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Spike-based COVID-19 vaccines induce potent neutralizing antibodies but their efficacy against SARS-CoV-2 variants decreases. OVX033 is a recombinant protein composed of the full-length nucleocapsid (N) protein of SARS-CoV-2 genetically fused to oligoDOM ® , a self-assembling domain which improves antigen immunogenicity. OVX033 including N as an antigenic target is proposed as new vaccine candidate providing broad-spectrum protection against sarbecoviruses. OVX033 demonstrated its ability to trigger cross-reactive T cell responses and cross-protection against three variants of SARS-CoV-2 (B.1 Europe, Delta B.1.617.2, and Omicron B.1.1.529) in a hamster challenge model, as evidenced by lower weight loss, lower lung viral loads, and reduced lung histopathological lesions., Competing Interests: Author TC is an employee of VFI. Author NC is co-founder and an employee of VFI. CP, JC, SV, EN, AV, DG-G, and FN are employees of Osivax. MC was employee of Osivax at the time of her participation in the study, and her current affiliation is Manpower France in Sanofi Marcy L’Etoile, France. AV, DG-G. and FN are shareholders and executive members of Osivax. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Primard, Monchâtre-Leroy, Del Campo, Valsesia, Nikly, Chevandier, Boué, Servat, Wasniewski, Picard-Meyer, Courant, Collin, Salguero, Le Vert, Guyon-Gellin and Nicolas.)

Autophagy is a fundamental housekeeping process by which cells degrade their components to maintain homeostasis. Defects in autophagy have been associated with aging, neurodegeneration and metabolic diseases. Non-alcoholic fatty liver diseases (NAFLDs) are characterized by hepatic fat accumulation with or without inflammation. No treatment for NAFLDs is currently available, but autophagy induction has been proposed as a promising therapeutic strategy. Here, we aimed to design autophagy-inducing particles, using the autophagy-inducing peptide (Tat-Beclin), and achieve liver targeting in vivo, taking NAFLD as a model disease. Polylactic acid (PLA) particles were prepared by nanoprecipitation without any surfactant, followed by surface peptide adsorption. The ability of Tat-Beclin nanoparticles (NP T-B) to modulate autophagy and to decrease intracellular lipid was evaluated in vitro by LC3 immunoblot and using a cellular model of steatosis, respectively. The intracellular localization of particles was evaluated by transmission electron microscopy (TEM). Finally, biodistribution of fluorescent NP T-B was evaluated in vivo using tomography in normal and obese mice. The results showed that NP T-B induce autophagy with a long-lasting and enhanced effect compared to the soluble peptide, and at a ten times lower dose. Intracellular lipid also decreased in a cellular model of NAFLD after treatment with T-B and NP T-B under the same dose conditions. Ultrastructural studies revealed that NP T-B are internalized and located in endosomal, endolysosomal and autolysosomal compartments, while in healthy and obese mice, NP T-B could accumulate for several days in the liver. Given the beneficial effects of autophagy-inducing particles in vitro, and their capacity to target the liver of normal and obese mice, NP T-B could be a promising therapeutic tool for NAFLDs, warranting further in vivo investigation.

