Biohybrid nanomaterials: bioinspired, DNA- and peptide-based assemblies for sensing, delivery and electronics

Resume : The study of complex molecular networks and supramolecular assemblies is a clear objective of the field so-called systems chemistry, which is expected to have a great impact in the area of origins-of-life research and as biohybrid materials in materials science. With regards to the origins of life, a pertinent question is whether artificial cells could be constructed from non-natural components. In order to provide clues about this question, we have started research lines on nucleic acid hybrids, replicating nucleopeptide networks and nucleolipid compartments. The study and combination of these components is an interesting approach because it allows exploring some properties of life without the restrictions of the historical pathway that Darwinian evolution took. Concerning the approach to biohybrid materials, we focus our work on supramolecular biohybrids for biomedical light management, which combine different photoactive molecules with peptide, protein and nucleic acid nanostructures. For example, we have recently studied the electrostatic co-crystallization of a cationic porphyrinoid and negatively charged tobacco mosaic virus (TMV), together with the capacity of the resulting crystals to photogenerate reactive oxygen species. In a different approach, electrostatically assembled zinc Pc-DNA origami complexes have been demonstrated. In this presentation we will shortly discuss some of these research lines.

Resume : Translation of genetic information into functional products (proteins) underpins all living systems, and is key to the process of evolution by natural selection. However, nature is limited to a small palette of building blocks (amino acids) - developing the capability to translate genetic codes into completely artificial, perfectly sequence-defined polymers would give access to many exciting new technologies, including the development of non-natural polymers that match and extend the functionalities of peptides and proteins and even nanomachines capable of programmed product synthesis in situ. On a more fundamental level, we could recreate complex biological behaviours such as gene regulation, bringing the creation of artificial living systems—the grand challenge of synthetic biology—a step closer. I will describe our efforts to use DNA nanotechnology and DNA-templated synthesis to develop such an 'artificial ribosome', focusing on two key barriers to progress and how we are addressing them. I will discuss how we have explored performing translation in solvents other than water to prevent degradation of fragile building blocks during synthesis, and our more recent attempts to activate building blocks in situ using DNA-templated catalysts. I will also discuss the new mechanisms we are investigating that will allow nanomachines made from DNA to sense when chemical reactions have happened, enabling great increases in efficiency. By combining these advances, we hope to make significant progress towards recreating one of nature's most complex molecular machines.

Resume : Modulation of temperature within biological systems can provide precise control over system dynamics. Plasmonic gold nanoparticles (AuNPs) have the innate capability to generate heat when excited with an external light source and can readily be conjugated with biological materials, making them suitable heat sources for a wide variety of biological applications. Recently, it has been demonstrated that femtosecond (fs)-pulsed lasers can be used to excite AuNPs and generate heat profiles that are confined to the immediate nanoscale region around the AuNP, allowing for local heating without increasing the bulk temperature of the system. When using the proper AuNP size and laser fluence, the local temperature increase can be turned so that it is pertinent to biological activity, yet, it is not yet known the level of precision and control afforded through this technique and little work has been done to implement confined nanoscale heat sources into complex biological systems. Here, we use 55 nm AuNPs in conjugation with moderate pulse fluences (1-14 J/m2) to increase the local temperature by (10-90°C) and monitor the temperature sensitive responses of various biological materials attached the surface of the AuNP. We use the denaturation of double-stranded deoxyribonucleic acid (ds-DNA), the kinetics of the enzyme alkaline phosphatase (AP), and the kinetics of the thermostable enzyme alcohol dehydrogenase (AdhD) as model temperature sensitive biological materials to evaluate the level of spatiotemporal control fs-pulsed heating of AuNPs can provide over biological activity.

Resume : The controlled organization of individual molecules and nanostructures with nanoscale accuracy is key for the fabrication of the next generation optoelectronic devices and the investigation of events in biological and chemical assays. In this regard, we controlled the assembly of different biomolecules, from proteins to DNA and aptamer sequences, on carbon nanotubes (CNTs), so as to couple CNTs electronic output to biomolecular function. In particular, we will report on the fabrication of reconfigurable and solution processable nanoscale biosensing devices with multisensing capability based on SWCNTs functionalized with specific, and different, aptamer sequences employed as selective recognition elements. Multiplexed detection of three different biomarkers indicative of stress and neuro-trauma conditions was successfully performed, and real-time detection was achieved in serum down to physiologically relevant concentrations.[1] We will then discuss the extension of this rationale into the development of real-time biosensors with engineered protein interfacing. We assembled different variants of a B-lactamase (BL) inhibitory protein (BLIP) onto SWCNT sidewalls in electronic device configurations with different, and controlled, protein orientations. We recorded the current responses in real-time for the detection of different concentrations of class BL enzymes,[2] that degrade antibiotics, in the context of investigating antimicrobial resistance. This allowed us to investigate the influence of the protein orientation in binding and biosensing device response/performance. [1] Nano Letters, 2018, 18, 4130-4135 [2] submitted, 2020

Resume : The lighting industry growth is fueled by the need of replacing old-fashion and inefficient bulbs by more efficient white inorganic light-emitting diodes (WILED). Even though WILEDs rule the artificial lighting market, drawbacks related to their sustainable development (use of inorganic phosphors based on toxic and rare-earth elements)1 and health concerns (eye photodamage and effects on human circadian rhythms)1 have placed hybrid light-emitting diode (HLED) in the spotlight. They consist of blue chips coated by an organic phosphor, such as polymers, quantum dots, dyes and, more recently, fluorescent proteins (FPs).2-3 FP-based bio-phosphors have led to bio-hybrid light-emitting diodes (Bio-HLED) that stand out combining best device performance and sustainable and eco-friendly emitters. First Bio-HLEDs achieved moderate efficiencies (50 lm/W) and stabilities (100 h) due to thermal- and photo-degradations of FPs.3 A new Bio-HLED design shielding the FP surface with a hydrophilic polymer allowing their integration into the network of a light-guiding and host polymer enhances device efficiency (~130 lm/W) and stability (>150 days).4 The control of the mechanical, thermal, and optical features in this bio-phosphor is paramount towards highly stable and efficient FP-phosphor for biogenic lighting systems. 1. US Department of Energy: Manufacturing Roadmap Solid-State Lighting Research and Development 2014. 2. V. Fernández-Luna et al. Angew.Chem. Int.Ed. 2018, 57,8826 –8836 3. M. D. Weber et al. Adv. Mater. 2015, 27, 5493–5498 (2015.; Patent WO2016203028A1 (2016) 4. A. Espasa et al. Nat. Comm 2020 ( in press); Patent WO2019115525A1 (2019)

