Vectorization of biologically active molecules for their selective delivery to target cells is a major objective since this strategy allows potentiating the activity of these molecules, but also limiting their side effects. In particular, selective addressing of antigens of vaccinal interest to dendritic cells represents an active field of investigation and is particularly promising for the development of new candidate vaccines against infectious agents or cancers. Indeed, the dendritic cells are the only cells of the innate immune system able to activate the naive T cells, which is indispensable to induce protective adaptive immune responses. According to their phenotype, ontogeny, localization and specialized functions, dendritic cells are divided into different subpopulations, including lymphoid, myeloid and plasmacytoid subsets, identified both in mice and humans. A large body of data shows that the level of adaptive immune responses, differentiation and specialization of CD4+ Th1, Th2, Th17, or regulatory T cells, as well as activation of CD8+ T cells are orchestrated by various sub-populations of dendritic cells. Therefore, the in vivo mobilization and activation of the latter by their direct targeting through specific surface markers, represents a critical way for the development of preventative and/or therapeutic vaccine candidates against cancers or infectious diseases. We have recently developed a vectorization technology to address one or more biologically active molecules, including antigens and/or adjuvants, in a highly versatile manner, to various target cells and in particular to dendritic cell sub-populations. Unlike other technologies developed to date, our strategy allows the addressing of biologically active peptidic or non-peptidic compounds to well-defined dendritic cell subsets. Indeed, these compounds can be polypeptides, sugars, lipids or oligonucleotides of large size. Application of this technology to vaccination will enable to deliver in a highly specific and well-controlled manner, necessary and sufficient amounts of antigens and adjuvants to the same sub-population of dendritic cells, selected as a function of the type of the adaptive immunity to be induced. This will allow inducing optimal and ultimately protective immune responses, without undesirable inflammatory responses, frequently related to administration of adjuvant. We have established the proof-of-concept and feasibility of this approach in different experimental models, ranging from synthesis and purification of vectorized molecules until in vivo induction of adaptive immune responses. All of these results gave rise to a European patent application in December 2009. Our current project aims to reinforce the results obtained, in order to substantiate claims of the patent and strengthen intellectual property protecting the developed technology. Furthermore, the objective of our project is the implementation of our technology for the development of effective therapeutic and/or prophylactic vaccines in priority areas in public health and economic level, such as adult pulmonary tuberculosis and cancers due to chronic Human Papillomavirus (HPV) infection. The present proposal will lead to a rapid valorization of this technology at the industrial level.

The cardiovascular diseases are among most serious health problems of modern human society. Despite various advanced therapeutic, pharmaceutic and surgical methods of their treatment, significant improvement of human health has not been reached yet due to insufficient efficiency of these methods, side effects, soreness and long recovery. Hence, searching for novel approaches for both diagnostics and treatment of the diseases is of highest importance. This proposal will explore a new approach based on combining latest achievements in laser- and nano-technologies. Contamination-free multi-component nanoparticles with novel properties (efficient light absorption at specific wavelength ranges, enhanced photoluminescence for detection, minimal cytotoxicity and biocompatibility, etc.) will be synthesized by ultrafast laser irradiation of materials in aqueous environment. They will be tested as nanotools for treating actions on biological objects (model systems provided by biologists) for dissolving thrombus and cholesterol and for fatigued muscles recovery. The nanoparticles will be injected to the model systems (simulating problematic blood vessels and overexpressed muscles) and activated by ultrafast lasers tunable in mid-IR spectral range to make laser action gentle but effective with a deep penetration to bio-tissues. Irradiation will be performed in single- and multicolor modes to inspect capabilities for nanoparticle activation. The results of nano-phototherapy will be analyzed with assessment of its efficiency. The detailed protocols of nano-phototherapy will be prepared to continue these studies with biologists in-vivo after the project implementation. Outcomes of this proposal will bring new knowledge on the prospects of non-invasive therapy of cardiovascular problems that will have a positive impact on both fundamental science and bio-medicine in the European area. Implementation of this research will enable me to become a recognized expert in the field.

A major challenge in biomolecular research is the investigation of biomolecular interaction at single-molecule level. Biomolecules possess heterogeneities of high physiological relevance that can only be unravelled with single-molecule tools. All current single-molecule imaging methods, however, require chemical modifications, such as fluorescent labelling or immobilization onto a surface, which might alter the biomolecule’s natural behaviour. Recently, I have developed a ground-breaking optical microscopy method – Nanofluidic Scattering Microscopy (NSM) – whose unprecedented resolution enabled me to bypass those limitations and to image individual small proteins in free motion without any label. Despite these attractive attributes, in its current form, NSM does not allow for quantitative study of interaction kinetics between individual molecules. Due to the inherently fast Brownian motion, the time that a molecule spends on average in the optically probed volume – a nanofluidic channel – is substantially shorter than the time required to record a statistically relevant number of association and dissociation events. In this project, we will develop an essential nanoscopic component – a rapid nanofluidic valve – that will enable to confine and release interacting biomolecules to and from the nanofluidic volume. The nanofluidic valves will be based on the principles of thermo-responsive polymer hydrogel in combination with nanoplasmonic heating. The integration of the nanofluidic valves with NSM will enable to track evolution of individual biomolecules at single molecule level, without the need of chemical modifications and in conditions that mimic an in-vivo state. The project will deliver a unique bioanalytical tool that will make key contributions to the fundamental understanding of biomolecular interactions, which is needed in basic research as well as in the pharmaceutical industry.

The project will investigate the current-induced vibrational heating and cooling of molecular junctions with relatively sharp molecular resonances. These systems are interesting because, depending on the atomistic details of the metal/molecule interface and the chemical structure of the molecule, an extremely rich variety of heating/cooling dynamics under an external bias is possible. A detailed study of the connection between the electronic structure (both at equilibrium and in presence of a bias) and the inelastic processes associated to the emission and absorption of molecular vibrations represents a necessary step toward the comprehension and control of the junction heating and cooling dynamics and thus of its stability. We will study a broad range of systems where the sharp DOS features are originated by two different physical mechanisms: i) structural and chemical details of the metal/molecule interface, and ii) destructive interference in the molecule. In the first case, we will focus on the effect of electrode shape on the heating and cooling of the molecule and consider a wide range of molecule-electrode couplings. The molecules we will consider are examples of these classes of systems, namely carbenes (strong coupling to the electrodes), bypiridine (intermediate coupling), PTCDA (weak coupling). In the case of destructive interference, we will focus on conjugated linear molecules where quantum interference features originate from the coupling of side-groups to the molecular backbone. Finally the project will extend the state-of-the-art approach for the calculation of the heating and cooling dynamics by introducing self-consistency between vibrations populations and the electronic structure. For this we will make use of a series of approximations in the derivation of the equations for vibration emission and absorption rates. We expect this new approach will reveal complex dynamics in systems with sharp resonances and close to vibrational instabilities.