Autophagic pathways cross with lipid homeostasis and thus provide energy and essential building blocks that are indispensable for liver functions. Energy deficiencies are compensated by breaking down lipid droplets (LDs), intracellular organelles that store neutral lipids, in part by a selective type of autophagy, referred to as lipophagy. The process of lipophagy does not appear to be properly regulated in fatty liver diseases (FLDs), an important risk factor for the development of hepatocellular carcinomas (HCC). Here we provide an overview on our current knowledge of the biogenesis and functions of LDs, and the mechanisms underlying their lysosomal turnover by autophagic processes. This review also focuses on nonalcoholic steatohepatitis (NASH), a specific type of FLD characterized by steatosis, chronic inflammation and cell death. Particular attention is paid to the role of macroautophagy and macrolipophagy in relation to the parenchymal and non-parenchymal cells of the liver in NASH, as this disease has been associated with inappropriate lipophagy in various cell types of the liver. Abbreviations : ACAT: acetyl-CoA acetyltransferase; ACAC/ACC: acetyl-CoA carboxylase; AKT: AKT serine/threonine kinase; ATG: autophagy related; AUP1: AUP1 lipid droplet regulating VLDL assembly factor; BECN1/Vps30/Atg6: beclin 1; BSCL2/seipin: BSCL2 lipid droplet biogenesis associated, seipin; CMA: chaperone-mediated autophagy; CREB1/CREB: cAMP responsive element binding protein 1; CXCR3: C-X-C motif chemokine receptor 3; DAGs: diacylglycerols; DAMPs: danger/damage-associated molecular patterns; DEN: diethylnitrosamine; DGAT: diacylglycerol O-acyltransferase; DNL: de novo lipogenesis; EHBP1/NACSIN (EH domain binding protein 1); EHD2/PAST2: EH domain containing 2; CoA: coenzyme A; CCL/chemokines: chemokine ligands; CCl 4: carbon tetrachloride; ER: endoplasmic reticulum; ESCRT: endosomal sorting complexes required for transport; FA: fatty acid; FFAs: free fatty acids; FFC: high saturated fats, fructose and cholesterol; FGF21: fibroblast growth factor 21; FITM/FIT: fat storage inducing transmembrane protein; FLD: fatty liver diseases; FOXO: forkhead box O; GABARAP: GABA type A receptor-associated protein; GPAT: glycerol-3-phosphate acyltransferase; HCC: hepatocellular carcinoma; HDAC6: histone deacetylase 6; HECT: homologous to E6-AP C-terminus; HFCD: high fat, choline deficient; HFD: high-fat diet; HSCs: hepatic stellate cells; HSPA8/HSC70: heat shock protein family A (Hsp70) member 8; ITCH/AIP4: itchy E3 ubiquitin protein ligase; KCs: Kupffer cells; LAMP2A: lysosomal associated membrane protein 2A; LDs: lipid droplets; LDL: low density lipoprotein; LEP/OB: leptin; LEPR/OBR: leptin receptor; LIPA/LAL: lipase A, lysosomal acid type; LIPE/HSL: lipase E, hormone sensitive type; LIR: LC3-interacting region; LPS: lipopolysaccharide; LSECs: liver sinusoidal endothelial cells; MAGs: monoacylglycerols; MAPK: mitogen-activated protein kinase; MAP3K5/ASK1: mitogen-activated protein kinase kinase kinase 5; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCD: methionine-choline deficient; MGLL/MGL: monoglyceride lipase; MLXIPL/ChREBP: MLX interacting protein like; MTORC1: mechanistic target of rapamycin kinase complex 1; NAFLD: nonalcoholic fatty liver disease; NAS: NAFLD activity score; NASH: nonalcoholic steatohepatitis; NPC: NPC intracellular cholesterol transporter; NR1H3/LXRα: nuclear receptor subfamily 1 group H member 3; NR1H4/FXR: nuclear receptor subfamily 1 group H member 4; PDGF: platelet derived growth factor; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PLIN: perilipin; PNPLA: patatin like phospholipase domain containing; PNPLA2/ATGL: patatin like phospholipase domain containing 2; PNPLA3/adiponutrin: patatin like phospholipase domain containing 3; PPAR: peroxisome proliferator activated receptor; PPARA/PPARα: peroxisome proliferator activated receptor alpha; PPARD/PPARδ: peroxisome proliferator activated receptor delta; PPARG/PPARγ: peroxisome proliferator activated receptor gamma; PPARGC1A/PGC1α: PPARG coactivator 1 alpha; PRKAA/AMPK: protein kinase AMP-activated catalytic subunit; PtdIns3K: class III phosphatidylinositol 3-kinase; PtdIns3P: phosphatidylinositol-3-phosphate; PTEN: phosphatase and tensin homolog; ROS: reactive oxygen species; SE: sterol esters; SIRT1: sirtuin 1; SPART/SPG20: spartin; SQSTM1/p62: sequestosome 1; SREBF1/SREBP1c: sterol regulatory element binding transcription factor 1; TAGs: triacylglycerols; TFE3: transcription factor binding to IGHM enhancer 3; TFEB: transcription factor EB; TGFB1/TGFβ: transforming growth factor beta 1; Ub: ubiquitin; UBE2G2/UBC7: ubiquitin conjugating enzyme E2 G2; ULK1/Atg1: unc-51 like autophagy activating kinase 1; USF1: upstream transcription factor 1; VLDL: very-low density lipoprotein; VPS: vacuolar protein sorting; WIPI: WD-repeat domain, phosphoinositide interacting; WDR: WD repeat domain.