Resume : Scattering-type scanning near-field optical microscopy (s-SNOM) has become one of the key techniques to overcome the ubiquitous diffraction limit of standard, far-field optical spectroscopy and microscopy. This AFM-based technology exploits the strong confinement of light at the end of a sharp, metallic AFM tip to generate a nanoscale optical hotspot at the sample directly below the tip. Importantly, the amplitude and phase of the light within the optical hotspot is strongly influenced by the dielectric properties (e.g. absorptivity) of the sample directly below the tip. Phase and amplitude-resolved detection of the back-scattered light as function of position can therefore be used to extract the local optical properties at the sample surface with <10 nanometer precision. The development of Fourier transform infrared spectroscopy on the nanoscale (nano-FTIR) can now be used to routinely perform nano-FTIR spectroscopy and hence allow chemical identification of materials on the nanometer length scale. This enables the identification of individual biomolecules and analyze complex bio and bio-inspired nanostructures and self-assembled monolayers. In addition, DNA-origami structures, biominerals (bone), embedded structural phases in polymers or functional semiconductor nanostructures can now directly be visualized and characterized on the sub-10nm scale. Even further, hyperspectral imaging has pushed this technology to the next level, recording nano-FTIR spectra at each pixel of a larger image on the sample. Subsequent data analysis enables the direct extraction of optical images at specific optical frequencies with 10 nanometer spatial resolution.

Resume : The prospect of finding an improved method for rapid human-genome sequencing might offer an entirely new way of preventive health care [1]. Recently solid-state nanopores/nanogaps have generated a lot of interest for ultrafast DNA sequencing. However, there are challenges to slow down the DNA translocation process to achieve a single-nucleobase resolution. A series of computational tools have been used in an attempt to study the DNA translocations in several model systems [2-4]. We have also studied the performance of a boron-carbide (BC3) based nanogap setup for DNA sequencing using the density functional theory and non-equilibrium Green’s function-based methods. The current variations under applied bias voltages due to change in the nucleotides orientation and lateral position found to be significant for nucleotides. In the case of BC3 nanogap, unique identification of all four nucleotides possible in the 0.3-0.4 V bias region. Furthermore, each of the four nucleotides exhibits around one order of current difference, which makes it possible to identify these nucleotides uniquely. Thus, our BC3 based nanogap may be promising for rapid human DNA sequencing. References: [1] B. Pathak, H. Löfås, J. Prasongkit, A. Grigoriev, R. Ahuja, R. H. Scheicher, Appl. Phys. Lett. 100 (2012), 023701. [2] J. Prasongkit, A. Grigoriev, B. Pathak, R. Ahuja, R. H. Scheicher, Nano Lett. 11 (2011), 1941- 1945. [3] J. Prasongkit, A. Grigoriev, B. Pathak, R. Ahuja, R. H. Scheicher, Journal of Physical Chemistry C. 117 (2013), 15421-15428. [4] R. L. Kumawat, P. Garg, S. Kumar, B. Pathak, ACS Applied Materials and Interfaces, 11 (2019), 219-225.

Resume : Going beyond the traditional concept of electronic devices, we convey the idea of making electronics edible. This unconventional approach exploits the electronic properties of natural and food-based materials for developing ingestible functional devices. Critical biomedical, pharmaceutical, and food industry applications are targeted by the proposed field. In this framework, we explore the potential of cost-effective and edible substance, honey, to be used as electrolytic gate viscous dielectric. Honey-gated organic field effect transistors (OFETs) based on both n & p type semiconductors are fabricated. A distinctive feature of these transistors is their long-term stability, reproducibility and low voltage < 1V operation in air. Devices exhibit forward-looking electronic performances, notably, electron and hole mobility–capacitance product of 3.5 × 10–3 μF/Vs and 23 × 10–3 μF/Vs, respectively, surpassing ones of the previously reported water-gated OFETs. Furthermore, the observed devices responsivity to humidity provides promising opportunities for sensing applications. We then demonstrate, for the first time, the implementation of honey-based integrated circuits: inverting logic gate and ring oscillator.

Resume : The sensing of biomarkers has been regarded as a powerful method for early clinic diagnosis, which is crucial for the treatment of a large number of diseases.1 However, developing an ultrasensitive and highly selective method to detect and quantify these biomarkers is still challenging, due to their generally low concentrations, small sizes and easy degradation.2 In this project, we propose to combine SERS (surface-enhanced Raman scattering)-based nanoassemblies with Dielectrophoresis (DEP) to design an ultrasensitive biosensor. The target molecules will induce the formation of Au nanoparticle (AuNP) dimers with small gaps (the hot-spot regions) where the Raman process of Raman reporters is enhanced, and then we utilize a DEP trap at the tip of nanopipette to preconcentrate AuNP dimers to get stronger Raman signals and lower limits of detection. In addition to high sensitivity and selectivity, this technique is also highly adaptable. By changing the aptamers attached on AuNPs, this biosensor will be able to detect various biomolecules that can be specifically recognized, which will lead to a more accurate diagnosis. Furthermore, this method is low-cost and highly responsive, which makes it suitable for point-of-care clinic diagnosis. References 1 E. Stern, A. Vacic, N. K. Rajan, J. M. Criscione, J. Park, B. R. Ilic, D. J. Mooney, Nat. Nanotech, 2010, 5, 138–142. 2 S. Li, L. Xu, W. Ma, X. Wu, M. Sun, H. Kuang, L. Wang, N. A. Kotov and C. Xu, J. Am. Chem. Soc., 2016, 138, 306–312.

