The CHLOR2NOU project aims to develop new monitoring tools for CLD and its TPs, to provide new knowledge on the fate and risk of CLD TPs, and to explore realistic alternative approaches for pollution remediation. The postulate of the non-degradability of CLDs commonly admitted for several decades has had a strong negative impact on pollution management by ruling out the possibility of CLD degradation. The representation of CLD in the FWI society and in the scientific community is therefore of paramount importance. The CHLOR2NOU project is divided into 7 Work Packages that bring together scientists from various background: the WP1 with the synthesis of CLD TPs, CLD baits and fluorescent macromolecular cages; the WP2 that deals with innovative analytical methods: (i) routine laboratory method for the detection of CLD TPs in environmental and food matrices, (ii) immunoassay using a CLD-selective antibody, (iii) a semi-high-throughput detection protocol based on the recognition of CLD by a fluorescent macromolecular cage; the WP3 dedicated to toxicological and ecotoxicological studies in order to define the toxicity profile of CLD TPs; the WP4 with several analytical campaigns to obtain a first estimate of the possible exposure to CLD TPs; the WP5 that aims at studying the fate of CLD TPs, in particular in FWI soils, while defining degradation indicators; the WP6 that is focused on the study of realistic agronomic and environmental conditions capable to favor CLD degradation; the last WP centered on the representation of CLD in the FWI society at large. A co-construction method will be used to help the population and stakeholders to better assimilate the scientific results.

Peptimprint relies on the complementarity and expertise of five teams specialized in the conception of bioactive compounds (Partner 1-IBMM-team amino acids), analytical development (Partner 1-IBMM-team analytical sciences), sol-gel process and hybrid materials (Partner 2-ICG-team CMOS), microfluidics devices (Partner 3-L2C-team POMM) and study of biomolecular interactions by biosensors development (Partner 4-IRCM-team criblage) Objectives The main objective of Peptimprint is to set-up a ground-breaking technology to prepare specific tridimensional and functionalized imprints of peptides and large proteins by sol-gel process, polymerizing original hybrid building blocks mimicking amino acids around the biomolecular template. Unfunctionalized blocks (e.g. tetraethoxysilane, dimethyldichlorosilane) will be used concomitantly to create the network. Once the template biomolecule removed, the hollow cavities will be able to capture these biomolecules with very high selectivity. The second objective of Peptimprint is the preparation of unprecedented imprinted devices for the detection, separation of biomolecules and their extraction and concentration in biological samples. Surface Plasmon Resonance (SPR) and Quartz Crystal Microbalance (QCM) chips as well as polydimethylsiloxane (PDMS) based microfluidics channels will be modified with hybrid biomimetic imprints. Scientific challenges Classical molecular imprinted polymers (MIPs) suffer important limitations. Firstly, the polymerization conditions (organic solvents, non-selectivity vs amino acid side-chains…) affect the structure of the template yielding a non-relevant MIPs. Secondly, the obtainment of a large cavities deprived of functional groups generate non-specific imprints that failed to mimic the diversity of weak interactions found in natural recognitions systems. Peptimprint approach is bringing down such barriers. The sol-gel process takes place at physiological pH, in aqueous conditions and at room temperature preserving the structural and functional integrity of the protein template. Moreover, the original hybrid monomers (amino acid mimics) used in Peptimprint will recapitulate all types of interactions (ionic/hydrogen bonding/hydrophobic/aromatic stacking) involved in real interactions between biomolecules. Proceeding much more slowly than photopolymerization used in classical MIPS, sol-gel approach of Peptimprint favours a self-organization of all the functionalized hybrid blocks around the template. First models and applications Several peptide and proteins templates covering a wide range of size (from 1.5 to 150 kDa) and functions will be used as models for Peptimprint. It includes vancomycin, C-peptide, human kallikrein1, antibody fragments and therapeutic antibody. Hybrid imprints of such proteins will be prepared on the surface of QCM and SPR devices and the interaction will be studied. On the other hand, models will be either adsorbed or covalently grafted on the silicon mold to cast hybrid-PDMS microchannels. The later will be used for electrophoric analyses. Expected impacts Fundamental knowledge on molecular imprinting will be gathered thanks to Peptimprint project including (i) a generic and straightforward sol-gel method and (ii) tailored reagents (hybrid amino acids) for the inorganic polymerization of functionalized imprints. From a technological point of view, if convincing analytical data are obtained using imprinted microfluidics devices or sensors, the development of relevant selective sensors for the detection and the quantification of biomarkers will be envisioned. Indeed, Peptimprint concept can be generalized to any other types of biomolecules (oligosaccharides, oligonucleotides etc.) or even objects (viruses, bacteria) enlarging the scope of biomedical applications. A strategic advisory board will be gathered (month 24) to discuss the technological transfer opportunities and to maximize the dissemination impact.

