Universidad Pablo de Olavide, Ctra. De Utrera Km.1. Seville, Spain; Centro de Investigación c Cooperativa en Biociencias, Bizkaia, Spain; Instituto de Ciencia de Materiales de Sevilla, d e Seville, Spain; Universidad de Sevilla, Seville, Spain; Centro Andaluz de Biología del f Desarrollo, Seville, Spain; Biomedal S.L, Seville, Spain apzadpar@upo.es

Abstract Tannic acid (TA), a natural polyphenol widely distributed in the plant kingdom, exerts a wide variety of biological activities including anticancer, antioxidant, antimicrobial and antiviral activities, as well as pro-longevity and protective effect against several damages and diseases. With respect to its anticancer activity, TA is especially promising to treat those types of tumors that overexpress the epidermal growth factor receptor (EGFR), a major target in cancer therapy [1,2], as TA modulates its activation and downstream signalling pathways triggering apoptosis [3]. Nevertheless, despite the evident benefits provided by delivering TA to EGFR, no specific delivery system to this receptor has been proposed yet. We have developed a method to synthesize stable TA nanoparticles in a single step. Additionally, our nanoparticles were (i) labeled with a fluorescent marker, propidium iodide (PI), to combine imaging and therapeutic applications, PEGylated to prevent protein adsorption, opsonization, nonspecific uptake, and to allow further functionalization [4-6], and conjugated to an antibody to EGFR. TA nanoparticles were synthesized by self-assembly of TA and poly(vinyl alcohol) through hydrogen bonding. The presence of both components in the nanoparticles was confirmed by FTIR, and their size determined both by electronic microscopy and dynamic light scattering (Figure 1 a). The entrapment efficiency of TA in nanoparticles was extremely high (> 90%), and was not affected by the addition of the fluorescent marker (PI) to the reaction medium during the synthesis process, rendering a loading capacity of 0,3%. PI was chosen as fluorescent marker given that this compound is not able to cross intact cell membranes itself. PI-loaded TA nanoparticles were then coated with polyethylene glycol (PEG) and a recombinant protein A, bearing a PEG-binding domain, which enables conjugating the antibody in a suitable orientation to interact with the receptor. To assess the cellular cytotoxicity of the nanoparticles, human squamous carcinoma (A-431) and human dermal fibroblast (HDF) cell lines were used. A-431 is an epidermoid carcinoma cell line widely used as model target to study the effect of therapeutic approaches directed to EGFR, owing to its high EGFR expression, while HDF is derived from the dermis of normal human skin. The cytotoxicity assay showed that nanoparticles were not toxic for the nontumoral HDF cell line whereas they were markedly toxic for the tumoral cell line A431 (Fig. 1b). To further investigate the receptor-specific uptake of our nanoparticles, A-431 cells were incubated with PI-labelled nanoparticles conjugated to the monoclonal antibody to EGFR, as well as with the corresponding nanoparticles without antibody and free PI in solution or nothing at all as control. Our results demonstrate that only TA nanoparticles conjugated to the antibody were able to enter the cell (Fig. 2). In summary, multifunctional tannic acid nanoparticles with extremely high entrapment efficiency, loaded with a fluorescent marker and targeted to EGFR were prepared. Our synthesis method offers a real and vastly improved alternative to the current techniques. The uptake assay demonstrated that our nanoparticles are able to enter the cells through a receptor-mediated mechanism, and are only toxic for the tumoral cells.

Acknowledgements: This work was supported by Junta de Andalucía (Proyecto de Investigación de Excelencia P10-FQM-6615 and PAIDI FQM- 319) and Ministerio de Economía y Competitividad, Gobierno de España (Proyecto de Excelencia CTQ2013-48396- P).

References 1. R. Roskoski, Pharmacol Res, 79, 2014, 34-74. 2. C. Yewale, D. Baradia, I. Vhora, S. Patil and A. Misra, Biomaterials, 34, (2013), 8690-8707. 3. M. Cichocki, M. Dałek, M. Szamałek and W. Baer-Dubowska, Nutr. Cancer, 2013, 66, 308314. 4. B. Maestro, I. Velasco, I. Castillejo, M. Arévalo-Rodríguez, A. Cebolla and J. M. Sanz, J. Chromatogr. A. 1208, ( 2008), 189–96. 5. R. Fernandez-Montesinos, P. M. Castillo, R. Klippstein, E. Gonzalez-Rey, J. A. Mejias, A. P. Zaderenko and D. Pozo, Nanomedicine, 4, (2009), , 919-930. 6. P. M. Castillo, M. de la Mata, M. F. Casula, J. A. Sánchez-Alcázar and A. P. Zaderenko, Beilstein J. Nanotechnol., 5, (2014), 1312-1319

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Figure 2. Fluorescence microscopy images of A431 cell cultures inoculated with PI-labeled TA nanoparticles conjugated to anti-EGFR. From left to right: Blue, green and red channels. Nuclei were stained with Hoechst (blue staining), monoclonal antibody was stained with AF488 (green staining), PI (red staining). Scale bar 25 µm.

PyridiniumÂąmediated incorporation of porphyrin derivatives into micro and nanoparticles for photodynamic therapy 1,2

1,2

M. Elisa Alea , Sara Duran , Asensio Gonzalez , JosĂŠ A. Plaza , LluĂŻsa PĂŠrez-GarcĂa 1

Departament de Farmacologia i QuĂmica TerapĂ¨utica, Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, EspaĂąa 2 Institut de NanociĂ¨ncia i Nanotecnologia UB (IN2UB), Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, EspaĂąa 3 Institut de CiĂ¨ncia de Materials de Barcelona (ICMAB Âą CSIC), Campus Universitari de Bellaterra, 08193 Bellaterra, EspaĂąa 4 Instituto de MicroelectrĂłnica de Barcelona (IMB-CNM Âą CSIC), Campus Universitari de Bellaterra, 08193 Bellaterra, EspaĂąa mlperez@ub.edu

Abstract In the last years, nanostructured systems have raised huge interest in the biomedical field because of 1 their biocompatibility and the potential application as delivery agents for therapy. One example is the use of such vehicles to target cells in cancer therapy. One of the most studied systems in drug delivery is gold nanoparticles (AuNP). The use of nanoparticles coated with photosensitizers to improve their 2 specificity in Photodynamic Therapy (PDT) has been reported. For the synthesis of organic and water soluble AuNP, different types of ligands have been studied as stabilizers, like water-soluble polymers, amino acid based amphiphiles or peptides. On the other hand, gemini surfactants display excellent properties in the preparation and stabilization of monodisperse gold 3 nanoparticles, but the combination of a gemini structure and pyridinium functionality has not yet been reported. In this work we describe a methodology for the synthesis of pyridinium-coated AuNP, based on the Brust-Schiffrin method, where the pyridinium amphiphiles act as both transfer agents and stabilizers. The synthesized AuNPs were characterized using UV-visible absorption spectroscopy, Transmission Electron Microscopy, Dynamic Light Scattering and Mass Spectrometry. On the other hand, polysilicon microparticles, which have already proved to have excellent biocompatibility and are able to be 4 internalized by cells, were also functionalized with the pyridinium ligands. 4

Furthermore, different porphyrins - which are one of the most studied photosensitizers for PDT - were loaded into these biocompatible micro and nanoparticles through non-covalent interactions with the pyridinium ligands. The presence of the porphyrins on the AuNP was assessed by UV-visible absorption spectroscopy and spectrofluorimetry, while in the case of microparticles the functionalization with the porphyrins derivatives was confirmed by fluorescence microscopy. Acknowledgements: This work was supported by the EU ERDF (FEDER) funds and the Spanish Government grants TEC2011-29140-C03-01/02 and TEC2014-51940-C2-1/2. MEA thanks the Universitat de Barcelona for a predoctoral grant (APIF). References [1] E. Boisselier and D. Astruc, Chem. Soc. Rev, 38 (2009) 1759-1782. [2] D. Bechet, P. Couleaud, C. Frochot, M.-L. Viriot, F. Guillemin, and M. Barberi-Heyob, Trends Biotechnol, , 26 (2008) 612Âą621. [3] L. Casal-Dujat, M. Rodrigues, A. YagĂźe, A. C. Calpena, D. B. Amabilino, J. GonzĂĄlez-Linares, M. BorrĂ s and LluĂŻsa PĂŠrez-GarcĂa, Langmuir, 28 (2012) 2368-2381. [4] NĂşria Torras, Juan Pablo Agusil, Patricia VĂĄzquez, Marta Duch, Alberto M. HernĂĄndez 3LQWR -RVHS 6DPLWLHU (QULTXH - GH OD 5RVD -DXPH (VWHYH 7HUHVD 6XiUH] /OXwVD 3pUH] GarcĂa, JosĂŠ A. Plaza, Adv Mater, DOI: 10.1002/adma.201504164. [5] E. D. Sternberg, D. Dolphin, and C. Brucker, Tetrahedron, 54 (1998) 4151-4202.

Figures

Figure 1. A) Porphyrin derivatives: x= (H or Zn), B) Complex pyridinium salt - porphyrin, C) Schematic representation of AuNPs with pyridinium salt and thiol- PEG- OH as ligand, D) UV-vis absorption spectra and TEM pictures of the AuNPs, E) SEM images of the polysilicon microparticles and F) Fluorescence microscopy images of the two derivatives porphyrins immobilized on polysilicon microparticles and their 3D image respectively.

Intracellular Reactive Oxygen Species (ROS) Sensing using Bi-Functional Microparticles for Cancer Theranostics 1,2

E. Amirthalingam,

J. Soriano, S. Durán, J. A. Plaza, I. Mora-Espí, A. González-Campo and L. 1 Pérez-García

1. Departament de Farmacologia i Química Terapèutica and Institut de Nanociència i Nanotecnología UB (IN2UB), Universitat de Barcelona, Barcelona, Spain 2. Institut de Ciència de Materials de Barcelona, ICMAB (CSIC), Campus de la UAB, Bellaterra, Spain 3. Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biociències, Universitat Autònoma de Barcelona, Bellaterra, Spain 4. Instituto de Microelectrónica de Barcelona, IMB-CNM (CSIC), Bellaterra, Spain mlperez@ub.edu Abstract The aim of this work is to develop a novel microtool for producing and sensing Reactive Oxygen Species (ROS) potentially useful in cancer treatment. For this study, we will be using two species acting as a photosensitizer: either Cytochrome C, an apoptotic agent containing heme group, or zinc porphyrin 1 derivatives. Both the species are known to have peroxidase activity and, upon light excitation, therefore they would release ROS necessary for initiating apoptosis leading to cell death. On the other hand, in order to sense ROS production, BODIPY 581/591 will be used: this fluorescent dye shows a shift in the fluorescence emission peak upon oxidation, allowing the detection and quantification of singlet oxygen 2 species. Our approach is to prepare and optimize a protocol for functionalizing bi-functional microparticles (build up of gold and polysilicon) with both BODIPY 581/591 and one selected photosensitizer. Finally, the photochemical behaviour of the bi-functionalized microparticles will be studied in an in vitro experiment, in order to test their capability and efficiency in sensing ROS in living cells. References [1] W. M. Sharman, C. M. Allen, J. E. Van Lier, Meth. Enzymol., 319 (2000), 376-400. [2] G. P. C. Drummen, L. C. M. V. Liebergen, J. A. F. O. D. Kamp, J. A. Post, Free Radic. Biol. Med., 33 (2002), 473-490. Acknowledgement Financial support from the Ministerio de Ciencia e Innovación (MICINN) (project TEC2011-29140-C03-02, TEC2014-51940-C2-2-R, TEC2011-29140-C03-01 and TEC2014-51940-C2-1-R) are acknowledged. E.A. thanks FI-DGR from Generalitat de Catalunya for a predoctoral grant.

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Simple synthetic routes for the functionalization of nanodiamonds as biocompatible carbon nanoallotropes for future sensing and (bio)targeting applications. 1

Julio Bastos , Zhao Jingjing and Cristina Palet 'HSDUWDPHQW GÂś(QJLQ\HULD 4XtPLFD 8QLYHUVLWDW 3ROLWqFQLFD GH &DWDOXQ\D 83& Av. Diagonal 647, 08028 Barcelona, Spain 2 Centre Grup de TĂ¨cniques de SeparaciĂł en QuĂmica, Unitat de Q.AnalĂtica, Departament de QuĂmica, Universitat AutĂ˛noma de Barcelona, 08193 Bellaterra, Catalunya, Spain julio.bastos@upc.edu 1

The progress in many fields of the modern technology within the last 20 years is determined by the rapid development and wide application of various nanomaterials such as nanoparticles (NPs) and nanodiamonds (NDs) [1Âą4]. NPs are object of great interest in modern chemical research due to their special properties such as electrical, magnetic, optical and others, which are distinct from both those of the bulk material and those of isolated atoms and molecules[5]. NDs are less toxic carbon nanoallotropes compared with wide extended carbon nanotubes. Their feasibility of applications due to their biocompatibility and high temperature resistant, make them an interesting matrix for the development of new low-dimensional hybrid carbon nanomaterials. This can be accomplished by the incorporation of NPs on the surface and further functionalization with organic molecules for specific (bio)recognition approaches[6Âą8]. The surface of NDs is partially oxidized. This means that it bears carboxylic groups and, in this sense, it may be considered as a sort of nano-analogs of carboxylic ion exchangers. In this communication, we present simple synthetic routes for the functionalization of NDs; firstly by an acidic activation stage. Secondly, by taking advantage of their ion exchange functionality of the loading of the NPs precursor and consequent NPs appearance due to chemical reduction or precipitation. Modification with silver (Ag+), platinum (Pt2+) and copper (Cu2+) proper ion salts is here presented and final materials are properly characterized by TEM analysis. In the case of the Cu@NDs, they are used as template nanoparticles. Galvanic displacement takes place between Au3+ and Cu0 due to the high difference of redox potentials is proposed in order to also prepare Au@NDs, This is advantageous for their further functionalization with organic molecules as antibodies and aptamers for specific (bio)recognition assays. References 1. 2. 3. 4. 5.