Resume : We present an innovative method to create an original nanostructured SPR biosensor in order to enhance sensitivity and limit of detection. Our strategy consists in taking advantages of high refractive dielectric materials which increase the sensitivity factor of a gold thin film. The approach consists in using iron oxide nanoparticles assembled onto a gold thin film to improve the sensitivity factor of the gold thin film as well as the specific surface of the sensor. The nanoparticles were easily grafted on the substrate by catalyzed alkyne azide cycloaddition (CuAAC) “click”. The biomolecular receptor was grafted at the surface of assembled nanoparticles by performing again the CuAAC “click” chemistry. Furthermore, polyethylene glycol (PEG) derivatives were grafted onto free areas of the gold thin film in between nanoparticles in order to avoid nonspecific adsorption of analytes. The biotin-streptavidin couple was used as an example to prove the efficiency of the sensor. Bovine serum albumin (BSA) was also used to demonstrate that nonspecific absorption is avoided. Finally, we successfully demonstrate that PEG molecules associated with nanoparticles enhance the accessibility of streptavidin to the biotin group, thus contributing to decrease the limit of detection.

Resume : The hydrogel is a three-dimensional cross-linked hydrophilic polymer with multiple advantages such as high biocompatibility, biodegradability, high water content and controllable drug release rate. Also, the aptamer is a either DNA or RNA nucleotides with affinities comparable to antibodies as well as high specificity and affinity towards the target with nanomolar to picomolar range. In this presentation, we will report DNA aptamer-incorporated hydrogels whose macroscopic volume is changed through an inducement of the binding events of aptamer-target recognition. We synthesized multi-arm PEG based hydrogels and potassium cation aptamers so that both can be covalently attached. By adding potassium cation, the macroscopic volume shrinkage of hydrogels is achieved due to the formation of the G-quadruplex structure between the linear DNA aptamers and potassium cations, leading to the decreased microscopic volume. This stimuli-responsive hydrogel will be applied to sustained drug delivery systems.

Resume : One of the most promising developments in the nanomedecine field is the design of smart activable nanosystems remotely releasing drugs upon externally applied stimuli (e.g., light, magnetic or electric fields). Among the various activable nanomaterials, carbon-based nanocomposites are still few investigated for remotely controlled drug delivery. In this work, we address the design of original carbon nanotube base composite nanoplatforms endowed with phototherapy combined with a drug release mediated by NIR laser excitation. These responsive carbon nanotubes are surrounded with a mesoporous silica shell having small pores which are chemically modified with isobutyramide (IBAM) grafts. These IBAM binders allow first the loading of an antitumor drug doxorubicin with a very high drug loading capacity (≥to 80%) followed by the adsorption of an additional tight human serum albumin (HSA) shell ensuring a biocompatible interface and drug gate keeping. Such smart photoresponsive platforms are shown to deliver the drugs upon several pulsatile NIR excitations with controlled T profiles according to the conditions used (concentrations, power laser etc…) These nanosystems are in fine integrated within a hydrogel mimicking/resembling the extracellular matrix and the biological response with cells on this resulting smart nanocomposite hydrogel scaffold is assessed upon NIR light irradiation. Such nanocomposites are hence highly promising as new components of implantable scaffolds by interfacing biomolecules with carbon nanotubes, which can respond in time and location to external stimuli for a better disease management.

Resume : The detection of carbon monoxide (CO) is of high significance as it binds to haemoglobin that restricts the transport of oxygen in body. Over the recent years, carbon monoxide has gained greater attention in therapeutic treatments due to its effectiveness in anti-inflammation and anti-microbial infection. The concentration of CO in these treatments needs to be controlled to prevent immediate and chronic poisoning. This research focuses on using an ultrasensitive nanopore-based probe to sense CO, to achieve a limit of detection (LOD) that is smaller than other conventional sensing techniques by orders of magnitude. A molecular ruthenium complex that connects a 10 kbp DNA and a 5 kbp DNA is employed for nanopore translocations. The CO molecules are administrated by CO-releasing molecules into the electrolytes. The binding of CO to the ruthenium metal centre cleaves the pyridyl-terminated 10 kbp DNA unit, which results in a significant change in the detected current signals. The concentration of CO is determined by the translocation frequency of CO-cleaved DNA strands (5kpb), and the LOD can be specified and then compared with that obtained by fluorescence-based sensors.

Resume : Oral vaccination for cancer immunotherapy provides an improved patient compliance and induction of mucosal immune response as well as humoral immune response. However, the delivery of antigen by oral route is limited due to harsh gastro-intestinal environment, intestinal epithelial barrier and lower uptake of the antigenic particles. In this study, we prepared an improved oral immunotherpy platform for simultaneous delivery of antigen and adjuvant using polymeric carrier (DA3). Encapsulation of adjuvant and antigen (DA3/Ad/Ag) was successfully carried out by formulating with DA3, which effectively protects antigen from enzymatic degradation and enhances colloidal stability. Permeation of DA3/Ad/Ag nanoparticles was increased by specific interaction with deoxycholic acid and apical sodium-dependent bile acid transporter (ASBT). In addition, their enhanced cellular uptake was observed in antigen presenting cells (APCs), which leads to successful maturation of immune cells and secretion of proinflammatory cytokines. Orally administrated DA3/Ad/Ag nanoparticles showed an enhanced permeation and induction of humoral, cellular and mucosal immune responses, which leads to desired cancer therapeutic effect. These results suggested that DA3-based oral vaccination platform for simultaneous co-delivery of antigen and adjuvant can be considered as a prospective candidate for cancer immunotherapy.

Resume : To boost the efficacy of chemo-photothermal cancer treatment by the intracellular rapid release of doxorubicin (DOX) combined with near-infrared (NIR)-triggered photothermal effect of indocyanine green (ICG), the poly(r-glutamic acid)-g-poly(lactic-co-glycolic acid) (r-PGA-g-PLGA)-based polymeric nanoassemblies (PNAs) with simultaneous incorporation of DOX and ICG were developed in this work by simple one-step nanoprecipitation. The obtained DOX/ICG-loaded PNAs were characterized by a compact r-PGA/DOX complexes-encapsulated PLGA-rich core covered with the co-assembly of amphiphilic D-α-tocopheryl polyethylene glycol succinate (TPGS), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)] (DSPE-PEG) and ICG molecules. The robust cargo-loaded PNAs not only promoted the photo-stability of ICG in PBS, but also reduced ICG leakage from PNAs. With the milieu pH being altered from 7.4 to 5.0, the massive disruption of ionic DOX/r-GA complexes and PLGA degradation considerably accelerated DOX release from payload-containing PNAs. The results of in vitro cellular uptake revealed that the DOX/ICG-loaded PNAs appreciably enhanced the cellular uptake of payloads by HeLa cells. Notably, through the intracellular acid-triggered rapid DOX release combined with the ICG-based NIR-activated hyperthermia and singlet oxygen generation, the combination therapy of DOX/ICG-loaded PNAs can use lower dosage of drugs to effectively inhibit proliferation of HeLa cells compared to the chemo or photothermal treatment alone, thus showing the great potential to improve efficacy of cancer treatment.