The “EPI-B-PLASMADIFF” project (36 months, 4 teams) will address scientific issues pertinent to the B-cell-lineage and their implications in immunopathology, with specific focus on antibody-secreting plasma cells. Antibody secreting cells are critical effector cells and long-lived sentinels for immune memory. B cell maturation should be tightly regulated to ensure efficient immune response without autoimmunity or immune deficiency. On the transcriptional level, the differentiation of B cells into plasma cells is associated with substantial and coordinated changes in the gene expression profile, which fall into two main categories: the loss of B cell-associated transcripts and the acquisition of plasma cell gene expression program. Although the role of the complex network of transcription factors involved in PCD has been investigated, the mechanisms regulating key plasma cell differentiation transcription networks remain poorly known. Little is known about the role of epitranscriptomic modifications in B to plasma cell differentiation and how it could regulate fundamental processes during normal plasma cell differentiation. In this proposal, we seek to characterize and understand the epitranscriptomic remodeling and the transcriptional, translational and epigenetic impacts during B to plasma cell differentiation with a particular focus on m6A modifications. We would like to reveal both the pathways involved in this remodeling and their downstream effects. The rationale of our project is in line with the identification of major epitranscriptomic changes during plasma cell differentiation, which was identified by mass spectrometry, and preliminary data supporting a role of the enzymes involved in m6A modification in the biology of plasma cells. We plan to identify upstream pathways mediating these changes and their downstream transcriptional and translational impacts. These studies will take advantage of a unique combination of powerful models and new technologies that are fully mastered by the four partners of the project.

Measuring the affinity of a ligand for its target is an issue of central importance in life science research and drug development. In particular, understanding how well a potential drug interacts with its target (receptor, enzyme, DNA…) provides valuable knowledge in the search for new pharmaceuticals. Collective progress in chemistry, biology, robotic, informatics, led to the advent of high throughput screening (HTS) and enabled the biological evaluation of large number of drug candidates. As far as ligand-receptor system is concerned, binding of a drug candidate is characterized by a “competitive binding assay”. Since the affinity of reference ligand for target receptor is generally high, measurement of signal requires a very good sensitivity and as a consequence, radioactivity is still the method of choice for pharmacological studies of receptor/ligand systems. However, radioactivity implies a lot of constraints linked to radioelement manipulation and storage, strongly limiting high throughput screening applicability. In this context, we aim at developing a new and universal technology involving non-radioactive detection and quantification of the reference ligand displaced by the competing candidates. Mass spectrometry offers sensitive and selective methodologies to quantify molecules in complex biological systems. In contrast to known methods, the purpose of the submitted project is to develop a generic MS-based competitive binding assay avoiding the synthesis, for each evaluated molecule, of the corresponding quantification standard. This point is a great advantage since the isotopic standard used for quantification will be the same for all biological systems. The novelty of such an approach thus lies in the joint use of mass spectrometry and original labeling chemistries. Several robust mass spectrometry technologies (MALDI-MS, ESI-MS and ICP-MS) will be investigated in conjunction with different types of new chemical tags (preformed ions, elemental tags, MS enhancers…). The strategies will be validated with soluble proteins (for instance p38 kinases) as well as membrane proteins (such as two GPCR model systems: CCKB and MCR1), involving either small heterocyclic drugs or peptides as native ligands. If successful results are obtained during the course of the proposed basic research project, the developed generic tag/MS-based quantification protocol will have a great impact in biosciences, in particular in research laboratories dealing with pharmacology.