6. 7. 8.

Xu P, Han X, Zhang B, et al. (2014) Multifunctional polymer-metal nanocomposites via direct chemical reduction by conjugated polymers. Chem Soc Rev 43:1349Âą60. doi: 10.1039/c3cs60380f Hussain F (2006) Review article: Polymer-matrix Nanocomposites, Processing, Manufacturing, and Application: An Overview. J Compos Mater 40:1511Âą1575. doi: 10.1177/0021998306067321 Paul DR, Robeson LM (2008) Polymer nanotechnology: Nanocomposites. Polymer (Guildf) 49:3187Âą3204. doi: 10.1016/j.polymer.2008.04.017 Kao J, Thorkelsson K, Bai P, et al. (2013) Toward functional nanocomposites: taking the best of nanoparticles, polymers, and small molecules. Chem Soc Rev 42:2654Âą78. doi: 10.1039/c2cs35375j 5XL] 3 0XxR] 0 0DFDQiV - 0XUDYLHY '1 ,QWHUPDWUL[ 6\QWKHVLV RI 3RO\PHUĂ&RSSHU Nanocomposites with Tunable Parameters by Using Copper Comproportionation Reaction. Chem Mater 22:6616Âą6623. doi: 10.1021/cm102122c Mochalin VN, Shenderova O, Ho D, Gogotsi Y (2011) The properties and applications of nanodiamonds. Nat Nanotechnol 7:11Âą23. doi: 10.1038/nnano.2011.209 Purtov K V, Petunin a I, Burov a E, et al. (2010) Nanodiamonds as Carriers for Address Delivery of Biologically Active Substances. Nanoscale Res Lett 5:631Âą636. doi: 10.1007/s11671-010-9526-0 Fu C-C, Lee H-Y, Chen K, et al. (2007) Characterization and application of single fluorescent nanodiamonds as cellular biomarkers. Proc Natl Acad Sci U S A 104:727Âą32. doi: 10.1073/pnas.0605409104

Nanoscale Electric Permittivity of Single Bacterial Cells at GHz frequency by Scanning Microwave Microscopy Maria Chiara Biagi1, Rene Fabregas1, Georg Gramse2, Marc Van Der Hofstadt1, Antonio JuĂĄrez1,3, Ferry Kienberger4, Laura Fumagalli5, Gabriel Gomila6 1Institut

de Bioenginyeria de Catalunya (IBEC), Barcelona, Spain 2Johannes Kepler University Linz, Linz, Austria 3 Departament de Microbiologia, Universitat de Barcelona, Barcelona, Spain 3 Keysight Technology Austria GmbH, Linz, Austria 5 School of Physics and Astronomy, University of Manchester, Manchester, UK 6 Departament dâ&#x20AC;&#x2122;Electronica, Universitat de Barcelona, Barcelona, Spain mcbiagi@ibecbarcelona.eu Abstract Information on the local electromagnetic properties of single cells in the GHz frequency range has led to important application in therapeutic and diagnostic, such as microwave imaging and selective hyperthermia treatment for cancer and other diseases. Furthermore, it is also fundamental to assess the potential hazardous effects of microwaves on biosamples. In recent years, the Scanning Microwave Microscope (SMM) has appeared as a unique tool to provide access to the electromagnetic properties of samples in the GHz frequency range and with nanoscale spatial resolution. Indeed, combining the good lateral resolution of near-field measurements, far below the wavelength of the source radiation, with the high penetration depth of microwaves, SMM allows to obtain maps of the (even internal) complex impedance of materials, with high lateral spatial resolution. Yet, the quantification of the intrinsic dielectric properties (i.e. complex permittivity) of the sample from the complex impedance measurements obtained in standard SMM imaging modes such as contact, intermittent contact or lift mode remains a challenge. This is because the experimental signal is greatly affected by the huge presence of non-local contributions (topographic cross-talk) due to the non-planar irregular shape of objects like single cells. The cross-talk can amount to up to an 83% of the measured image contrast, thus preventing the direct use of the images to quantify the local complex permittivity the sample. We present here a methodology to quantify and remove it, which consequently allows the derivation of impedance images revealing only the intrinsic dielectric response of the sample. This intrinsic contribution is then suitable for a quantitative analysis and it enables, combined with 3D finite element numerical calculations, to extract and map the complex permittivity of the sample. We have applied this procedure to a single bacterial cell (E. coli) and obtained for the first time its complex permittivity at ~19 GHz, in dry and humid conditions.

Nanomedicines for the Treatment of Tuberculosis Samantha MK Donnellan, Karen Stevenson, Helinor Johnston, Andrew Owen, Vicki Stone School of Life Sciences, Heriot Watt University, Edinburgh, United Kingdom smd31@hw.ac.uk There is an urgent need to develop effective treatments for the disease tuberculosis (TB), caused by the organism Mycobacterium tuberculosis (Mtb). The burden of the disease is 1 enormous; with a third of the world’s population estimated at being infected with latent TB it 2,3 continues to kill up to two million people annually . Mortality rates of multi-drug resistant TB are increasing and there are the continuing problems of treating HIV infected patients who are prone to secondary infection, especially TB. Development of novel anti-TB drugs is timely and expensive. The objective of this project was to create a rapid screening assay for investigating the anti-mycobacterial activity of different nanomaterials. A related surrogate organism was used, Mycobacterium avium subsp. paratuberculosis (Map) causing paratuberculosis in ruminants, as it could be handled in a lower containment level facility (L2), making it a relevant and cheaper alternative to Mtb to develop and optimise the assay. Map has a cell wall similar in structure and chemical composition to that of Mtb, which hinders the entry of drugs and is resistant to a similar spectrum of drugs. Map has been transformed with a plasmid carrying the gene for Green Fluorescent Protein (GFP) to create a reporter strain 4 (Map-GFP), thus allowing both growth and viability to be tracked by fluorescence . The antimycobacterial properties of nano-preparations of silver, copper (II) oxide, zinc oxide and first line anti-TB drugs (solid drug nanoparticles [SDNs]) have been investigated using the developed screening assay. Growth was monitored over 7 days and a dose response was measured, showing the effects of the different nanoparticles at various concentrations. A method to analyze the data to compare between compounds has also been developed. SDNs, silver and copper (II) oxide were found to be highly anti-mycobacterial at a range of concentrations tested, whereas zinc oxide offered poor anti-mycobacterial activity. Imaging displaying uptake of SDNs and the metal/metal oxide NPs by infected macrophage-like cells has been achieved. This project has many stages involved and will contribute towards the development of nanomedicines for the treatment of TB.

References

Smith, J. Nanoparticle delivery of anti-tuberculosis chemotherapy as a potential mediator against drug-resistant tuberculosis. Yale J. Biol. Med. 84, 361–9 (2011).

WHO. Global Tuberculosis Report 2014. (2014). at <http://www.who.int/tb/publications/global_report/en/> (Accessed October 2015).

WHO. Global Tuberculosis Report 2013. (2013). at <http://apps.who.int/iris/bitstream/10665/91355/1/9789241564656_eng.pdf> (Accessed October 2015).

Williams, S. L., Harris, N. B. & Barletta, R. G. Development of a firefly luciferase-based assay for determining antimicrobial susceptibility of Mycobacterium avium subsp. paratuberculosis. J. Clin. Microbiol. 37, 304–9 (1999).

Doxorubicin loaded – PAS masked human ferritin nano-cages for pancreatic tumor targeting. Giamaica Conti1, Cristina Anselmi2, Elisabetta Falvo3, Elisa Tremante4, Alessandro Arcovito5, Massimiliano Papi6, Marcella Pinto2, Nadav Elad7, Alberto Boffi3,4,8, Veronica Morea4, Patrizio Giacomini5, Pierpaolo Ceci4, Giulio Fracasso2 1- Department of Neurological Biomedical and Movement sciences, University of Verona, Italy 2- Department of Medicine, Immunology Section, University of Verona, Italy, (giulio.fracasso@univr.it) ϯͲDepartment of Biochemical Sciences “A. Rossi Fanelli”, University “Sapienza”, Rome, Italy 4-CNR – National Research Council of Italy, Institute of Molecular Biology and Pathology, Rome Italy 5-Immunology Laboratory, National Cancer Institute Regina Elena, via delle Messi d’oro 156,Rome Italy 6-Istituto di Fisica, Università Cattolica del Sacro Cuore, Largo F. Vito 1, 00168, Rome, Italy 7-Department of Chemical Research Support, Weizmann Institute of Science, Rehovot, Israel 8-Center for Life Nano Science at Sapienza, Istituto Italiano di Tecnologia (IIT), Rome, Italy Abstract Targeted delivery of anti-cancer therapeutics to cancer cells ("site-specific drug delivery") aims at enhancing accumulation of the drugs within the tumor while guiding them away from potentially endangered healthy tissues. Selective delivery to disease sites is particularly critical for many current cancer therapeutics that are distributed non-specifically in the body, thereby inducing dangerous side effects. Among the currently investigated nano-carriers for targeted delivery, protein-cage molecules based on heavy chain ferritins (HFts) are attracting growing interest due to their exceptional characteristics: [1] • HFts protein-cage are of human origin and therefore the risks of immunoresponse and inflammatory response which may hamper the patients’ treatment are substantially reduced; • HFts exploit transferrin receptor 1 (TfR1), also called CD71, for their internalization • It is long known that TfR1 is up to 100 fold more expressed in cancer than in normal cells. • HTf nano-cages may be loaded with drugs/trackers following two strategies: cross-linking to the exterior surface and/or positioning of the drug/tracker in their internal cavity (about 8 nm in diameter). In the present work we report the use of a new protein nanosystem based on the heavy chain of the human protein ferritin which is genetically engineered to a repetitive sequence of the tripeptide ProlineAlanine Serine (PAS peptide, PASylation), which is endowed with high masking properties.[2] The nanosystem was loaded with the drug Doxorubicin (DOXO) and exploited for the specific targeting of the tumor cells. In “vitro” and “in vivo” assays have been performed to evaluate its killing efficacy. To assess in vitro the efficacy of our drug loaded nanosystem (HFt-PAS40-DOXO) to intoxicate and to kill cancer cells we have performed XTT assays on a human pancreatic cancer cell line, PaCa44 cells; the aim was to demonstrate that after encapsulation the drug, DOXO, preserved its pharmacological activity. As summarized in Figure 1 the IC50 of the drug alone was 0.24 µM and the IC50 of our nanosystem was quite superimposable (i.e. IC50 of 0.11 µM for HFt-PAS40-DOXO). Moreover, it is important to point out that our nanocage in the form of drug-free compound (i.e. HFt-PAS40) did not show any sign of toxicity in our assay (i.e. viability 100%). In “in vivo” experiments performed on a xenogeneic mouse model of pancreatic cancer (i.e. PaCa44 cells injected s.c., in female CD1 Nude mice) we observed a reduction of 84% in the tumor volume when the mice were i.v. treated with HFt-PAS40-DOXO (Fig. 2); moreover our DOXO-loaded nansystem was more efficacious in reduce the tumor volume than a new drug Aldoxorubicin (formerly INNO-206) [3] a tumor-targeted doxorubicin conjugate. Finally no signs of toxicity were detected in the treated mice (Fig.3). References

[1] K. Fan et al., WIREs nanomed Nanobiotechnol, 5, (2013) page :287-298. [2] M. Schlapschy el al., Protein Eng Des Sel., 26(8), (2013) page :489–501. [3] SP Chawla, JAMA Oncol., Sep 17, (2015) page:1-9.

Figure 1 “In vitro” efficacy of our HFt-PAS40-DOXO nanosystem on PaCa44 cells (XTT assay).

Figure 2 “In vivo” efficacy (i.e. the reduction of the tumor volume) of our HFt-PAS40-DOXO nanosystem in a xenogeneic mouse model of pancreatic cancer.

mouse weight tumor weight subtracted (grams)

CTRL

INNO-206

HFt-PAS40-DOXO

Figure 3 Analysis of the tumor weight of untreated and treated mice.

Effect of Nanoparticle-Enzyme Interaction on Cellulase Activity Manuel Hafner, Christian Becker Institute of Biological Chemistry, Department of Chemistry, University of Vienna, Währinger Straße 38, 1090 Vienna, Austria manuel.hafner@univie.ac.at Abstract Protein-nanoparticle conjugates become increasingly important for diverse research areas such as drug delivery, food chemistry, medical applications and biotechnology [1, 2]. In biotechnology, enzymenanoparticle conjugates have been reported to be useful because they seem to increase the application spectrum of most commercially interesting enzymes regarding temperature, pH and activity [3, 4]. In this context, cellulase is a highly interesting model since it is used for degradation of cellulose into oligo- and monosaccharides to obtain a readily available source of energy. In order to design cellulase-nanoparticle conjugates, two different nanoparticle materials were chosen, silica and magnetite. Silica is widely used in medical and food applications because of its low toxicity as well as its easy availability. Magnetite offers an easy separation method due to its magnetic properties [4]. All nanoparticles form a corona made of available lipids, sugars and proteins upon contact with any biological medium. This behavior, the corona effect, describes a strong physisorption of these molecules on nanoparticles [5, 6]. Here we use this effect to generate an easy to produce cellulasenanoparticle conjugate and report on the effects this physisorption has on cellulase and its activity. In addition, the effect of nanoparticle exposure on enzyme activity (without being adsorbed) was measured. Immobilization of the enzyme on the nanoparticle results in significant changes of the secondary structure of the enzyme and leads to a decrease in activity of up to 40 %, depending on enzyme concentration. In contrast to this finding, a significant increase in activity of up to 85% was found for enzyme that was incubated with nanoparticles, but did not adsorb to it. Both effects depend on the material of the nanoparticles and are not observed with other enzymes such as trypsin.