Resume : A well-known interest of using nanoparticles to deliver drugs is the controlled delivery of the contained therapeutic agent followed by internal or external stimuli. But these nanosystems are even more interesting if they combine several modalities. In our group, such combination has been used to obtain imaging and magnetic hyperthermia. The system consisted in an iron oxide nanoparticle encapsulated in a stellate mesoporous silica shell (IO@STMS). The shell was then functionalized with quantum dots and coated with Human Serum Albumin (HSA)[1]. The HSA was fixed through the IBAM strategy, demonstrated to allow the non-covalent but stable binding of several types of macromolecules to silica particles such as protein, nucleic acid, polysaccharide and polypeptide[2]. The aim of our work is to modify the previous IO@STMS system to deliver interesting proteins for tissue regeneration. To do so, it is hypothesized that the combination of this system with thermo-responsive (bio)polymers will allow the release of the protein through magnetic hyperthermia-induced conformational change. The IBAM strategy would still be used either to fix the new coating or to ensure protein adsorption. [1] F. Perton, M. Tasso, G. A. Muñoz Medina, M. Ménard, C. Blanco-Andujar, E. Portiansky, M. B. Fernández van Raap, D. Bégin, F. Meyer, S. Begin-Colin, D. Mertz, Applied Materials Today 2019, 16, 301-314 [2] D. Mertz, P. Tan, Y. Wang, T. K. Goh, A. Blencowe, F. Caruso, Adv. Mater. 2011, 23, 5668-5673

Resume : Biological systems, where weak non-covalent interactions of simple building blocks form functional complex machinery, gave rise to biologically inspired self-assembling materials, based for example on nucleic acids and peptides. Although self-organized systems based on molecules from both families have already been explored, artificially synthesized hybrid molecules are novel and exhibit noticeable importance in material science and biomedical research. In our system, two short complementary DNA segments are attached to an amphiphilic peptide sequence previously investigated by our lab1-3. We demonstrate the self-assembly of our system into a number of different topologies, fibers and multi-lamellar spherical structures, and we present the co-assembly pathway leading from one type of the aggregates to another. In addition, we find that when forming the onion-like structure, the conjugates exhibit conformational changes of DNA hybridization. We studied the stability of these spheres and the effect of metal ions on the DNA hybridization under different environmental conditions. To the best of our knowledge, this study proposes the first systematic analysis of structural and functional characteristics of small double stranded nucleic-acid-peptide conjugates. References: 1. B. Rubinov, N. Wagner, M. Matmor, O. Regev, N. Ashkenasy and G. Ashkenasy, ACS nano, 2012, 6, 7893-7901. 2. D. Ivnitski, M. Amit, O. Silberbush, Y. Atsmon-Raz, J. Nanda, R. Cohen-Luria, Y. Miller, G. Ashkenzsy, N. Ashkenasy, Angew. Chem. Int. Ed. 2016, 55, 9988-9992. 3. J. Nanda, B. Rubinov, D. Ivnitski, R Mukherjee, E. Shtelman, Y. Motro, Y. Miller, N. Wagner, R.C.Luria, G. Ashkenzsy, Nature Commun. 2017,8,434. 4. A. Chotera, H. Sadihov, R. Cohen‐Luria, P. Monnard, and G. Ashkenasy, Chem. Eur. J., 2018, 24, 10128-10135

Resume : α-Chymotrypsin uses three cooperative amino acids constitutive of its catalytic triad, serine (Ser), histidine (His) and aspartate (Asp) to cleave the peptide bond through covalent catalysis. The mechanism differs from the uncatalyzed hydrolysis of the amide bond, a challenge when designing protease mimics. Despite the limited success of the DNA catalyzed hydrolysis, DNA is however an ideal scaffold due to its chemical stability, its ease of access by solid phase synthesis (SPS) and its perfectly controlled folding. Moreover, grafting additional organic functionalities would expand its catalytic repertoire. Using the convertible and/or functionalized phosphoramidite approaches, we aim to mimic the active site of this serine protease with functionalized oligonucleotides (FuON) covalently modified at precise coordinates along their backbone with amino acids side chain-like residues (alcohol, imidazole or aspartate function for Ser, His or Asp respectively) (1). From a toolbox of twelve C5'-modifed thymidine phosphoramidites (varying both linkers and functions), families of FuON bearing up to three modifications could be obtained by SPS. They are then shaped into DNA secondary structures (3-way junction, bulges, hairpins) in an attempt to reproduce the active conformation of the triad. FuON combinations would then lead to heterogeneous libraries with skeletal and topological variety increasing the chances to screen for catalytic mimics. The first DNA catalysts libraries would be presented alongside the efforts to overcome the lack of a binding site. Their future insertion into 2D and 3D DNA nanostructures will also be discussed. (1) Addamiano, C.; Gerland, B.; Payrastre, C.; Escudier, J. M.; Molecules 2016, 21, 1082

Resume : Atherosclerosis is the major cause of cardiovascular disease including causing heart attacks, ischaemic heart disease and strokes. It is a chronic inflammatory disease that is described by the retention of foam cells, formed from oxidized low-density lipoproteins (LDL), building up the plaque and as a result narrowing or possibly blocking arteries. It is also possible to cause rupture when the fibrous cap is punctured which leads to ischaemic heart disease and strokes. With the recent advancements in nanotechnology, nanogels targeting and sensitive to the plaque will be investigated and developed. Nanogels are hydrogel nanoparticles with the properties of both hydrogel and nanomaterials. Nanogels offer size, higher loading capacity and stability advantages as a drug deliverer over common biological drugs. They can also be modified to be sensitive to stimuli, allowing a trigger for the release of the payload. In this work, nanogels will be modified to be responsive to matrix metalloproteinase and to incorporate anti-oxidizing paraoxanase-1 enzyme as the payload to prevent oxidation of LDL. In collaboration with Dr Ramzi Khamis group from Hammersmith Hospital, they have identified developed monoclonal antibody IgG3? (LO1). This antibody acts as targeting ligand towards malondialdehyde-conjugated LDL found in copper oxidized LDL. Therefore, this work demonstrates the potential of the combination of targeting, response and release as strategy to atherosclerosis therapy.