In the development of therapeutic nanoparticles (NP), there is a large gap between in vitro testing and in vivo experimentation. Despite its prominence as a model, the mouse shows severe limitations for imaging NP and the cells with which they interact. Recently, the transparent zebrafish larva, which is well suited for high-resolution live-imaging, has emerged as a powerful alternative model to investigate the in vivo behavior of NP. Poly(D,L lactic acid) (PLA) is widely accepted as a safe polymer to prepare therapeutic NP. However, to prevent aggregation, many NP require surfactants, which may have undesirable biological effects. Here, we evaluate 'safe-by-design', surfactant-free PLA-NP that were injected intravenously into zebrafish larvae. Interaction of fluorescent NPs with different cell types labelled in reporter animals could be followed in real-time at high resolution; furthermore, by encapsulating colloidal gold into the matrix of PLA-NP we could follow their fate in more detail by electron microscopy, from uptake to degradation. The rapid clearance of fluorescent PLA-NP from the circulation coincided with internalization by endothelial cells lining the whole vasculature and macrophages. After 30 min, when no NP remained in circulation, we observed that macrophages continued to internalize significant amounts of NP. More detailed video-imaging revealed a new mechanism of NP transfer where NP are transmitted along with parts of the cytoplasm from endothelial cells to macrophages., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: After the golden age of antibiotic discovery, bacterial infections still represent a major challenge for public health worldwide. The biofilm mode of growth is mostly responsible for chronic infections that current therapeutics fail to cure and it is well-established that novel strategies must be investigated. Particulate drug delivery systems are considered as a promising strategy to face issues related to antibiotic treatments in a biofilm context. Particularly, poly-lactic acid (PLA) nanoparticles present a great interest due to their ability to migrate into biofilms thanks to their submicronic size. However, questions still remain unresolved about their mode of action in biofilms depending on their surface properties. In the current study, we have investigated the impact of their surface charge, firstly on their behavior within a bacterial biofilm, and secondly on the antibiotic delivery and the treatment efficacy., Results: Rifampicin-loaded PLA nanoparticles were synthetized by nanoprecipitation and characterized. A high and superficial loading of rifampicin, confirmed by an in silico simulation, enabled to deliver effective antibiotic doses with a two-phase release, appropriate for biofilm-associated treatments. These nanoparticles were functionalized with poly-L-lysine, a cationic peptide, by surface coating inducing charge reversal without altering the other physicochemical properties of these particles. Positively charged nanoparticles were able to interact stronger than negative ones with Staphylococcus aureus, under planktonic and biofilm modes of growth, leading to a slowed particle migration in the biofilm thickness and to an improved retention of these cationic particles in biofilms. While rifampicin was totally ineffective in biofilms after washing, the increased retention capacity of poly-L-lysine-coated rifampicin-loaded PLA nanoparticles has been associated with a better antibiotic efficacy than uncoated negatively charged ones., Conclusions: Correlating the carrier retention capacity in biofilms with the treatment efficacy, positively charged rifampicin-loaded PLA nanoparticles are therefore proposed as an adapted and promising approach to improve antibiotic delivery in S. aureus biofilms.

Poly(lactic acid) (PLA) nanoparticles (NPs) are widely investigated due to their bioresorbable, biocompatible and low immunogen properties. Interestingly, many recent studies show that they can be efficiently used as drug delivery systems or as adjuvants to enhance vaccine efficacy. Our work focuses on the molecular mechanisms involved during the nanoprecipitation of PLA NPs from concentrated solutions of lactic acid polymeric chains, and their specific interactions with biologically relevant molecules. In this study, we evaluated the ability of a PLA-based nanoparticle drug carrier to vectorize either vitamin E or the Toll-like receptor (TLR) agonists Pam 1 CSK 4 and Pam 3 CSK 4 , which are potent activators of the proinflammatory transcription factor NF-κB. We used dissipative particle dynamics (DPD) to simulate large systems mimicking the nanoprecipitation process for a complete NP. Our results evidenced that after the NP formation, Pam 1 CSK 4 and Pam 3 CSK 4 molecules end up located on the surface of the particle, interacting with the PLA chains via their fatty acid chains, whereas vitamin E molecules are buried deeper in the core of the particle. Our results allow for a better understanding of the molecular mechanisms responsible for the formation of the PLA NPs and their interactions with biological molecules located either on their surfaces or encapsulated within them. This work should allow for a rapid development of better biodegradable and safe vectorization systems with new drugs in the near future.

Current influenza vaccines manufactured using eggs have considerable limitations, both in terms of scale up production and the potential impact passaging through eggs can have on the antigenicity of the vaccine virus strains. Alternative methods of manufacture are required, particularly in the context of an emerging pandemic strain. Here we explore the production of recombinant influenza haemagglutinin using the ciliated protozoan Tetrahymena thermophila . For the first time we were able to produce haemagglutinin from both seasonal influenza A and B strains. This ciliate derived material was immunogenic, inducing an antibody response in both mice and non-human primates. Mice immunized with ciliate derived haemagglutinin were protected against challenge with homologous influenza A or B viruses. The antigen could also be combined with submicron particles containing a Nod2 ligand, significantly boosting the immune response and reducing the dose of antigen required. Thus, we show that Tetrahymena can be used as a manufacturing platform for viral vaccine antigens., (Copyright © 2019 Jawinski, Hartmann, Singh, Kinnear, Busse, Ciabattini, Fiorino, Medaglini, Trombetta, Montomoli, Contreras, Le Grand, Coiffier, Primard, Verrier and Tregoning.)