References

[1] S. Al-Zuhair, Biofuels, Bioproducts and Biorefining, 1 (2007) 57-66. [2] P. Fortina, L.J. Kricka, D.J. Graves, J. Park, T. Hyslop, F. Tam, N. Halas, S. Surrey, S.A. Waldman, Trends in Biotechnology, 25 (2007) 145-152. [3] C. Mateo, J.M. Palomo, G. Fernandez-Lorente, J.M. Guisan, R. Fernandez-Lafuente, Enzyme and Microbial Technology, 40 (2007) 1451-1463. [4] P. Tartaj, M.P. Morales, T. González-Carreño, S. Veintemillas-Verdaguer, C.J. Serna, Journal of Magnetism and Magnetic Materials, 290±291, Part 1 (2005) 28-34. [5] J. Klein, Proceedings of the National Academy of Sciences, 104 (2007) 2029-2030. [6] M.P. Monopoli, D. Walczyk, A. Campbell, G. Elia, I. Lynch, F. Baldelli Bombelli, K.A. Dawson, Journal of the American Chemical Society, 133 (2011) 2525-2534.

CHANGES OF ENDOTHELIUM NANO-MECHANICS IN RESPONSE TO THE VASCULAR DYSFUNCTION 1

Magdalena Jaglarz , Marta Targosz-Korecka , Katarzyna Malek-Zietek ,(OĪELHWD %XF]HN , 2 2 2 1 Aleksandra Gregorius , Barbara Sitek , Stefan Chlopicki and Marek Szymonski 1

Center for Nanometer-scale Science and Advanced Materials, Department of Physics of Nanostructures and Nanotechnology, Institute of Physics, Jagiellonian University, prof. Stanislawa Lojasiewicza 11, 30-348 Krakow, Poland 2 Jagiellonian Centre for Experimental Therapeutics (JCET),Bobrzynskiego 14, 30-348 Krakow,Poland

magdalena.jaglarz@uj.edu.pl

Endothelium plays an essential role in regulating blood pressure and vascular homeostasis. Endothelial cells (ECs) are in direct contact with the blood flow. Vascular dysfunction can lead to development of diseases of affluence, e.g., hypertension, diabetes and various inflammations. The mail purpose of the presented study was to validate the observation in vitro to classical ex vivo models of endothelial dysfunction. Characterization of the endothelial nano-mechanics was performed for the non-fixed aorta in a murine model of hypertension induced by N-nitro-L-arginine methyl ester (LNAME) as well as in diabetes model (db/db mice). We have conducted analysis of changes in the structure and elasticity of an inner layer of the mouse aorta, in particular changes involving endothelium in healthy and pathological tissues. Elastic properties (reduced Young's modulus) of the inner tissue surface were characterized using nanoindentation spectroscopy with a colloidal AFM tip. Based on the nanoindentation measurements, the distribution of the elasticity parameters is determined for different areas of the aortic wall. Both models used in our studies are associated with the development of endothelium dysfunction, which affects the ability to contraction/diastole of the entire vessel. The morphology of the tissue (probed with a sharp AFM tip) and the qualitative analysis the tissue elasticity indicate that the above-mentioned diseases induce nano-mechanical changes in the inner layer of the aorta. Additionally, the complementary methods were used in order to characterization of the structure of the aortic wall (E-SEM) and biochemical analysis of the NO production.

Acknowledgements: The research was supported by 1.1.2 PO IG EU project 320267 )13 ³(ODVWLFLW\ SDUDPHWHU DQG VWUHQJWK of cell to cell interaction as a new marker of endothelial cell dysfunction LQ K\SHUJO\FHPLD K\SRJO\FHPLD´

Studying synthetic micro swimmers near surfaces and using them to build self-assembled machines Jaideep Katuri, Juliane Simmchen, Laurent Helden, Clemens Bechinger and Samuel Sanchez Max Plank Institute for Intelligent Systems, Heisenberg straße 3, Stuttgart, Germany katuri@is.mpg.de Silica particles asymmetrically coated with Pt, which move in self generated chemical gradients, are model systems in the study of active matter[1, 2]. While there is a relatively good understanding of the motion of single active particles[3], the influence of the surfaces near which they swim is not well understood. Natural swimmers like bacteria show interesting behaviours near surfaces that is different from their bulk properties. There is reason to believe that surface properties can have a significant influence on synthetic swimmers as they propel by phoresis, an interfacial effect. Here, we use catalytically active spherical Janus micro-motors in order to investigate experimentally their motion on substrates of different properties. We use Total Internal Reflection Microscopy (TIRM) to estimate the height at which they swim and establish a relationship between swimming height and swimming speed. We further explore ways to modify this interaction and thereby obtain desired behaviors like directional control. Our understanding of the interaction of swimmers with the surfaces enables us to exploit surface properties to design nano-patters on the surface, much smaller than the radius of the swimmers themselves, to influence their directional behaviour. Further, we are able to use the tendency of selfpropelled particles to have stable orientations against obstacles to self-assemble rotors by using a combination of passive and active components. The speed and direction of the rotors can be well controlled leading to very reliable micro-machining systems.

References 1. Howse, J.R., et al., Self-Motile Colloidal Particles: from Directed Propulsion to Random Walk. Phys. Rev. Lett., 2007. 99(4): p. 048102. 2. Sánchez, S., Soler, L. and Katuri, J. (2015), Chemically Powered Micro- and Nanomotors. Angew. Chem. Int. Ed., 54: 1414±1444. doi: 10.1002/anie.201406096 3. Ebbens, S.J. and J.R. Howse, Direct Observation of the Direction of Motion for Spherical Catalytic Swimmers. Langmuir, 2011. 27(20): p. 12293-12296.

Figures

Fig 1. SEM and schematic representation of a self propelled Janus particles near a nano-patterned surface. Minute details of the surface can be used to control the propulsion behaviour of Janus particles.

Fig 2. Schematic representation of the two possible docking events leading to a Janus particle stably propelling the micro-gear.

0XOWLSOH[ 3RO\PHU SHQ OLWKRJUDSK\ DQG LWV DSSOLFDWLRQ LQ %LRORJ\ Ravi Kumar1,2, Michael Hirtz1, and Harald Fuchs1,2 1Institut

für Nanotechnologie (INT) & Karlsruhe Nano Micro Facility (KNMF), Karlsruher Institut für

Technologie (KIT), 76021 Karlsruhe, Germany. 2Physikalisches

Institut & Center for Nanotechnology (CeNTech), Universität Münster, 48149 Münster,

Germany. ravi.kumar@kit.edu, michael.hirtz@kit.edu Abstract: Polymer pen lithography (PPL) is a hybrid technique of dip-pen nanolithography (DPN) and conventional microcontactprinting (µCP). It has the capability of patterning large area (cm²) with precise spatial control utilizing the several ten-thousands of tips on a square centimetre sized PPL stamp as pens, without denaturing or damaging delicate organic and biologically active compounds. The ink transfer mode is alike microcontact printing or pen spotting approaches, depending on the ink / substrate combination. Multiplexing, i.e. patterning more than one ink compound in close proximity onto the surface is highly demanded in biological applications and PPL is an ideal technique to create this kind of large area multiplex bioactive surfaces with high resolution. Here we present the application of multiplex PPL for patterning multiple azide ink and multiple single strand oligonucleotide array. The azide multiplex pattern are with different fluorescent azide (e.g. Tamra Azide, Alexa 488 Azide, cy3 azide etc.) and bio-active ink DNP (2,4-Dinitrophenol) azide on an alkyne-terminated surface with copper catalyzed azide-alkyne cycloaddition (CuAAC). The co-localization of the labelled IgEFc‫ܭ‬RI complex on the mast cells¶ PHPEUDQH LV also observed. The multiplexed ssDNA array also observed by hybridization with complementary ssDNA. Applications of this improved multiplexing strategy offers direct routes in e.g. large area patterning for substrates used in the study of mast cell activation in the future and better understanding of gene expression and interaction with multiplexed DNA arrays. References: [1] S. Sekula-Neuner, J. Maier, E. Oppong, A. C. B. Cato, M. Hirtz, and H. Fuchs, Small, 2012, 8, 585± 591. [2] F. Huo, Z. Zheng, G. Zheng, L. R. Giam, H. Zhang, and C. A. Mirkin, Science (80-. )., 2008, 321, 1658±60 [3] F. Brinkmann, M. Hirtz, A. M. Greiner, M. Weschenfelder, B. Waterkotte, M. Bastmeyer, and H. Fuchs, Small, 2013, 9, 3266±3275.

Figure 1: Fluorescence microscope image of co-localization of the labelled IgE-Fc‫ܭ‬RI complex in Mast cell on multiplex azide pattern. a) Tamra azide pattern b) DNP azide pattern showing IgE-Fc‫ܭ‬RI complex labelled with alexa 647 c) merged image of a and b. The scale bar is 20 µm.

Figure 2: Fluorescence microscope image of 5x5 multiplex azide pattern with four different florescent azide. The Cy3 Azide is in orange color (Mix of signal from FITC and TexasRed channel), Tamra Azide in red, Alexa 488 azide in green and Alexa 647 azide in pink. The scale bar is 20 µm.

Coarse-grained molecular dynamics simulation of water diffusion in the presence of carbon nanotubes Isabel Lado Touriñoa, Arisbel Cerpa Naranjoa, Viviana Negrib, Sebastián Cerdánb, Paloma Ballesterosc a

Department of Industrial Engineering, Universidad Europea de Madrid, C/Tajo s/n, 28670 Villaviciosa de Odón, Spain, b LIERM, Institute of Biomedical Research “Alberto Sols”, CSIC, 28029 Madrid, Spain c Laboratory of Organic Synthesis and Molecular Imaging by Magnetic Resonance, Faculty of Sciences, UNED, 28040 Madrid, Spain misabel.lado@uem.es Abstract Computational modeling of water diffusion in anisotropic media entails vital relevance to understand correctly the information contained in the magnetic resonance images weighted in diffusion (DWI) and of the diffusion tensor images (DTI). The effectiveness of DTI may be improved by the design of new Contrast Agents increasing the quality, resolution and specificity of the magnetic resonance images. We have previously shown that paramagnetic carbon nanotubes (CNTs) are able to perturb the diffusion of surrounding water molecules in an anisotropic manner, with larger effects in the longitudinal than in the transversal directions, constituting at present the first contrast agent for DTI [1]. In the present work we investigated the validity of a coarse-grained (CG) model to simulate water diffusion in a medium containing CNTs as models of anisotropic water diffusion behavior. The CG approach was chosen as it allows handling time and length scales of systems beyond what is feasible with traditional all-atom models. We report on the influence on water diffusion of various parameters such as length and concentration of CNTs, comparing the CG results with those obtained from a classic force field calculation. We show that water diffusion outside the CNTs follows Fick´s law, while water diffusion inside the CNTs is not described by a Fick´s behavior. Calculated water diffusion coefficients decreased in the presence of CNTs in a concentration dependent manner. We also observed smaller water diffusion coefficients for longer CNTs. Using the CG methodology we were able to demonstrate anisotropic diffusion of water inside the nanotube scaffold, but we could not prove anisotropy in the surrounding medium, suggesting that grouping several water molecules in a single diffusing unit may affect the diffusional anisotropy calculated. The methodologies investigated in this work represent a first step towards the study of more complex models with reasonable savings in computation time [2]. References [1] V. Negri, A. Cerpa, P. López.Larrubia, L. Nieto-Charques, S. Cerdán, P. Ballesteros, Nanotubular paramagnetic probes as contrast agents for magnetic resonance imaging based on the diffusion tensor, Angew. Chem. 122 (2010) 1857. [2] I. Lado Touriño, A. Cerpa Naranjo, V. Negri, S. Cerdán, P. Ballesteros, J. Mol. Graphics Modell. 62 (2015) 69.

Figure 1. Models used for CG (A, B, C) and all-atoms simulations (D). In GG models, grey and blue beads represent CNTs and water molecules respectively: A) 1 CNT, B) 2 CNTs, C) 4 CNTs, D) allatoms model.