Resume : Nature has the amazing ability to achieve a perfect control over the monomer sequence in biological macromolecules. This microstructural precision leads to sophisticated structures with remarkable functions such as catalysis, molecular recognition, and information storage. Inspired by the precision of biological macromolecules, the design of synthetic sequence-defined polymers constitutes an emerging topic to generate materials with next-generation performances. Despite considerable progress on the control over the primary structure, the controlled folding and assembly of synthetic polymers into predetermined three-dimensional shapes have not been achieved so far. Here we use computational chemistry approaches to decipher the structure and dynamics of sequence-defined oligomeric chains used for supported catalysis. These oligomers incorporate structuring groups and three functional moieties for the selective oxidation of alcohols. The oligomeric chains under study differ by the monomer sequence and the number of molecular spacers between the catalytic moieties. In combination with the experimental catalytic activity, our computational study brings insights into structure-activity relationships for targeted applications in catalysis.

Resume : Peptoids represent an emergent class of synthetic polymers differing from peptides by the side chain position on the nitrogen atom instead the alpha carbon to amide moiety. Peptoids can adopt secondary structures in solution such as helix. However, unlike peptides and proteins, no data concerning the gas phase structure of peptoid ions is to date present in the literature. In our laboratory, we combine ion mobility mass spectrometry (IMMS) and computational chemistry to provide data on the 3D structures adopted by gaseous ions. Nevertheless, the ionization/desolvation processes related to the use of MS can generate huge structural modifications. Thus, the conservation of helical structure during the transfer from the solution to the gas phase represents a great challenge since most of the time, a charge solvation effect is observed during the transfer to the gas phase. Currently, a specific attention is given to design original peptoids with a localised charge to avoid possible charge solvation effect and potentially adopt helical structure in gas phase. Its backbone possesses (S)-phenylethyl side chains (Nspe), a bulky chiral group known to form helix in solution, and a (S)-N-(1-carboxy-2-phenylethyle) (Nspc) at C terminus. This sequence could optimize the interaction between the negative charge located on the C terminus and the helix macrodipole. Moreover, the influence of Nspc position and acetylation are investigated.

Resume : An alternative method for cancer therapy, which has attracted a significant attention in the past few years is magnetic hyperthermia. In the presence of RF magnetic field, the magnetic nanoparticles generate heat, which increase the temperature in tumors in a controlled manner, leading to killing the tumor cells. The interaction between the nanoparticles and the biological system depends almost by the surface modification of magnetic nanoparticles. In this context, cysteine coating, not only enhances colloidal stability, but also makes nanoparticles more biocompatible. Moreover, surface functionalization of magnetic nanoparticles with antitumoral will increase significantly the chances in the cancer treatment by hyperthermia. In this study, we report the synthesis of superparamagnetic nanoparticles (SPION NPs) with different surface modification, as well as their influence in the hyperthermia study. Biological tests on A375 human melanoma cells and metastatic murine melanoma (B16F10) confirmed the internalization of magnetic nanoparticles delivering Doxorubicin, which is used as a chemotherapic in the treatment of cancer. Also, IC50 value was determined for both cell types. Based on the obtained results, new perspectives on development of new biocompatible and bio-functional SPION NPs for magnetic hyperthermia are highlighted.

Resume : Small molecules that interact with DNA are of interest in the pursuit of functional DNA-templated self-assemblies.[1] In particular, tetraphenylethene (TPE) compounds have recently attracted a growing interest in this context thanks to their Aggregation-Induced Emission (AIE) effect that can be exploited for turn-on sensing applications.[2] After a previous work on imidazolium-TPE derivative for G-quadruplexes recognition,[3] we wish to explore the generation of new TPE derivatives by acylhydrazone conjugation. After a successful preparation of TPE derivative displaying four quaternary ammoniums, we evaluated its self-assembly by fluorescence and UV-visible spectroscopies. AIE effect was evidenced and correlated to nanoparticles formation as observed by Dynamic Light Scattering (DLS) and Transmission Electronic Microscopy (TEM). Further DNA recognition studies, carried out by UV-visible, fluorescence and Circular Dichroism (CD) spectroscopy analyses, clearly demonstrate the interaction with ssDNA and dsDNA and point toward a model involving intercalation that will be presented in this poster.[4] References: [1] Surin M., Polym. Chem., 2016, 7, 4137 – 4150 [2] Kwok R. T. K. et al, 2015, Chem. Soc. Rev., 44, 4228 – 4238 [3] Kotras C. et al, Front. Chem., 2019, 7, 493 [4] Coste M. et al., 2020, manuscript in preparation

Resume : Mammoplasty is a plastic surgery to reshape or modify human breasts through silicone implants. It is one of the most common surgeries in women around the world because is used in mammary reconstruction as a post-treatment of breast cancer, as well as for aesthetic reasons. Like any surgery, augmentation mammoplasty involves inherent risks such as bruising, rupture or emptying of implants, capsular contracture, and bacterial infections. To reduce these post-operative risks, medicaments and biomolecules can be localized and controlled release by drug delivery systems based in cyclodextrins (CDs). CDs have been used as efficient delivery carriers due to their characteristic cavity and their ability to form drug-reversible complexes. In that sense, the aim of this project is the functionalization of commercial breast implants of silicone, with smooth and roughness surfaces, by β-CD/citric acid and 2-hydroxypropyl-β-CD/acid citric in situ polymerization as an alternative for the post-operative risks treatment in mammoplasty. Functionalized samples were characterized by different physicochemical techniques, and its stability was studied. Five different molecules were used as model to test the obtained samples as controlled local drug delivery system for mammoplasty: zafirlukast, utilized for capsular contracture prevention, levobupivacaine for local anesthesia, rose Bengal that possesses cytotoxic properties in cancer cells, and KR-12 and GL13K antimicrobial peptides to prevent bacterial infections. In this regard, the release kinetic mechanism was obtained by the Korsmeyer–Peppas model. Further works will be focused on antibacterial tests using Staphylococcus aureus and Staphylococcus epidermis, two of the most common finded strains in post-mammoplasty bacterial infection.

Resume : Antimicrobial resistance to antibiotics is one of the biggest threats for health worldwide. This process occurs naturally but it is accelerated by the incorrect usage of antibacterial treatments. In developing countries this problem is aggravated due to socioeconomic and behavioural factors such as poor drug quality, unhygienic conditions and wrong use of antibiotics. Sensitive diagnostic platforms are usually very resource demanding since they require specialised infrastructures and equipment as well as expert technicians. The aim of this project is the design of a low cost, fast, sensitive, portable and user-friendly device that could be used in both developed and developing countries for the distinction between bacterial and viral infections. To this end, a paper-based analytic tool known as Lateral Flow Immunoassays (LFIAs). Semiconducting Polymer Nanoparticles (SPNs) synthesized from F8BT will emit a fluorescent signal in the presence of two biomarkers highly expressed in bacterial and viral infections, C-reactive protein (CRP) and Procalcitonin (PCT). The use of SNPs is expected to increase sensitivity in two orders of magnitude compared with traditional nanoparticle colorimetric LFIAs. Different targeting approaches will be explored and a smartphone device for the analysis of the strips will be engineered. This project aims to increase sensitivity whilst maintaining low-cost and user-friendly properties of LFIAs in the battle against antibiotic resistance.

Resume : Reconstruction of critical bone size defect needs specific material to fill the lost and to promote neo-osteoformation. To be perfectly integrated, and able to favorize bone tissue regeneration at the site of implantation, such substitute must possess combined bio-properties to mediate an efficient dialogue with the surrounding cells. Specific sequences derived from bone sialoprotein and bone morphogenetic protein, - as RDG like and BMP-2 like peptides – as emerged as interesting protagonists to act as proadhesive and osteocompetent partners able to induced an appropriate cell bone response. These peptidic sequences, inspired from natural protein, have attracted much attention, especially due to their high structure stability and their absence of immunogenicity. Commercial bioglass and natural bone purified by BIOBank, appears as interesting materials to serve as primary scaffold to design bone substitute. While the grafts generated from natural bone appear to be the best alternative to repair human bone defects thanks to their natural structure and properties; bioglass exhibited also interesting specificities, due to their capacity to release ions during their decomposition and generate locally apatite like matter. In this work, different peptides has been synthetized, characterized and grafted onto the two materials. Osteocompetent cells contacting responses are analyzed, following various parameters, as the synergistic effect of the co-adsorption of the two peptides.

Resume : Recently, there have been increasing interests in mesoporous silica especially in the field of controlled drug delivery. It is increasingly important to control the degree of bioactivity and rate of biodegradation of such mesoporous silica. In this study, manufactured the microporous(2nm~50nm) silica Nanorods by perfectly filling the TEOS solution using the pumping system in porous anodic alumina oxide(AAO)-the porous scaffold-with the most widely used metal alkoxide, Tetraethylorthosilicate(TEOS-si based), and their drug-delivery performance were investigated.In addition, in order to use in intraocular drug delivery system, paramagnetic mesoporous nanoparticles having SPION (superparamagnetic iron oxide nanoparticle) cores were also fabricated by coating SPION nanoparticles with the same silica precursor sol which was used for the fabrication of the mesoporous silica nanorods.SPIONs can achieve the highest drug targeting efficiency, an external magnetic field locally applied to the target.The porosity and biodegradation kinetics of the fabricated mesoporous nanorods and magnetic nanoparticles were analyzed using SEM, TEM, XRD, FTIR. The performance of the mesoporous silica nanorods as drug delivery carrier and intraocular drug delivery efficiency of the fabricated magnetic nanoparticles were evaluated.

Resume : Many researchers are studying efficient gene transfer methods to improve cell function and treat diseases.Retrovirus is the most commonly used vector for gene transmission, and if it does not develop an immune response, it will be incorporated into the gene and released in a stable manner. However, the problem is that it is injected only into the cells that are being split, and the diffusion rate is slow indicating a low transfer rate. Therefore, nonviral vectors, including cationic polymers, liposomes, dendrimers, and inorganic materials, attract significant attention in spite of their low efficacy.But because of the low transmission efficiency, there is a continuous effort to modify and refine these gene delivery systems either by adopting different strategies or by installing various functional attributes in the existing systems.Efforts to use polymeric materials are among them. In this study, in order to overcome the low gene transfer efficiency, a problem with retrovirus vectors, the transmission efficiency was improved by removing the reaction force to target cells using a functional silane – based platform.Three types of composite platforms were prepared by coating a complex surface layer on a substrate using precursor solutions synthesized from functional silanes. Triethylorthosilicate (TEOS), polydimethyl siloxane (PDMS), 3-glycidoxypropyltrimethoxysilane (GPTS) and 3-aminopropyltriethoxysilane (APTS) were used for the platform due to their biocompatibility and strong chemical transfection ability.Acid catalyzed sol-gel reaction in the mixed binder system was used for the synthesis varying the ratio of functional silanes.After transfusion of retrovirus vectors, cell number and trypan blue exclusion method were used to analyze cell survival.Using FACscan (Becton Dickinson, Franklin Lakes, NJ, USA), the ratio of eGFP positive cells was measured and compared with the gene transfer efficiency.

Resume : Fighting microbial biofilms on biomaterials is usually addressed by incorporating antimicrobial agents. Nevertheless, as usual in the natural life, intrinsic properties of the material surface can be also a complementary approach. They may drastically reduce the quantity of adhered microorganisms, that are thus to be further treated by antimicrobial agents. Mechanical properties of material surfaces recently emerged as a possible way to impact biofilm formation. However, many questions have not been elucidated so far. We have especially studied whether hydrogel and non-hydrogel soft and stiff films may impact biofilm formation and microbial behavior in a similar or different way. Microbial mobility, adhered quantity and production of organelles such as pili have been specifically investigated. The surface properties, especially mechanical ones, have been thoroughly characterized. The study has been conducted with yeast (Candida albicans) and bacteria species (Escherichia coli). It demonstrated significant differences of microbial behavior and response between films of similar elasticity but different hydration beside impacts of softness versus stiffness properties on amount and mobility of the adhered cells. This confirm relevance of using some soft coatings to prevent biofilm formation on a material but also clarifies the risk to get opposite effects as desired.