Gold nanoparticles functionalized with serine protease inhibitors as a novel approach to RosaceaÂˇV WKHUDS\ David LimĂłn, MarĂa JosĂŠ FĂĄbregas, Josefa Badia, Laura BaldomĂ , Ana C. Calpena, LluĂŻsa PĂŠrez-GarcĂa. Universitat de Barcelona, Av. Joan XXIII s/n 08028, Barcelona, Spain Institut de NanociĂ¨ncia i Nanotecnologia IN2UB, Universitat de Barcelona, 08028, Barcelona, Spain dlimonma7@alumnes.ub.edu Abstract It has recently been discovered the role of the serine protease Kallikrein 5 in the development of Rosacea, a chronic autoimmune inflammatory skin disease [1]. Kallikrein 5 is overexpressed and altered in Rosacea patients, producing the pro-LQIODPPDWRU\ SHSWLGHV Â´FDWKHOLFLGLQVÂľ OHDGLQJ WR VHFUHWLRQ RI ,/-8, vasodilation, and facial erythema. Gold nanoparticles are highly interesting drug delivery systems for their stability and low toxicity, besides their properties for efficiently delivering pharmaceuticals to tumors and sites of inflammation through the Enhanced permeability and retention effect [2,3]. Therefore, as a novel approach to RosaceaÂˇV treatment, highly water soluble gold nanoparticles were successfully synthesized and further functionalized with serine protease inhibitors (Figure 1). The potential of this new approach is demonstrated by the in vitro inhibition of human Kallikrein 5, which showed to be dose-dependent. The dose required to inhibit enzymatic activity to 50% (IC 50) varied depending on the inhibitor attached to the nanoparticles and the type of functionalization (Figure 2). Nanoparticles can be successfully internalized in human keratinocytes (HaCaT) when working at the concentrations in which they inhibit Kallikrein activity, as observed by Fluorescence Microscopy. Cytotoxicity of nanoparticles is also minimal at this range of concentrations as cell viability ranges from 70% to 80%. The inflammatory response observed in Rosacea, mediated by the TLR-2 receptors, was successfully induced in human keratinocytes leading to a higher enzymatic activity of Kallikrein 5 and a higher production of IL-8. Nanoparticles either with serine protease inhibitors or without functionalization effectively reduced the production of IL-8 in a dose dependent way (Figure 3). The correlation of this IL-8 reduction with the decrease of enzymatic activity was also demonstrated by evaluating the Kallikrein 5 activity in human keratinocytes in presence of these nanoparticles. Furthermore, the high water solubility and stability of these gold nanoparticles make them a suitable option for therapy. All these results show these nanoparticles to be a completely novel potential treatment of Rosacea. References [1]

Yamasaki K, Di Nardo A, Bardan A, Murakami M, Ohtake T, Coda A, et al. Increased serine protease activity and cathelicidin promotes skin inflammation in rosacea. Nat Med 2007;13:975Â˛80.

[2]

Iyer AK, Khaled G, Fang J, Maeda H. Exploiting the enhanced permeability and retention effect for tumor targeting. Drug Discov Today 2006;11:812Â˛8.

[3]

Amirthalingam E, Rodrigues M, Casal-Dujat L, Calpena AC, Amabilino DB, Ramos-LĂłpez D, et al. Macrocyclic imidazolium-based amphiphiles for the synthesis of gold nanoparticles and delivery of anionic drugs. J Colloid Interface Sci 2015;437:132Â˛9.

Acknowledgements This work was supported by the EU ERDF (FEDER) funds and the Spanish Government grants TEC2011-29140-C03-01/02 and TEC2014-51940-C2-1/2.. D.L. thanks the CONACYT (MĂŠxico) for a predoctoral grant.

Figures

Figure 1. Schematic representation of gold nanoparticles functionalized covalently with AEBSF (A). with both alcohol and carboxyl groups (B). with AEBSF electrostatically (C). and with alcohol groups (D).

Figure 2. Inhibition of Kallikrein 5 activity with AEBSF, and different kinds of gold nanoparticles schematized in Figure 1.

Figure 3. Inhibition of IL-8 with different kinds of gold nanoparticles schematized in Figure 1.

The role of glycocalyx in cellular interactions between lung carcinoma cells and the endothelium Katarzyna Malek-Zietek, Marta Targosz-Korecka, and Marek Szymonski Center for Nanometer-Scale Science and Advanced Materials, NANOSAM, Faculty of Physics, Astronomy, and Applied Computer Science, Jagiellonian University, Prof. Stanislawa Lojasiewicza 11, 30-348 Krakow, Poland

katarzyna.malek@uj.edu.pl

A detailed understanding of the tumor extravasation mechanism is crucial for designing any therapeutic treatment against circulating tumor cells (CTCs) in the bloodstream. Adhesive interactions between cancer cells and the endothelial cells (ECs) strongly depend on the structure of the glycocalyx. Therefore, we characterized the endothelial glycocalyx (eGC) of Primary Pulmonary Artery Endothelial Cells (PHAEC) and the glycocalyx layer of lung carcinoma cells (A549) cultured on fibronectin-rich surface. Nanoindentation spectroscopy with a spherical AFM probe was used to determine the eGC thickness and eGC stiffness before (reference) and after treatment of cells with heparinase I and heparin. Later, the adhesive interactions between lung cancer cells and ECs have been studied. A novel approach was employed to attach the living cancer cell to the tipless AFM probe. The interactions were validated for living cells containing either a native or enzymatically digested glycocalyx. Multiparameter analysis of the measured force-distance curves demonstrated that reduction of the glycocalyx layer by heparinise I caused stronger adhesive interactions between A549 cells and PHAEC. Adding to the measurement system heparin instead of heparinase I caused renewal of the glycocalyx and decrease of adhesive interactions between cells. Therefore, we conclude that the structure of the eGC strongly affects the adhesion process between CTCs and the endothelium. Acknowledgments: 7KH UHVHDUFK ZDV VXSSRUWHG 32 ,* (8 SURMHFW Âł(ODVWLFLW\ SDUDPHWHU DQG VWUHQJWK RI FHOO WR FHll LQWHUDFWLRQ DV D QHZ PDUNHU RI HQGRWKHOLDO FHOO G\VIXQFWLRQ LQ K\SHUJO\FHPLD K\SRJO\FHPLDÂ´ program POMOST FNP.

New bacteriorhodopsin mutants with highly stable M intermediate as candidates for BR-based optical devices Guillem Marco Garcia, Esteve Padrós Morell, Tzvetana Lazarova Center for Biophysical Studies, Unitat de Biofísica, Departament de Bioquímica i Biologia Molecular, Facultat de Medicina, Universitat Autònoma de Barcelona, Bellaterra (Barcelona) Spain guillemmarco@gmail.com Abstract In the field of electronics, there are two approaches to build and improve electronic components. First, in the top-down approach, large materials are processed to obtain small components and this way is used for example to construct the classical silicon transistors or for graphene components. Second, the bottom-up approach consist in using molecules as small building blocks. According to the Moore’s law [1] the number of transistors by chip is doubled every two years and the components will reach the molecular level around the year 2030 [2]. A hybrid field between electronics and biochemistry, the bioelectronics, was developed to avoid the collapse of Moore’s law, and consists in using biological molecules, natural or modified, as a small building blocks for electronic components. This is a multidisciplinary effort, involving areas like biology, biochemistry, chemistry, electronics or bioethics and several candidates are proposed, like chloroplasts [3], photosystem I [4], rhodopsin [5] or modified E. coli organisms [6]. Bacteriorhodopsin is a light-driven proton pump found in the membrane of extreme halophilic archaeon Halobacterium salinarum and has several characteristics to be a good candidate as biological material for biotechnological applications, namely abundance in the nature, efficient solar absorption, resistant up to 140ºC [7], long-term stability, tolerating to a wide pH range, resistant to enzymatic digestion, stable at high ionic strength or stability in apolar solvents like hexane [8]. The absorption of light by the BR triggers three molecular events that could be used to classify BR applications: charge separation, chromophore protonation and proton transport (see Figure 1). BR undergoes several changes during the proton transport forming photointermediates with a characteristic maximum wavelength absorption in the called photocycle (Figure 2). One of them, the M intermediate shows the biggest spectral shift, about 150 nm and this photochromism could be used for information processing and storage. For building memories is necessary two interconvertible and differentiable stable states and basal and M state are good candidates, following the model on figure 3. Here we report some properties of two BR mutants, K129C and K129C/F71C, placed on extracellular side of helix D and loop BC, in aqueous solution and on film. Whereas in the wild type suspensions the life time of M is at about several milliseconds and low temperatures (240K) are required to be trapped upon illumination [9], these BR cysteine mutants are able to produce stable M even at room temperature. We found that by controlling pH and humidity of the mutants’ films the half-life of M decays can be enlarged up to 200 minutes. The stable M product is a key parameter for the suitability of BR to be used in optical technology [10]. The perturbation of the active site, the conformation of the protein and the stability to thermal stress of K129C and K129C/F71C were also studied and the potential of the two mutants as biomaterials candidates for nanotechnology is discussed. References 1.Moore, G.E., Electron Devices Meeting, 1975 International. 1975. 2.Robert, R.B., Molecular and Biomolecular Electronics. 1994, American Chemical Society. p. 1-14. 3.Greenbaum, E., The Journal of Physical Chemistry, 1990. 94(16): p. 6151-6153. 4.Lee, I., J.W. Lee, and E. Greenbaum, Physical Review Letters, 1997. 79(17): p. 3294-3297. 5.Birge, R.R., Annual Review of Physical Chemistry, 1990. 41(1): p. 683-733. 6.Vilanova, C., et al., Journal of Biotechnology, 2011. 152(3): p. 93-95. 7.Shen, Y., et al., Nature, 1993. 366(6450): p. 48-50. 8.Bamberg, E., et al., Proceedings of the National Academy of Sciences of the United States of America, 1981. 78(12): p. 7502-7506. 9.Lazarova, T., et al., Biophys J, 2000. 78(4): p. 2022-30. 10.Seitz, A. and N. Hampp, Journal of Physical Chemistry B, 2000. 104(30): p. 7183-7192.

Figures 1.

Imidazolium-based nanostructured supramolecular gels for drug delivery 1,2

1,2

Mafalda Rodrigues , David Limón , Ezhil Amirthalingam , Gowtham Sathyanarayanan , Romen 3 2,4 5 1,2 Rodriguez-Trujillo , Anna Calpena , David B. Amabilino , Lluïsa Pérez-García 1

Departament de Farmacologia i Química Terapèutica, Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, España 2 Institut de Nanociència i Nanotecnologia UB (IN2UB), Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, España 3 Institut de Ciència de Materials de Barcelona (ICMAB-CSIC), Campus Universitari de Bellaterra, 08193 Bellaterra, España 4 Departament de Farmàcia i Tecnologia Farmacèutica, Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, España 5 School of Chemistry, University of Nottingham, University Park, NG7 2RD mlperez@ub.edu

Abstract Low molecular weight gelators are promising materials for use as drug delivery systems. In our group we have developed a family of bis-imidazolium gemini-type amphiphiles which were studied as possible gelators. It was found that with the appropriate water/ethanol ratio, thermo reversible gels are formed. These gels could be obtained with the amphiphilic molecules with meta and para substitution (Figure1).[1] Furthermore, the effect of chain length in the geling ability was also assessed for the amphiphile molecules with the meta substitution. The obtained gels were thoroughly characterized by infrared spectroscopy, X-ray powder diffraction, NMR. To determine the tridimensional structure of the gels, these were analysed by SEM, AFM and TEM. The images obtained show that the amphiphilic components of the gels self-assemble into fibbers, approximately 100 nm wide, and fuse and cross forming a network, which can be seen by SEM (Figure 2). The rheological behaviour was also assessed to determine the viscosity of the gels and their possible application as delivery agents. As found in literature, there are examples of imidazolium-derived ionofores that are able to form gels with ibuprofen and release it.[2] Based in this, some model anionic drugs were incorporated successfully into the gel. The gels containing the drugs were further characterized using the above mentioned techniques and showed important differences: the presence of the drugs appears to contribute to the stability of the gel, which may indicate that the type of interaction of the drug with the gel influences the stability. These findings suggest that these bis-imidazolium amphiphilic molecules have great potential to be used as vehicles in drug delivery application.

Acknowledgements: This work was supported by the EU ERDF (FEDER) funds and the Spanish Government grants TEC2011-29140-C03-01/02 and TEC2014-51940-C2-1/2. D.L. thanks Conacyt for a predoctoral grant. E. A. thanks Generalitat de Catalunya for a predoctoral grant. References

[1] M. Rodrigues, A. C. Calpena, D. B. Amabilino, M. L. Garduño-Ramírez, L. Pérez-García, J. Mat. Chem. B, 2 (2014) 5419-5429 [2] L. Viau, C. Tourné-Péteilh, J.-M. Devoisselle, A. Vioux, Chem. Commun., (2010) 228±230

Figures

Figure 1: Structure of the bis-LPLGD]ROLXP DPSKLSKLOHV %U DQG %U.