Resume : Fluorescence-based assays should be feasible in aqueous media for effectively detecting the biological factors. However, numerous sensors have limited signal transductions and low fluorescence quantum yields due to the inherently reduced excited state energy of fluorophores in aqueous solution, which reduces their sensitivity. This necessitates a new smart sensing platform with an amplified fluorescence response for analytes in aqueous solution. Herein, a new building block which self-assembles in aqueous media, giving the hydrophobic π-extended conjugated system at the core and hydrophilic groups at the periphery, was devised for the first time. We demonstrated that the self-assembly of conjugated units in a micelle provides a channel for efficient energy or electron migration and enhances the optical properties, thereby distinguishing this sensing platform from the numerous fluorescence-based tools previously developed for sensitive detection. This new system affords exceedingly sensitive detection of heparin via amplified fluorescence quenching, even under actual bioanalytical conditions.

Resume : Telomeres, the end parts of the chromosomes, contain repeated 6-nucleotide sequences (TTAGGG)n able to form so called G-quadruplex structures (G4) based on the stacks of guanine quartets. G4s are important cell regulatory elements whose detection is a challenging task. Their pi-electron systems can be studied by optics spectroscopy [1]. Specific optical properties of telomeric DNA (absorption, autofluorescence, autophosphorescence) and their environment-dependent changes allow using it as auto-luminescent sensing biomaterial. Here we have studied the spectral properties of telomere fragment d[AGGG(TTAGGG)3] (Tel22). Optical absorption (at 300K), fluorescence and phosphorescence (at 78K) of Tel22 samples folded into the G4 structure and unfolded by heating to various temperatures in the range 323-363K were investigated. The band at 295 nm associated with G-quadruplexes was observed in optical absorption of all samples. Only the band associated with G4 was observed in fluorescence spectra of the folded sample and the samples heated below 333K; no G4-associated fluorescence band was observed for samples heated above 343K. Phosphorescence of all samples was the emission of complex formed by A and T bases [1]. These effects are associated with thermal defolding of G4. [1]. V.M. Yashchuk, V.Yu. Kudrya, I.Ya. Dubey, K. I.Kovalyuk, O.I. Batsmanova, V.I. Mel’nik, G.V. Klishevich. Luminescence of telomeric fragments of DNA macromolecule. Mol. Cryst. Liq. Cryst. 2016, V. 639, p. 1-9.

Resume : Food freshness can be assessed by monitoring the level of biogenic amines in the food product. Electrochemical sensors that could detect early onset of spoilage (food freshness sensors) can reduce food waste for supermarkets and consumers. In this respect, carbon-based screen printed electrodes (C-SPE) have been functionalized with three different metallo-porphyrins (Co, Mn and Zn) and their response to several common biogenic amines was recorded via cyclic voltammetry. Standard samples of putrescine, cadaverine and histamine and mixtures of these biogenic amines were prepared and electrochemically tested with the fabricated sensors. Real-world samples (commercially available meat samples) were also used for the extraction of mixtures of the biogenic amines and tested in the same conditions as the standard samples. The C-SPE modified with metallo-porphyrins showed increased selectivity towards histamine as well as a decrease in the oxidation potential due to an improved electron transfer between the molecular complex trichloroacetic acid-histamine and the metallo-porphyrins. This interaction was modelled using HyperChem® software and the activation energy for each metallo-porphyrin was estimated. The limit of detection for histamine has been calculated as 0.7 ppm. Keywords: histamine detection, porphyrin functionalization, screen printed electrode, electrochemistry, food freshness. Acknowledgements: Core Program PN-2019-OPTRONICA VI.; Institutional Performance Programme-2019.

Resume : Nowadays a great attention is payed to regenerative medicine and tissue engineering. This discipline is based on the use of a bioactive 3D biomaterial for helping cells to grow and organize themselves in space like it is the case in organs. A logical trend is to develop biodegradable materials with a controlled porosity in order to exhibit a high surface reactivity with the biological environment. In this study, a 3D printer was combined to an electrohydrodynamic system to form a wet direct-writing technique able to produce a functional scaffold. This resorbable and hybrid nanostructure was realized from a polymer solution containing bioactive molecules and nanoparticles to regenerate both cement and bone from damaged dental tissues. The talk will first present the optimization of the solution properties (with or without) the proteins and the hydroxyapatite (HA) nanoparticles. Then, different designs of scaffolds produced and studied will be detailed before ending with results related to the bioactivity of these hybrid nanostructures. The whole work shows that cells cultured with the scaffold are able to differentiate and proliferate in specific cell types, depending on their contact with proteins (released from the scaffold) or with HA nanoparticles.

Resume : Our study aims to gather the maximum amount of information on the changes produced at the structural level (rotation, torsion, conversion) during the Cis-Trans transition (isomerization reaction) on one hand, and to examine the influence of different dopants or substituents on these kinetic parameters on the other hand. The results obtained through the optimization of molecules gave us the different distances and angles according to the methods and bases applied with a C2v symmetry. We were able to determine the total energies, the energy gap ΔE (HOMO-LUMO) of the two conformers Cis and Trans (the HF and DFT at 6-31G and 3-21G** levels) and finally a comprehensive analysis on the topological charges. The analysis of the results show that for the six molecules, the Trans conformer is energetically very stable compared to the Cis one, this stability is confirmed by the obtained values for the total energy ETOTAL. The increase in the stability energy leads to a less important HOMO-LUMO energy gap. The analysis of the optimized geometrical parameters of the six molecules using the AM1 and PM6 methods, are in agreement with the experimental structures characterized by X-ray diffraction. Finally, we were able to determine the reaction profiles of the Cis-Trans isomerization reactions of the polyacetylene in the gas phase, and to calculate the activation energy (Ea), as well as the diagrams of energies E (eV) based on the coordinates of the isomerization reaction of its molecules. All calculations performed in this study are carried out using the Hyperchem software. Keywords: substituted polyacetylene , semi empirical, HF (Ab-initio), DFT (B3LYP)