Figure 2: 6(0 LPDJHV RI WKH JHOV REWDLQHG ZLWK %U $ DQG %U %

Pirimidine analog-capped gold nanoparticles for gene delivery 1

J.M. Oliva-Montero , L. Velarde , J.A. Sánchez-Alcázar , M.J. Sayagués , A.P. Zaderenko

Universidad Pablo de Olavide, Ctra. De Utrera Km.1. Seville, Spain. Centro Andaluz de Biología del Desarrollo,CSIC-UPO, Seville, Spain. 3 Instituto de Ciencia de Materiales de Sevilla, CSIC-US, Seville, Spain. * apzadpar@upo.es 2

Abstract A major drawback related to the use of chemotherapeutic drugs in cancer therapy refers to their lack of selectivity and, consequently, undesirable side effects. Due to the combined impact of cancer, together with adverse side effects of many conventional chemotherapeutic agents, a significant effort is devoted to the design of new therapeutic approaches. Recent advances in gene therapy point to the potential of this strategy to treat severe malignancies, such as brain tumor glioblastoma [1]. In this sense, posttranscriptional blocking of specific proteins by small interfering RNA (siRNA) is a promising technique with an enormous therapeutic potential in cancer treatment [2,3], although this approach is only capable of temporarily blocking protein expression. To solve the transient nature of siRNA, it is necessary to use plasmid DNA [1]. Some major problems associated with plasmid DNA therapy are that DNA does not cross the cell membrane due to its negative charge¸ DNA shows poor stability and endosomal escape in vivo, and lacks selectivity for tumor cells. These drawbacks can be overcome using delivery systems to direct plasmid DNA to its target cells. Gold nanoparticles are particularly promising as delivery systems due to their poor toxicity, compared to more conservative targeting systems such as polycationic polymers [4] and their ability to be used both in cancer diagnosis and therapy. Gold nanoparticles can be effectively used as contrast agents, as carriers for chemotherapeutic drugs and to destroy cancer cells by acting as heat mediators in hyperthermia treatments [5]. We have developed and optimized an in situ method to obtain 4,6-diamino-2-mercaptopirimidine (APY)-capped gold nanoparticles (AuNPӨAPY). Our nanoparticles are stable under physiological conditions (Fig. 1A) and show high affinity for DNA (Fig. 2B). Additionally, the presence of amine groups in the nanoparticles surfaces enables covalent binding to targeting antibodies [6] (Fig 1.C)]. Nanoparticles were characterized by UV-Vis, FTIR, DLS and TEM (Fig. 1B). Furthermore, in vitro tests revealed their lack of cytotoxicity, and ability to be selectively targeted to cancer cells overexpressing EGFR receptor. Funding: This work was supported by Junta de Andalucía (Proyecto de Investigación de Excelencia P10-FQM-6615). References [1] Guerrero-Cázares H. et al. ACSnano 8(5), (2014) 5141-5153. [2] Aagaard, L.; Rossi, J.J. Adv Drug Deliv Rev, 59 (2007) 75-86. [3] Masiero M.; Nardo G.; Indraccolo S. Future Medicine, 28 (2007) 143-166. [4] Nimesh S.; Gupta N.; Chandra, R. Nanomedicine, 6(4) (2011) 729-746. [5] Sultana S.; Khan M. R.; Kumar M.; Kumar S.; Ali M. J. Drug Target 21 (2013) 107-125. [6] Castillo P.M. et al. Nanomedicine 4(8) (2009) 919-930.

Figures

Figure. 1. AuNPÓ¨APY Nanoparticles (A) pH stability, (B) TEM image, (C) Dark Field & Fluorescence image in cell cultures.

Figure. 2. Nanoparticle-DNA plasmid complex:: (A) Representative diagram, and ( B) Gel retardation assay.

Design of a Bio-Multi-Functional device for viscosity measurements Laura Ortega TaĂąĂĄ, Joan Cid Vidal, Ă ngeles IvĂłn RodrĂguez Villarreal, Jordi Colomer Farrarons, Pere Miribel CatalĂ D2In, Electronics Department Universitat de Barcelona, MartĂ i FranquĂ¨s 1, 08028 {lortega,pmiribel}@el.ub.edu Abstract In this work is presented a new envisaged micro-rheometer device based on a Lab-on-a-Chip solution, which is focused to biomedical applications, Figure 1. The fluid viscosity is an important and useful characteristic of fluids in many fields. In industry, for example, is applied to analyze oils as lubricants [1], fuels in aviation for save landings [2] and polymers in manufacturing processes [3]. In medicine, blood viscosity is an important factor in the analysis of cardiovascular diseases and in the effectiveness of several therapies [4, 5]. The device is not only conceived to work with blood. We developed a Lab-on-a-chip device has the capability to extract the viscosity of any type of fluid, from Newtonian fluids, such as water, to nonNewtonian fluids, such as blood. Traditionally, the viscosity analysis of fluids has been carried out with huge, bulky, intricate and expensive equipment only available in laboratories and for whose use a complex training is required [6-8]. Viscometers are capable of study only Newtonian fluids and rheometers are adequate to analyze Newtonian and non-Newtonian fluids. To solve this situation, a full-custom electronic module has been implemented. The combination of the electronics with the implementation of an array of electrodes in the microfluidic channel allows extracting the speed of the fluid front. The extraction of the position and the speed are the input variables to extract the viscosity. The full set-up to validate the concept is based on a low pressure pump, the micro full custom device, depicted in Figure 2, which has been manufactured using soft lithography with PET (polyethylene terephthalate) and PDMS (polydimethylsiloxane). In order to extract the measurements, a user friendly interface for the control of the device has been also implemented, Figure 3, where some control parameters of each experiment can be introduced, the pressure introduced by the pressure pump is plotted and an indicator of the experiment is showing when the fluid gets in contact with any of the electrodes in the microdevice. Knowing the distance between electrodes, and measuring the time needed for the fluid to flow from one electrode to the following, the software computes the velocity of the fluid and with it the viscosity is obtained. The system has been validated with an optical setup, presented in Figure 4, and thanks to these experiences we have developed a patent for a portable micro-rheometer, working with small volume of sample (<50Âľl) [9]. Calibrated with Newtonian fluids, such as water and ethylene glycol at different concentrations, and tested with blood as a non-Newtonian fluid, we have observed the characteristic behavior of every fluid, as shown in Figure 5, and with blood we have tested that the analysis is safe for biological samples. From the results, we can say that our device is very precise since the standard deviation between values has been always under 0.034mPaÂˇs, and it is also accurate since the viscosity values obtained with the set-up differ from those in the literature by a 10%. References [1] Ochoa, B., Kruspe, T., & Goodbread, J. (2014, June). A New Sensor for Viscosity and Fluid Density Measurement for Oil Well Drilling Applications. InSensors and Measuring Systems 2014; 17. ITG/GMA Symposium; Proceedings of (pp. 1-6). VDE. [2] http://www.anton-paar.com/?eID=documentsDownload&document=10067&L=5 > @ .DĂĽ\V 5 5HNXYLHQĆĄ 5 9LVFRVLW\ DQG GHQVLW\ PHDVXUHPHQW PHWKRGV IRU SRO\PHU melts. Ultragarsas" Ultrasound", 66(4), 20-25. [4] Evans, P. A., Hawkins, K., Lawrence, M., Williams, R. L., Barrow, M. S., Thirumalai, N., & Williams, P. R. (2008). Rheometry and associated techniques for blood coagulation studies. Medical engineering & physics, 30(6), 671-679 [5] http://www.anton-paar.com/?eID=documentsDownload&document=20072&L=5

[6] http://www.malvern.com/en/products/product-range/kinexus-range/kinexus-pro-plus/default.aspx [7] http://www.anton-paar.com/no-en/products/group/rheometer/ [8] http://www.tainstruments.com/product.aspx?siteid=11&id=29&n=3 [9] Method, apparatus and micro-rheometer for measuring rheological properties of Newtonian and nonNewtonian fluids (No. EP 15382248.1) Figures

Figure 1 Lab-on-a-chip solution

Figure 2 Full set-up

Figure 3 Control software of the device

Figure 4 Validation with the optical microscope

Viscosity vs Shear rate 4

Viscosity (mPa路s)

3,5 3 2,5

Water

Et40%

1,5

Et25%

Et20%

0,5

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Synthesis of nanomaterials with multiple applications in Biomedicine Isabel Ortiz de Solórzano, Laura Usón, and Jesús Santamaría Institute of Nanoscience of Aragón (INA) and Department of Chemical Engineering and Environmental Technology. University of Zaragoza, c/ Mariano Esquillor, Edif I+D, 50018 Zaragoza, Spain Email: isaortiz@unizar.es

Nowadays, Biomedicine is one of the main investigation areas and different Nanotechnology-based approaches are helping in the development of new therapeutic and diagnosis tools applied in this field. Nanotechnology takes advantage of the specific properties of nanomaterials due to their reduced size. Surface atoms are responsible of different properties of nanomaterials including superparamagnetism, quantum confinement, plasmonics, catalytic, and so on. Some nanomaterials have special optical and electronic properties. They are capable to absorb irradiated light and convert it in thermal energy or to emit light at shorter wavelength than the excitation wavelength. Plasmonic Aubased nanoparticles, semiconductor Copper Sulfide, Carbon NanoDots and Rare Earth-based UpConversion Nanocrystals are examples of nanomaterials which show characteristic optical and thermal properties having an impressive potential in Optical Coherence and Photoacoustic Tomography, Photodynamic Therapy, NIR-triggered Hyperthermia, Bio imaging and Drug Delivery. On the other hand, magnetic nanomaterials like Iron oxide nanoparticles can be used in magnetic hyperthermia applications as a result of their ability to convert dissipated alternating magnetic energy into thermal energy[1] Moreover, these nanoparticles, as contrast agents, might play a key role in the development of In vivo Magnetic Resonance Imaging, which permits visualize molecular characteristics of physiological or pathological processes in a non-invasively way.[2] Apart from inorganic nanomaterials, polymeric nanoparticles and scaffolds are prominent in drug delivery and tissue engineering applications, respectively. Poly (lactic-co-glycolic acid) based nanoparticles have been extensively investigated because of their biocompatibility and biodegradability. They act as potential carriers for several classes of drugs being their targeting capability influenced by morphological characteristics, surface chemistry and molecular weight.[3] Mass production is a serious concern in the synthesis of all these nanostructured materials due to their large demand and, for that reason, continuous synthesis procedures are sought. When using discontinuous (batch) synthesis several drawbacks usually emerge such as variations in the physicochemical characteristics of the resulting products. In order to overcome these disadvantages microfluidic reactors, electrospinning and pyrolysis are synthesis techniques that have been used to control precisely temperature and residence times, rendering, in a continuous way, nanoparticles with narrow particle-size distributions.

References [1] Challa S.S.R Kumar, Faruq Mohammad, Magnetic nanomaterials for hyperthermia-based therapy and controlled drug delivery, Advanced Drug Delivery Reviews, Vol. 63, 9, (2011), 789-808. [2] Miriam Colombo, Susana Carregal-Romero, María F. Casula, Lucía Gutierrez, María P. Morales, Ingrid B. Böhm, Johannes T. Heverhagen, Davide Prosperi and Wolfgang J. Parak, Biological applications of magnetic nanoparticles, Chem. Soc. Rev., 41 ,(2012) 4306±4334. [3] Indu Bala, Sarita Hariharan, M.N.V. Ravi Kumar, PLGA Nanoparticles in Drug Delivery: The State of the Art, Critical Reviews in Therapeutic Drug Carrir Systems, Vol. 21, 5, (2004)

Cross-fertilization of Key Enabling Technologies. Insights from Nano-enabled Medical Devices. 1

Cristina Paez-Aviles , Esteve Juanola-Feliu , Josep Samitier

1,2,3

Department of Electronics, Bioelectronics and Nanobioengineering Research Group (SIC-BIO), University of Barcelona, Martí i Franquès 1, Planta 2, 08028 Barcelona, Spain. 2 IBEC-Institute for Bioengineering of Catalonia, Nanosystems Engineering for Biomedical Applications Research Group, Baldiri Reixac 10-12, 08028 Barcelona, Spain 3 CIBER-BBN-Biomedical Research Networking Center in Bioengineering, Biomaterials and Nanomedicine, María de Luna 11, Edificio CEEI, 50018 Zaragoza, Spain cpaezeviles@el.ub.edu

Abstract Technological convergence has become a current phenomenon that can generate products resulting from different technical backgrounds or industry sector [1], [2]. In this context, not only the convergence, but the coordinated effort of involving two or more disciplines could generate new product properties and technology features important to achieve social impact and technological progress. In this context, the cross-fertilization of Key Enabling Technologies (KETs) could improve the overall performance of the technological and biomedical systems. Current examples are the biocompatible materials, embedding reliable and targeted biosensors, high speed data communication, and even energy autonomy [3]. The convergence of rapid advances in areas such as microelectronics, microfluidics, microsensors and biocompatible materials allows the availability of cheaper and faster biodevices for continuous monitoring when compared with standard methods [4]. The concept cross-cutting KET is relatively new and was introduced by the European Commission. KETs are technologies that have a strategic importance to the future competitiveness and prosperity. Given this foregoing, since 2009, the EC have been stimulating a common strategy on behalf of the development of these technologies for their potential impact in strengthening Europe's industrial and innovation capacity [5]. At the healthcare domain, crosscutting KETs could improve the overall performance of the technological and biomedical systems given that the convergence of technologies in Nanotechnology, Biotechnology, Micro/Nano-electronics and Advanced Materials allow the development of new medical devices of small dimensions [6]. The aim of this study was to identify the principal actors and the market trends when developing nano-enable medical devices as the result of the integration of different KETs. For this purpose we selected 10 signed projects under Horizon 2020 Framework that were included in the category ³NMP 25±2014/2015: Accelerating the uptake of nanotechnologies, advanced materials or advanced manufacturing and processing technologies by SMEs´ We found in total 99 projects, from which 10 are related to nano-enabled medical devices development, 27 are health related projects, and 62 are related to other industries (construction, food, automobile, etc.). These projects were obtained from the dataset of the European Union Open Data Portal. This research engine provides information for each project, the specific programme, topic, title, start and end date, objectives, total cost, coordinators and participants. Market trends of these projects include the tissue engineering market, imaging diagnosis, orthopedic, prosthesis and dental implant market. We used the text mining tool VOSViewer for visualize this trends (Figure). The countries that are developing nano-enabled medical devices include Spain, Italy, Sweden, Netherlands and Ireland. Next steps in this research involve the study of the knowledge diversity background of each SME. The scope of this research encompasses the scientific practitioners and innovation managers in the strategies of managing and developing innovative devices. It also has a scope to companies, research organizations, and other organizations that are involved in the sector and that aim to foster the interdisciplinary integration of technologies and collaboration. The insights obtained from this research can be used for policy recommendations that can influence the process of future cross-fertilization of enabling technologies.

References [1] [2] [3]

[4] [5] [6]

S. Gauch and K. Blind, Technol. Forecast. Soc. Change, vol. 91, pp. 236Âą249, Feb. 2015. F. Harianto and M. Pennings, Res. Policy, vol. 23, pp. 293Âą304, 1993. J. Colomer-Farrarons, P. Miribel-Catala, E. Juanola-)HOLX DQG - 6DPLWLHU Âłin Novel Advances in Microsystems Technologies and Their Applications, L. Francis and K. Iniewski, Eds. CRC Press, 2013, pp. 497 Âą 534. E. Juanola-Feliu, P. L. Miribel-CatalĂ , C. P. AvilĂŠs, J. Colomer-Farrarons, M. GonzĂĄlezPiĂąero, and J. Samitier, Sensors (Basel)., vol. 14, no. 10, pp. 19275Âą306, Jan. 2014. ECSIP consortium, Copenhagen, 2013. C. Paez-Aviles, M. Gonzales-PiĂąero, and E. Juanola-Feliu, LAP Lambert Academic Publishing, 2015.

Figures

Figure: Visualization of principal domains form the H2020 selected projects developing nanoenable medical devices (VOSViewer).

Multifunctional, reusable, self-powered nano-bots for cleaning polluted water Jemish Parmar1, Diana Vilel2, Lluís Soler3 and Sámuel Sánchez1, 2, 4 1. Institute for Bioengineering of Catalonia (IBEC), Baldiri I Reixac 10-12, 08028 Barcelona, Spain. 2. Max-Planck Institute for Intelligent Systems, Heisenbergstr. 3, 70569 Stuttgart, Germany. 3. Institute of Energy Technologies, Universitat Politècnica de Catalunya, Diagonal 647, 08028 Barcelona, Catalonia, Spain. 4. Institució Catalana de Recerca i Estudis Avançats (ICREA), Psg. Lluís Companys, 23, 08010 Barcelona, Spain. jparmar@ibecbarcelona.eu Abstract Artificial nano-micromotors capable of performing multiple tasks while swimming in solution is a growing fascinating field with potential applications in the near future (1). Recent advances demonstrated dye and chemical warfare agent degradation, heavy metal and oil removal capabilities of nano/micromotors.(2-4) It is reported that Fe/Pt motors can efficiently degrade organic pollutant via Fenton oxidation reaction.(2) However, key features such as their lifetime, reusability and sizes of Fe/Pt motors, which would make the Fenton degradation process cost-effective were not studied. In this study, self-propelled Fe/Pt motors were fabricated by rolling up nanofilms of Fe/Pt evaporated on specifically designed square-shaped photoresist patterns. The motors were left swimming in the hydrogen peroxide containing polluted water for one hour to degrade the dye. They were collected from the solution using a magnet for the next cleaning cycle and re-used for over 12 times to degrade the dye, followed by UV-Vis spectrometer. Tubular motors can continuously swim for more than 24 hours and remain active even after 7 weeks storage. The iron leaching from the motors surface decreased 10 times after the first use and remains extremely low in subsequent uses, which could potentially reduce slug formation in the large-scale system in the future. This study could help building artificial swimmers containing a fast and efficient system for household and industrial wastewater treatment, which could replace the current bacterial system for sewage treatment. References [1] S. Sanchez, L. Soler, J. Katuri. Angew.Chem.Int.Edit. 2015, 54 (5), 1414-1444 [2] L. Soler, V. Magdanz, V. M. Fomin, S. Sanchez and O. G. Schmidt, ACS Nano, 2013, 7, 9611-9620. [3] L. Soler and S. Sanchez, Nanoscale, 2014, 6, 7175-7182. [4] W. Gao and J. Wang, ACS Nano, 2014, 8, 3170-3180.

An Instantaneous Portable Anemia Detection Device. 2

Jaime Punter Villagrasa , Joan Cid Vidal , Ángeles Ivón Rodríguez Villarreal , Jordi Colomer 1 1 Farrarons , Pere Miribel Català 1

D2In, Electronics Department Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona, Spain jpunter@el.ub.edu 2 Department of Hemotherapy and Hemostasis, CDB, IDIBAPS, Hospital Clínic, Villarroel 170, 08036 Barcelona, Spain 3 Centre de Recerca Matemàtica, Campus Bellaterra, UAB, Edifici C, 08193, Barcelona, Spain Abstract In this work is presented a new anemia low cost POC device concept, depicted in Figure 1.c, which works with a commercial sensor without the need on any kind of microfluidic platform, Figure 1.b, and a drop of whole blood, up to 50 µL. This device is based on the use of the Electrochemical Impedance Spectroscopy (EIS) and the electrical behavior of blood [1] in order to detect the haematocrit (HCT (%)) relation with impedance measurement. In order to detect this measurement a potentiostat 3-electrodes configuration, an oscillator and an rms-to-dc converter are implemented, Figure 1.a. Based on our previous study in [1], with a huge range of EIS analysis, with a detection in magnitude and phase of the response of the impedance detected in the electrodes, is defined the frequency of operation that defines the oscillator. The rms-to-dc converter extracts the Vrms measurement of the signal which is proportional to the HCT (%) in a DC voltage level Figure 2. For its validation, we analysed 24 consecutive blood samples from patients hospitalized at Hospital Clínic in Barcelona. We performed a complete blood count (CBC) of the blood samples with a haematology analyser, the Advia 2120 from Siemens AG, which reported the hemoglobin and hematocrit results as g/dL and percentage (%), respectively. We tested all samples with the prototype within 2 h of blood collection and CBC. As it is an instantaneous detector with a time response of several milliseconds, to evaluate system precision and accuracy, every whole blood sample was tested 5 times consecutively using fresh sensors and fresh sub-samples. Our simple low-cost POC approach has been designed, fixing then a frequency of operation for the hematocrit detection [2] where the response is more sensitive. This device is quite different from other solutions [3]-[5]. Instead of other solution like [6], which presents a subjective response that depends of the user interpretation, our approach presents a numerical response, and with a simple processing the device, working as an event-detector, is able to state if the user has a Critical, Severe, Moderate, very Moderate or absence of anemia without the need of a visual interpretation of the results. References [1] Punter-Villagrasa, J.; Cid, J.; Colomer-Farrarons, J.; Rodriguez-Villarreal, I.; Miribel-Catala, P.Ll. Towards an Anaemia Early Detection Device Based on 50µl Whole Blood Sample. IEEE Transactions on Biomedical Engineering 2014. DOI: 10.1109/TBME.2014.2364139. [2] Punter-Villagrasa, J.; Cid, Páez-Avilés, C.;J.; Rodriguez-Villarreal, I.; Juanola_feliu, E.; ColomerFarrarons, J.; Miribel-Catala, P.Ll. An Instanatneous Low Cost Point-of-Care Anemia Detection Device. Sensors 2015, 15(2), 4564-4577;http://dx.doi.org/10.3390/s150204564. [3] Pop, G.A.; Bisschops, L.L.; Iliev, B.; Struijk, P.C.; van der Hoeven, J.G.; Hoedemaekers, C.W. Online blood viscosity monitoring in vivo with a central venous catheter, using electrical impedance technique. Biosensors and bioelectronics 2013, 41, 595–601. [4] Pradhan, R.; Mitra, A.; Das, S. Impedimetric characterization of human blood using three-electrode based ECIS devices. J. of Electrical Bioimpedance 2012, 3, 12–19. [5] Ramaswamy, B.; Yin-Ting, T.Y.; Si-Yang, Z. Microfluidic device and system for point-of-care blood coagulation measurement based on electrical impedance sensing. Sensors and Actuators B: Chemical 2013, 180, 21–27. [6] Tyburski E, Gillespie S. Disposable platform provides visual and color-based point-of-care anemia self-testing. J. Clin. Invest. 2014;124(10):4387–4394.

Acknowledgment This work has been funded by project DADDi2 DADDi2 (Dispositivo Aut贸nomo Desechable para el Diagn贸stico Diag de la Diabetes, TEC-2013-48506 48506-C3-3-R) from the Spanish Ministry of Economy. Figures

Figure 1. POC device. a)Device Device prototype electronics ; b) Sensor and drop of bllod; c) Envisaged prototype. (Reproduced from [1] with kind permission permissio from IEEE Publishers and [2]) [2]

Figure 2. Measured output dc voltage (VRMS (mV)) mean value (n = 5) as a function of blood samples hematocrit (Hematocrit (%)) [2].

Dithranol-Loaded PLGA Microspheres Conjugated with a Hydrogel Patch: Potential Application for Treatment of Psoriasis Raha Rahbari Swansea University, United Kingdom

Controlled drug delivery devices are becoming increasingly important for development of modern therapeutics and new ways of drug administration. Poly lactic-co-glycolic acid (PLGA) polymer has been extensively studied as a drug delivery matrix in the quest to design and fabricate devices for drug delivery. This project aims to develop a medical product using a novel polymer encapsulated drug-delivery approach for treatment of the common skin condition, psoriasis. Combining anapproved generic drug dithranol, with a new microsphere encapsulation technology and a controlled release system, consisting of a hydrogel patch, we present a novel drug delivery system. This system offers significantly improved safety, convenient application and optimised and controlled delivery of conventional pharmaceuticals. It paves the way for potential smart application of other generic drugs for a wide range of skin conditions. Biodegradable PLGA microsphere encapsulants were fabricated using an emulsification solvent evaporation technique. Dithranol was encapsulated inside the PLGA microspheres. Mass spectrometry was used to qualitatively detect dithranol, within microspheres from its corresponding accurate mass (dithranol has a mass / charge ratio of 226.2). To assess encapsulation efficiency, UV spectroscopy was used to compare the original concentration of dithranol initially used to load the microspheres, to the amount of dithranol extracted from the fabricated microspheres. Results showed encapsulation efficiency of dithranol of 99 ± 0.002% within the PLGA microspheres. The PLGA microspheres had an average size distribution of 46μm diameter and released of 28.45 ± 5.38% of their total dithranol content in vitro over 24 hours. Viability assays and toxicology studies of the PLGA microspheres suggested that these carriers are non-toxic and safe to use for in future medical applications. The results of this study suggest that the use of dithranol-loaded microspheres, in conjugation with a hydrogel patch, is feasible as a novel drug delivery system. This approach would create new opportunities for customised release of pharmaceutical and active ingredients.

Figure: SEM images of dithranol-encapsulated PLGA-microspheres in combination with hydrogel (A) Magnification (500x), shows the microspheres clearly embedded in the hydrogel support matrix (B) Magnification (110x).

SP1, a Protein from Aspen Trees, as a Nanopore M. Akerman1,3, N. Attias2,3, L. Nesiel2,3, K. Liu1,3, A. Karmi1,3, Y. Nevo2,3, D. Rotem1,3, O. Shoseyov2,3, D. Porath1,3 1. Institute of Chemistry. 2. Institute of Plant Science. 3. Center for Nanoscience and Nanotechnology, The Hebrew University of Jerusalem, Israel rotem.dvir@mail.huji.ac.il Nanopores have been used as stochastic sensors for the detection of analytes that range from small molecules to DNA, RNA, and proteins. In this approach, individual analyte molecules modulate the ionic current flowing through a single nanopore. SP1 (stable protein 1) is a ring-shaped, highly stable homododecamer protein. It was originally isolated from Aspen trees (P. euphratica), where its level is altered in response to stress. SP1 is stable under extreme conditions (high temperature, protease resistance, wide pH range tolerance, stability to detergents and organic solvents). Its pore diameter is relatively large (3-4 nm) and it can also be manipulated to suit for detection of varied analytes (DNA, RNA, Proteins). It was recently shown that SP1 can be embedded into lipid bilayers, thus creating a nanopore. The protein can be modified in order to change the charge distribution on its surface to further increase its stability in the lipid bilayer; this could be done via site directed mutagenesis and/or chemical modifications to increase surface hydrophobicity. In parallel, we trapped SP1 protein on top of a solid state nanopore drilled in a Si membrane. This hybrid nanopore combines the advantage for both protein nanopores (that have atomically precise structure and the potential for genetic engineering) and the solid-state nanopores (that offer durability, size and shape control, and are also better suited for integration into wafer-scale devices).