Resume : Electroporation induced by the “point discharge” effect is an effective technique for bacteria inactivation. Rapidly monitoring the electroporation-induced inactivation process is important for screening nanomaterials with high antimicrobial performance. In this study, we develop a facile strategy to in situ monitor the electroporation induced antimicrobial mechanism based on the surface-enhanced Raman scattering (SERS) effect of the Au-nanotip arrays. Owning to the high local-electric field (∼107 V m−1) generated on the Au nanotips, the bacteria are rapidly electroporated and effectively inactivated with ≥99.9% reduction in bacteria colony counts by only applying an external voltage of +0.8 V for 10 s. The related inactivation mechanism is directly verified by the formation of the Prussian blue (PB) nanocrystals by leaking of the uptaken [Fe(CN)6]3− ions from the cleavage area on the cell membrane. These [Fe(CN)6]3− ions react with Fe2+ to form PB nanocrystals onsite as soon as they leak out. The characteristic peak of PB in the cellular Raman-silent region provides a collective monitoring approach for the destruction of microorganisms. The present strategy not only develops a facial method for future use in evaluating electroporation materials, but also paves a rapid way for offering accurate information on some antibacterial and antitumor processes.

Resume : Robust and accurate detection of pathogenic bacteria in water and food is an important issue. Escherichia coli (E. coli) O157:H7 is an important cause of waterborne and foodborne illness leads to serious symptoms in humans. According the estimation of World Health Organization (WHO), 1.5 million children lose their lives because of diarrheal diseases ever year, which 88% caused by polluted water with E.coli. So in order to avoid the toxic effects of the pathogen, highly sensitive and selective identification should be done. In this study, a new, rapid, sensitive nucleic acid and gold nanoparticle based approach is established for the monitoring of E.coli, using a specific sequence of E.coli thiolated probe immobilized on the surface of gold electrode. Square wave voltammetry is carried out in the solution of toluidine blue for the cDNA hybridization and targeting. To investigate the morphology of the electrode and particle size estimation, Field scanning electron microscope (FE-SEM) is applied. The geno-assay selectively differentiates the complementary sequence from target sequences with one, double and three base mismatch sequences and detects the target DNA with sensitivity and selectivity.

Resume : Self-assembling peptides (SAPs) are a promising material for regenerative medicine as they can easily be chemically functionalized and show a multitude of different morphologies that are determined by the peptide sequence. Nanofiber-forming peptides can mimic essential features of the extracellular matrix (ECM) such as its fibrous morphology, mechanical properties, and chemical cues, which makes them interesting as scaffolds for tissue engineering. A gradual distribution of biological signals provides an essential role in guiding tissue orientation and function such as during embryogenesis as well as in cell-migration. Therefore, biomimetic scaffolds capable of imitating biomolecular gradients may be essential for tissue regeneration. Here, we present two approaches of forming a bioactive gradient in order to mimic the function of the ECM. First, by incorporating a photocleavable amino acid into the SAP, we can generate bioactive gradients on 2D substrates. Second, a pH-switchable depsi peptide is grafted to a polypeptide backbone forming a hybrid material. Under physiological conditions, the grafted peptides undergo a molecular rearrangement that induces nanofiber formation and subsequent gelation of the bulk material. By co-assembly of angiogenesis promoting peptides to this hybrid material we can create a gradual distribution of the bioactive peptides within a 3D space.

Resume : We have studied the surface of hydroxyapatite (HAP) coatings enriched with Mg and either Si or Ti deposited by RF magnetron sputtering on Ti6Al4V. This alloy, commonly used as material for orthopedic prostheses, was coated with HAP in order to improve biocompatibility and bioactivity of the material surface. HAP coatings have been furtherly functionalized by adsorption of a self-assembling peptide (SAP) on the HAP surface, with the aim of increasing the material bioactivity. The selected SAP is a self-complementary oligopeptide able to generate extended ordered structures by self-assembling in watery solutions. Samples were prepared by incubation of the HAP coatings in SAP solutions and subsequently analyzed by X-ray Photoelectron Spectroscopy (XPS), Fourier Transform Infra Red (FTIR) and Near Edge X-ray Absorption Fine Structure (NEXAFS) spectroscopies, in order to determine the amount of adsorbed peptide, the peptide stability and the structure of the peptide overlayer on the HAP coatings as a function of the HAP substrate and of the pH of the mother SAP solution. Experimental data yield evidence of successful immobilization of the SAP on the HAP surface; peptide overlayers show ordered structure and molecular orientation. The thickness of the SAP overlayer depends on the composition of the HAP coating.

Resume : Two-dimensional nanomaterials are emerging as a promising new class of materials due to their unique physical, chemical, optical and electronic properties, that make them suitable for a diverse range of applications. The current challenge is to develop a methodology that allows the orderer and controlled self-assembly of 2D nanomaterials from a single monomer. In this project we report a new approach to achieve two-dimensonal materials with hierarchical organisation using cyclic peptides as supramolecular building blocks. These peptides are able to form giant and dynamic nanosheets by sequencial 1D to 2D self-assembly. The primary amino acid sequence encodes the information for the adoption of the beta sheet motif, that arranged into amphiphilic nanotubes which consequently are self-assembled into 2D supramolecular nanosheets. We investigate the structure-assembly relationships of this innovative strategy and its functionalization.

Resume : A highly promising approach to influence and control the photophysical properties of conjugated polymers is directing their molecular conformation by templating. We report here the templating effect of single-stranded DNA oligomers (ssDNAs) on cationic polythiophenes. By means of electrostatic and other non-covalent interactions, the ssDNAs assemble with the polymer, directing its backbone to different conformations depending on the sequence of nucleic bases. We have comprehensively characterized the optical behaviour and structure of the polythiophenes in conformationally distinct complexes and addressed the effect on the ultrafast excited-state relaxation. This, in combination with molecular dynamics simulations, allowed us an unprecedented atomistic-level understanding of the structure-property correlations. We identify that optimal templating is achieved with (ideally 10-20) consecutive cytosine bases, having numerous π-stacking interactions with the thiophene rings and side groups of the polymer. Moreover, we show what specific side chains are necessary to force the polymer backbone into a rigid assembly with ssDNA, with highly ordered chains and unique optical signatures. Our insights are an important step forward in a pioneering approach to structural templating and optoelectronic control of conjugated polymers and organic materials in general.