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Low-noise Wideband pre-amplifier for fluorescence microscopy 1

Andreu Sanuy , Pablo Loza-Alvarez , Jordi Andilla , David Gascón 1

Univeristy of Barcelona, Av. Diagonal 645, Barcelona, Spain ICFO – The Institute of Photonic Sciences, Av. Carl Friedrich Gauss 3, Castelldefels, Spain

asanuy@ecm.ub.edu Abstract (Arial 10) A fluorescence microscope is an optical microscope that uses fluorescence instead of reflection to study properties of organic substances [1]. The specimen is illuminated with light of a specific wavelength which is absorbed by the fluorophores, causing them to emit light of longer wavelengths. Typical components of a fluorescence microscope are a laser light source, the excitation filter, the dichroic mirror, and the emission filter (see figure 1). The emission of light through the fluorescence process is nearly simultaneous with the absorption of the excitation light due to a relatively short time delay between photon absorption and emission, ranging usually less than a microsecond in duration. An unfortunate consequence of low emission levels in most fluorescence microscopy applications is that the number of photons that reach the eye or camera detector is also very low. In most cases, the collection efficiency of optical microscopes is less than 30 percent. In order to generate sufficient excitation light intensity to produce detectable emission, powerful light sources are necessary. They have become essential in scanning confocal microscopy, a technique that has proven to be a powerful tool in rendering very sharp fluorescence images through rejection of non-focused light removed from the specimen focal plane. Confocal microscopes accomplish this task through point or line scanning with coincident imaging through a conjugate aperture. Optical sections of the specimens can be stored in a host computer and reconstructed into the final image, which is then displayed on the monitor [2]. Confocal Laser Scanning Microscopy (LSM) is based on a conventional optical microscope in which instead of a lamp, a laser beam is focused onto the sample and an image is built up pixel-by-pixel by collecting the emitted photons, usually with a PMT. Thus, LSM combines point-by-point illumination with simultaneous point-by-point detection. The images of a mouse intestine section in Figure 2, noticeably illustrate the gain in resolution in confocal LSM imaging over conventional wide field imaging [3]. Two-photon excitation LSM can be a superior alternative to confocal microscopy due to its deeper tissue penetration, efficient light detection, and reduced phototoxicity. The high flux of excitation photons typically required is usually obtained using a femtosecond laser. One of the most common ultra-short pulse lasers is the Ti-sapphire laser that has typical pulse widths of approximately 150 femtoseconds and a repetition rate of about 80 MHz [4]. In order to get good resolution at the images reconstructed at the LSM observations, the size of these images becomes an important factor. The typical image size in a LSM observation is at least of 512 x 512 pixels so, several hundred thousand laser shots impact to the specimen to observe. In order to avoid undesired effects like photobleaching or phototoxicity, it is crucial to minimize as much as possible the laser power and pixel dwell time. In the other hand, demands on recordings of fast biological processes require fast acquisition speeds thus very high readout speeds. One of the limits in the readout is the bandwidth of the electronic circuitry, and especially when there is some averaging applied at each pixel samples, in order to reduce the photonic fluorescence noise. Most of the current electronic circuits used in the readout systems of fluorescence microscopy are low bandwidth systems (below 100 MHz). This could seems enough according to the common data acquisition (DAQ) systems used to store the pixel-by-pixel images that can operate up to 10 MS/s. Some techniques are used to “collect” multiple fluorescence pulses in order to do not loose information due to the DAQ sampling rate as integration circuits, that their output is proportional to the amount of the input signal during a period of time [5]. In this work we present the validation of a low-noise wideband pre-amplifier to improve the collection speed while maintaining the dynamic range of the acquisition [6].

Using the integration circuitry, the PMT of the fluorescence microscopes, can be read with low bandwidth pre-amplifiers. But, analyzing the images obtained, it can be appreciated a loos in the high frequencies at the FFT spectrum (see figure 3) whereas the same specimen illuminated at the same conditions by using a low-noise wideband amplifier of 500 MHz, with a dedicated integrator scheme, there is no loose at the FFT spectrum (see figure 4) References [1] Fluorescence microscope. Wikipedia, https://en.wikipedia.org/wiki/Fluorescence_microscope. [2] Guy Cox, “Biological confocal microscopy”, Material Today, 2002, pag 34 - 41. [3] Hellen C. Ishikawa-Ankerhold, “Advanced Fluorescense Microscopy Techniques – FRAP, FLIP, FLAP, FRET and FLIM”, Molecules, 2012, pag 4047 - 4132. [4] W Denk, “Two-photon laser scanning fluorescence microscopy”, Science, 1990, Vol. 248 no. 4951 pp. 73 - 76. [5] Op amp integrator. Wikipedia, https://en.wikipedia.org/wiki/Op_amp_integrator. [6] A. Sanuy, “Wideband (500 MHz) 16 bit dynamic range current mode PreAmplifier for the CTA cameras (PACTA)”, JINST, 2011, doi 10.1088-1748-0221/7/01/C01100. Figures

Figure 1. Confocal Laser Scanning Microscopy (LSM) block diagram scheme

Figure 2. Mouse intestine section image with Alexia Fluor 350 WGA, Alexa Fluor 568 phalloidin SYTOX Green

Figure 3. (Left) FFT spectrum of the image (Right) image zoom of the mouse intestine wall with 200kHz bandwidth readout system

Figure 4. (Left) FFT spectrum of the image (Right) image zoom of the mouse intestine wall with 500MHz bandwidth readout system

Mesoporous Nanowires as Carriers for Drug Delivery A. Serrà1, N. Gimeno1, E. Gómez1, M. Mora2, M. Ll. Sagristá2, E. Vallés1 1

Ge-CPN, Dept. Química Física, Institut de Nanociència i Nanotecnologia (IN2UB), Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona.

Dept. Bioquímica i Biologia Molecular (Facultat de Biologia), Universitat de Barcelona, Avda. Diagonal 643, 08028 Barcelona e.valles@ub.edu

Abstract Cancer treatment involving chemotherapy is typically accompanied by toxic side effects, thereby limiting the amount of the drug that can be given to a patient (all of the tumor tissue may not be exposed to a lethal dose of drug). The use of nanostructured materials as carriers could significantly improve the pharmacological properties of drugs by achieving their accumulation in the target tissue. The result will be an improved efficiency and reduced side effects [1-2]. Recently, mesoporous materials have demonstrated to be promising candidates for cell-specific delivery because of its high specific pore volume and surface area that allow reaching high drug loadings per weight unit. Moreover, recent developments with mesoporous nanostructures have highlighted its potential as drug carriers because of: i) its relatively straightforward inside-out tuning of the vehicles; ii) its high flexibility and iii) a large drug loading potential, allowing to coordinate drug and nanostructure concentrations and minimize the toxicity induced by the nanostructured carrier. Besides, their potential as sustained drug release systems makes these nanostructures very promising candidates for intelligent cancer therapy and other nanomedical applications [3-4]. On the other hand, magnetic nanomaterials have emerged as a new type of materials that could be used in cancer treatment. These materials allow performing a new type of therapy based on the mechanical destruction of cells, which causes the cell death [5]. Herein, we propose synthesizing magnetic mesoporous nanorods (100 nm in diameter and approximately 2 µm in length) to be used as nanocarriers, which combine the high surface area and high loading capability, with the possibility to control the release of the drug and to increase its efficiency by applying a magnetic field (mechanical destruction). Therefore, our study analyses the release rate of the drug and the mechanical destruction of cells. So the chemotherapeutic efficiency will be tested in stirring conditions of the nanocarrier and in non-stirring conditions. It is important to note that our proposal may reduce dramatically the amount of both the nanocarrier and the drug to obtain efficient results. In addition, mesoporous nanorods will be compared with compact nanorods.

Electrodeposition was used as a synthetic route for preparing mesoporous and compact nanorods using microemulsions with ionic liquids and aqueous solutions as electrochemical media, respectively. This proposal combines the use of a soft template (microemulsion) for pore definition, and a hard template (polycarbonate membranes) for nanorods definition, allowing the control of the length, the diameter and the pore size by controlling the deposition time, the hard template and the microemulsion structure, respectively [6]. Moreover, it is important to note that this method allows obtaining mesoporous nanorods of any metallic material by an easily and friendly procedure. After synthesis and washing, the magnetic nanostructures have been functionalized in order to provide them with an hydrophilic surface that minimizes the aggregation of the magnetic nanorods, that favors the interaction with the cells, and that, specially, promotes the drug retention (Figure 1). Finally, the usefulness of the prepared materials was also studied by using fluorimetry techniques. Now, we are testing the cytotoxic activity of our materials in HeLa cells (Human cervical adenocarcinoma cells, ATCC CCL-2).

References [1] N. Bertrand, Jun Wu, X. Xu, N. Kamaly, O. C. Farokhzad, Advanced Drug Delivery Reviews, 66 (2014) 2. [2] D.S. Spencer, A. S. Puranik, N. A. Peppas, Current Opinion in Chemical Engineering, 7 (2015) 84. [3] J. L. Vivero-Escoto, I. I. Slowing, B. G. Trewyn, V. S.-Y. Lin, Small, 6 (2010) 1952. [4] C. Argyo, V. Weiss, C. Bräuchle, T. Bein, Chemsitry of Materials, 26 (2014) 435. [5] D.-H. Kim, E. A. Rozhkova, I. V Ulasov, S. D. Bader, T. Rajh, M. S. Lesniak, V. Novosad, Nature materials, 9 (2010) 165. [6] A. Serrà, E. Gómez, E. Vallés, Electrochimica Acta, 174 (2015) 630.

Figures

Figure 1: Schematic representation of our proposal.

Controlled linear movement of nanoparticles in suspension by homogeneous magnetic field gradients 13 12 1 Carlos David Amaya Jaramillo , José Javier Serrano Olmedo , Milagros Ramos Gómez , Francisco 1 del Pozo Guerrero

1. Center for Biomedical Technology (CTB), Campus Montegancedo28223 Pozuelo de Alarcón, Madrid, Spain 2. Center for Biomedical Research Network -Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Av. Monforte de Lemos, 3-5. Pabellón 11. Planta 0, Madrid, Spain 3. Secretary of Higher Education, Science, Technology and Innovation (SENESCYT),Alpallana E7-123 Pasaje Martín Carrión Edificio "Caminosdel Parque" LOCAL No. 3, Quito, Ecuador carlos.amaya@ctb.upm.es Abstract The product of alternating gradient force and constant filed magnet is the main phenomenon to attract objects but, if the constant magnetic field and gradient magnetic are large enough it is possible to move magnetic nanoparticles and guide them in one directionand crash the MNP (magnetic nano-particle) against cells and produce cell death, this is possible due to the shape of Helmholtz coils and produce a gradient magnetic and control it by circulating current in the coils. The distance traveled by magnetic nanoparticles depends on: current and amount of nanoparticles clustered. We have designed and build an experimental set-up from a combination of a constant magnetic field and a time varying, homogeneous gradient magnetic field (Fig. 1). The higher the particle mass is the higher the linear momentum it acquires so that the mechanical interaction with cell (soft material) is more destructive. However, too large particles, for instance, equal or higher than the typical cell size, probably would cause only displacements of the cells, if they are free, since very large speeds are not easy to achieve inside a viscous fluid like water. On the opposite side, small particles could be too small if they simply punch the external cell membrane, or other inner cell membranes, causing the aperture of a hole, so small that the cell restores itself before any further damage is caused by, for instance, excessive interchange of fluids or ions through it. Then, our particles must be of size about or slightly higher than 1um that is the upper limit of nanoparticles used in Biolistic[1]. A large particle size is better because the magnetization is larger for the same magnetizing external field. Therefore, we suggest that particles from300 nm to 2µm. High magnetization can lead to the creation of clusters, filaments in shape, of mechanical characteristics depending on several conditions like the particle concentration, the viscosity of the media, the presence of other material in suspension and the strength of the magnetic field. In case of low concentration, the filaments are made of a few particles[2]. To control movement of MNP, we use a square current of 7A frequency 1 and 50 [Hz]in the coils (Fig. 2), and produce a magnetic field (Fig. 3) to move MNP in a concentration of 50 [µg/ml] from 30 to 60 [min]. This instrument could be useful to: cause cell death due to, damage caused by the movement of the MNPs interacting with cells; or treatment for thrombosis produced by the movement of the MNP to weaken the material that obstructs the blood flow. 132 1N1 cells were preincubated with different types of MNPs and introduced in the instrument described above to test the efficiency ofhomogeneous magnetic field gradient to promote cell death. Cell viability was assessed using the calcein and propidium iodide assay. (Fig. 4) Figure 4 and figure 5 show the resultsobtained by homogeneous magnetic field gradient application after treating 1321N1 and MC3T3-E1 cells with MNPs. The highest cell death rates were obtained preincubating cells with2Pm MNPs. References [1] B. R. Frame, H. Zhang, S. M. Cocciolone, L. V. Sidorenko, C. R. Dietrich, S. E. Pegg, S. Zhen, P. S. Schnable, and K. Wang,Vitr. Cell. Dev. Biol. - Plant, vol. 36, no. 1, ³3URGXFWLRQ of transgenic maize from bombarded type II callus: Effect of gold particle size and callus morphology on transformation HIILFLHQF\´, (2000), pages. 21±29. [2]

E. Zhang, M. F. Kircher, M. Koch, L. Eliasson, S. N. Goldberg, and E. Renström, ACS Nano, vol. 8, no. 4,³'\QDPLF 0DJQHWLF )LHOGV 5HPRWH-&RQWURO $SRSWRVLV YLD 1DQRSDUWLFOH 5RWDWLRQ´, Apr. (2014), pages. 3192±3201.

Figures

Front view Top View Figure 1: Helmholtz coil inside permanent magnetic field

Figure 3: Magnetic field produced by coils Figure 2: Gradient magnetic field produced by coils Control

1Hz

50Hz

Figure 4: Cell viability after magnetic field treatments. 132 1N1 cells were stained with propidium iodide to visualize dead cells (red) at different irradiation frequency. Scale bar 500Pm. A B

Figure 5:A. Viability of 1321N1 cells exposed to magnetic field treatments controlled by a current square of 7A frequency from 1 to 50 [Hz] for 60 min. B. Cell viability in MC3T3-E1 cell line after 30 and 60 min of exposure to 50Hz magnetic fields, as evaluated by calcein /propidium iodide assay

Methodology and evaluation of nanomechanical changes of endothelium in model of hyperglycemia: from in vitro to ex vivo experiments Marta Targosz-Korecka, Katarzyna Malek-Zietek, Magdalena Jaglarz, Marek Szymonski

Research Centre for Nanometer-Scale Science and Advanced Materials, NANOSAM, Faculty of Physics, Astronomy, and Applied Computer Science, Jagiellonian University, Krakow, Poland marta.targosz-korecka@uj.edu.pl

Most of the vascular complications in diabetes are related to hyperglycemia and result from the impairment of endothelium [1,2]. Endothelial cells (ECs) lining the lumen of every blood vessel are directly exposed to the action of high lucose concentration. Resulting endothelial dysfunction, can be cause changes of mechanical properties of ECs. In this work we used Atomic Force Spectroscopy to measure nanomechanical properties of ECs during hyperglycemia and after normalization of the glucose levels. ECs were exposed to short (5 min - 24 h) and long-term (1 day Âą 1 month) hyperglycemic conditions. It was observed that hyperglycemic conditions result in a significant stiffening of the ECs and that elastic changes are anti-correlated with NO production, that is the main parameter used for the description of ECs phenotype. For very short-term hyperglycemia (45 min), it was observed that changes in the nanomechanical properties of ECs are reversible, i.e. ECs return to their initial state after glucose normalization. For longer duration of hyperglycemia (up to 3h), ECs elasticity parameter slowly returns to the reference value, but for 24 h exposures the changes are already LUUHYHUVLEOH ZKLFK FDQ EH LQWHUSUHWHG DV WKH RQVHW RI ÂłVWLIIQHVV PHPRU\Â´ REVHUYHG IRU ORQJ WHUP hyperglycemia [3]. The studies carried out on the ECs in vitro model was verified ex vivo on the mouse diabetic model (db/db mouse model).

References [1] De Vriese A.S, Verbeuren T.J, Van de Voorde J, Lameire N.H, Vanhoutte P.M, Endothelial dysfunction in diabetes. British Journal of Pharmacology 130 (2000):963-974 [2] Brownlee M, Advanced protein glycosylation in diabetes and aging, Annu. Rev. Med, 46(1995):22334 [3] Targosz-.RUHFND 0 %U]H]LQND * 0DOHN . 6WÄ&#x160;SLHÄ&#x201D; ( 6]\PRQVNL 0 6WLIIQHVV PHPRU\ RI (A.hy926 endothelial cells in response to chronic hyperglycemia. Cardiovascular Diabetology 2013, 12:96 Acknowledgments: The research was supported by 32 ,* (8 SURMHFW Âł(ODVWLFLW\ SDUDPHWHU DQG VWUHQJWK RI FHOO WR FHOO interaction as a new marker oI HQGRWKHOLDO FHOO G\VIXQFWLRQ LQ K\SHUJO\FHPLD K\SRJO\FHPLDÂ´ program POMOST FNP.

Caco-2 cells as an in vitro model to determine detrimental effects on the intestinal barrier. Studies with SiO2- and ZnO-NPs at sub-toxic doses Laura Vila, Alba García, Ricard Marcos, Alba Hernández Grup de Mutagènesi, Departament de Genètica i de Microbiologia, Facultat de Biociències, Universitat Autònoma de Barcelona, Bellaterra, Spain; CIBER Epidemiología y Salud Pública, ISCIII, Spain. laura.vila@uab.es

Abstract Engineered nanoparticles (NPs) are used in many commercial products due to their desirable characteristics for many industrial applications. In addition, some of them are being used as additives in [1],[2],[3] food and packaging, increasing human exposure . This increased use of NPs and the lack of complete knowledge on their potential hazard in the gastrointestinal tract, as a main protection barrier, require further investigations. ZnO-NPs are already used in food packaging products due to their antimicrobial and UV-absorbent [4] properties . Furthermore, amorphous SiO2-NPs are used as food additive and as a component in milk [4] [5] [6] powders, instant soups, etc. to improve flow-ability . The human Caco-2 cell line is derived from colonic epithelial adenocarcinoma cells. This cell line has the capability to differentiate into small intestine enterocyte after reaching confluence, when it is grown under normal cell culture conditions. After 21 days of differentiation the cells become to be polarized [7] [8] and acquire tight junctions, microvilli and membrane transporters . This is considered a very useful model to observe uptake of nutrients and pharmaceuticals and, for these reasons, differentiated Caco-2 cells have become a model for in vitro studies related with the uptake and transport through barriers. Intestinal cell toxicity of ZnO and SiO2-NPs were evaluated in differentiated Caco-2 cells. Moreover, intestinal integrity and paracellular permeability were also evaluated after 24 hours of NPs incubation at sub-toxic doses in order to mimic a realistic environmental exposure. The nanomaterials were characterized for their morphology and size by TEM (Fig. 1) and DLS/LDV. The analysis of cytotoxicity show non-toxic effects of amorphous silica in differentiated Caco-2 cells, while ZnO-NPs led to significant reduction of Caco-2 cells viability. The use of sub-toxic doses of both NPs demonstrates that the integrity and permeability remain properly after 24 hours of exposure (Fig. 23). Hence, neither of these two NPs at sub-toxic doses were able to damage our system of differentiated Caco-2 model.

References [1] Borm PJA, Robbins D, Haubold S, Kuhlbusch T, Fissan H, Donaldson K, Schins RPF, Stone V, Kreyling W, Lademann J. Part Fibre Toxicol. 2006, 3:11. [2] Nel A, Xia T, Madler L, Li N. Science. 2006. 311:622-627. [3] Di Pasqua AJ, Sharma KK, Shi YL, Toms BB, Ouellette W, Dabrowiak JC, Asefa T. J Inorg Biochem. 2008. 102:1416-1423. [4] Chaudhry Q, Scotter M, Blackburn J, Ross B, Boxall A, Castle L, Aitken R, Watkins R. Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2008. 25:241-258. [5] Schmid K, Riediker M. Environ Sci Technol. 2008. 42:2253-2260. [6] Dekkers S, Krystek P, Peters RJ, Lankveld DX, Bokkers BG, van Hoeven-Arentzen PH, Bouwmeester H, Oomen Ag. Nanotoxicol. 2011. 5:393-405. [7] Chantret I, Barbat A, Dussaulx E, Brattain MG, Zweibaum A. Cancer Res. 1988. 48:1936-1942. [8] Chopra DP, Dombkowski AA, Stemmer PM, Parker GC. Stem Cells Dev. 2010. 19:131-142.

Figure 1. TEM images of SiO2-NPs (A) and ZnO-NPs (B) in dried form. A

Figure 2. Integrity and permeability analysis of SiO2-NPs. Integrity was evaluated using trans epithelial electrical resistance (TEER; A). Permeability was evaluated analyzing the concentration of paracellular compost named Lucifer Yellow (LY) in basolateral medium (B) A

Figure 3. Integrity and permeability analysis of ZnO-NPs. Integrity was evaluated using trans epithelial electrical resistance (TEER; A). Permeability was evaluated analyzing the concentration of paracellular compost named Lucifer Yellow (LY) in basolateral medium (B) A

Versatility of polymeric nanocapsules for the encapsulation of topical active ingredients of different physico-chemical nature 1*

Gemma Vilar, Jessica Romero, Lorena García-Fernández, Elisabet Fernández-Rosas, Jaume 1 2 1 Oliva, Ignacio Umbert, Socorro Vázquez-Campos. 1

Leitat Technological Center, C/ Pallars 179-185, 08005, Barcelona, Spain. Institute of Dermathology I. Umbert, Clínica Corachan, Plaza Manuel Corachán 4, 08017, Barcelona, Spain. *These authors have contributed equally in this work gvilar@leitat.org, svazquez@leitat.org

Abstract The physicochemical characteristics of active ingredients (AI) as well as the properties of vehicles are responsible for its differential absorption and distribution in the targeted organs/tissues. There is a need for the development of new strategies to increase the bioavailability of a number of AI for topical applications. One of these strategies includes formulation of AI within nanometric particulate carriers; as an example polymeric nanoparticles [1 have shown very promising results for topical use. They offer many advantages including a controlled- and sustained-release of the active ingredients, enhancement on their skin permeation [2], improvement of bioavailability and protection of the encapsulated AI from its environmental degradation [3]. The aim of this work was to develop safe, biodegradable nanocapsules (NCs) with high versatility and efficiency in the encapsulation of different topical active ingredients. To that aim, polymeric nanocapsules have been developed, in particular monodisperse polyester nanocapsules with an average size of 200 nm. This type of polymers are at the front line of attention because of their attractive safety profile. Since their degradation products are easily metabolized by the Krebs cycle and therefore easily eliminated. Furthermore, systemic toxicity associated with these nanoparticles for drug delivery is low [4]. Active compounds with different physicochemical nature were encapsulated inside of polymeric NCs. The encapsulation method for each AI was designed and optimized according to their different physicochemical properties. The active ingredients to be encapsulated were chosen for their use in antiaging or inflammatory skin disorders treatment and for their low solubility, limited skin penetration and/or their skin irritation potential. This is the case of Retinoic acid, Hydroquinone or Indomethacin. Both, the synthesis and purification processes were optimized and the amount of encapsulated AI was quantified by appropriate analytical methods. Depending on the physicochemical nature of the active ingredient, different encapsulation yields were obtained, but overall, they were more than 50%. Analytical characterization showed the efficiency of the encapsulation process. Different release profiles were obtained depending on the nature of the active and a sustained release was observed until 24h, after that, the release rate decreased. In vitro studies on skin cells have also been performed to demonstrate the biocompatibility and low cytotoxicity of the developed nanocapsules. In addition, a comparative study of the cytotoxic prolife of the encapsulated versus non-encapsulated AI was performed. Nanoencapsulated indomethacin shows a low cytotoxic response on skin cells with respect to direct application of the compound. Finally, the efficacy of the developed nanocapsules was evaluated by in vitro methods based on the analysis of the main biological activity of each active ingredient, showing promising results for the development of novel drug delivery systems for anti-aging or inflammatory disease treatments.

References [1] Zhang, Z., Tsai, P-C., Ramezanli, T., Michniak-Kohn, B.B., Wiley Interdiscip Rev Nanomed Nanobiotechnol., 5(3) (2013) 205. [2] Alvarez-Román, R., Naik, a., Kalia, Y. N., Guy, R. H. & Fessi, H. J. Control. Release 99 (2004) 53. [3] Cho, H. K., Jeong, S. H., Cho, D. C., Yeum, J. H., Cheong, I. W. Arch. Pharm. Res, 37 (2014) 423. [4] Anderson, J. M. & Shive, M. S., Adv. Drug Deliv. Rev., 64 (2012) 72.

Figures

A) TEM and SEM images of polymeric NCs and DLS and FT-IR spectra of polymeric NCs. B) Release assays of AI from polymer 1 (P1) and polymer 2 (P2) NCs. C) Cytotoxicity tests of the developed NC on skin cells: Comparative study of the cytotoxic prolife of the encapsulated AI versus non-encapsulated AI.D) Biocharaterization of the NC: Evaluation of the cell-availability of the AI (encapsulated and nonencapsulated) and in vitro efficacy studies. Microscopy images of application of non- and encapsulated Retinoic Acid on Human Dermal Fibroblast (HDF).

Nobel TiO2/Au fuel-free nanomotors based on active Brownian motion under visible light Varun Sridhar † , Xu Wang † , Samuel Sánchez † ‡ * †

Max Plank Institute for Intelligent Systems, Heisenbergstraße 3, 70563 Stuttgart, Germany ‡ Institut de Bioenginyeria de Catalunya (IBEC), Baldiri i Reixac 10-12, 08028 Barcelona, Spain *Institució Catalana de Recerca i Estudis Avancats (ICREA), Pg. Lluís Companys 23, 08010 Barcelona, Spain xuwang@is.mpg.de Nano- and micromotors are ultra-small devices designed to perform selected mechanical movements in response to specific stimuli. In addition, these nano- and micromotors offer extraordinary potential for environmental and biomedical applications [1]. Most of these devices have been mainly focused on chemically powered motors which are propelled by the catalytic decomposition of hydrogen peroxide by platinum. However, the requirement of the hydrogen peroxide fuel impedes many practical applications of such catalytically propelled micro and nanomotors, and has led to the exploration of alternative chemical fuels. Here, we present new nanomotors with hollow cap structures which are propelled under visible light [2]. The motors are fabricated by electron beam deposition of TiO2 and Au on soft templates. These nano caps have TiO2 on their concave layer and Au on their convex layer, displaying enhanced Brownian motion which is controlled with visible light under a wide field illumination. Visible light is used to activate the particles by surface plasma resonance induced electron transfer from Au to TiO2, thereby initiating the free radical formation on the surface of TiO2 which in turn propels the particles due to diffusiophoresis. This study could bring a promising ‘green’ system using natural fuel sources, such as sun light. Furthermore, these nanomotors open new doors for the fabrication of motors bio and environmental compatible and their use for the development of new applications in environmental and biomedical fields.

References 1. Soler, L. & Sanchez, S. Catalytic nanomotors for environmental monitoring and water remediation. Nanoscale, 6 (2014) 7175-7182. Guix, M., Mayorga-Martinez, C. C. & Merkoci A. Nano/Micromotors in (Bio)chemical Science Applications. Chem. Rev., 114 (2014) 6285–6322. 2. Hong, Y., Diaz, M., Córdova-Figueroa, U. M. & Sen, A. Light-Driven Titanium-Dioxide-Based Reversible Microfireworks and Micromotor/Micropump Systems. Adv. Funct. Mater., 20 (2010) 1568– 1576.

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