The European autophagy consortium Driving next-generation autophagy researchers towards translation (DRIVE) held its kick-off meeting in Groningen on the 14 th and 15 th of June 2018. This Marie Skłodowska-Curie Early Training Network was approved under the European Union's Horizon 2020 Research and Innovation Program and is funded for 4 years. Within DRIVE, 14 European research teams from academia and industry will train 15 PhD students through applied, cross-disciplinary and collaborative macroautophagy/autophagy research. The goal of DRIVE is to stimulate applied approaches in autophagy research and provide training towards translation, while advancing our knowledge on autophagy in specific physiological and pathological states. The strong focus on translation will prepare the PhD students to be at the forefront to exploit autophagy for the development of therapies directly benefitting patients. Thereby, DRIVE will contribute to filling the educational gap that currently exists between academia and industry, and will prepare its PhD students for alternative and highly flexible professional paths.

In view of preparing antibiotic-loaded structures that can be used as dressing to prevent or contain wound infections, this study evaluates biodegradable nanofibrillar matrices obtained by jet-spraying and containing ciprofloxacin (CIF). The matrices were prepared from different blends of poly-(ε-caprolactone) (PCL) and poly-d,l-(lactic acid) (PDLLA) in view of controlling mechanical properties, biodegradation and antibiotic release rate. The effect of CIF incorporation was assessed in regard of matrices fiber diameter, mechanical properties and degradation while antibiotic release from the polymer blends of different PCL/PDLLA ratios was measured in buffers of different pH to better mimic the wound context. Finally, antibiotic activity of the nanofibrillar matrices and their ability to be colonized by skin cells were evaluated. Non-woven nanofibrillar matrices could be obtained from various polymer blends by jet-spraying and CIF crystals incorporation was easily obtained. The crystals were dispersed in the fibers, without complete embedding. Antibiotic incorporation resulted in a slight increase of fiber diameter and did not modified the mechanical properties of the various matrices composed of different polymer blends. Unlike fiber diameter, degradation and mechanical properties of the fibrillar matrices, CIF release profiles were not controlled by the polymer blend ratios. However, sustained release was observed over more than 23days. Due to the antibiotic pH-dependent solubility, burst release was more prominent in acidic conditions, which mimic the pH of undamaged skin. Finally the incorporated antibiotic was efficient in inhibiting bacterial growth of E. coli and B. subtilis whereas human fibroblasts were able to colonize the CIF-loaded matrices., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Intradermal delivery of antigen represents a potent route of immunization that involves multiple blood- and skin-derived dendritic cell subpopulations endowed with specialized functions and dynamics in their ability to prime naïve CD4 + T cells in the draining lymph nodes. However, their individual contributions to the generation of CD4 + T follicular helper (T FH ) cells and germinal centers (GCs) remain to be understood. We found that intradermal immunization of mice with a particle-based vaccine induced robust T FH and germinal center B-cell responses in skin draining lymph nodes, which were completely abrogated when skin cell emigration was prevented. However, in this later condition, both lymph node-resident and blood-derived inflammatory cells access the antigen in the draining lymph nodes but are not able to induce T FH cell differentiation. Rather, only skin-derived dendritic cells up-regulated key genes related to T FH cell development in the draining lymph nodes. Depletion of Langerhans cells partially abrogated T FH and germinal center B-cell responses. Thus, after intradermal immunization, only skin-derived migratory dendritic cells, including Langerhans cells, permit the generation of T FH cells and germinal centers. Identifying the relative contributions of tissue and lymphoid organ dendritic cell subsets in generating humoral immune responses is of great importance for the development of tailored vaccines., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Polyelectrolyte complexes (PECs) constituted of chitosan and chondroitin sulfate (ChonS) were formed by the one-shot addition of default amounts of polyanion to an excess of polycation. Key variables of the formulation process (e.g., degree of depolymerization, charge mixing ratio, the concentration, and pH of polyelectrolyte solutions) were optimized based on the PECs sizes and polydispersities. The PECs maintained their colloidal stability at physiological salt concentration and pH thanks to the complexation of polyelectrolytes with zinc(II) ion during the nanoPECs formation process. The PECs were capable of encapsulating an antiretroviral drug tenofovir (TF) with a minimal alteration on the colloidal stability of the dispersion. Moreover, the particle interfaces could efficiently be functionalized with anti-OVA or anti-α4β7 antibodies with conservation of the antibody biorecognition properties over 1 week of storage in PBS at 4 °C. In vitro cytotoxicity studies showed that zinc(II) stabilized chitosan-ChonS nanoPECs were noncytotoxic to human peripheral blood mononuclear cells (PBMCs), and in vitro antiviral activity test demonstrated that nanoparticles formulations led to a dose-dependent reduction of HIV-1 infection. Using nanoparticles as a drug carrier system decreases the IC50 (50% inhibitory concentration) from an aqueous TF of 4.35 μmol·L(-1) to 1.95 μmol·L(-1). Significantly, zinc ions in this system also exhibited a synergistic effect in the antiviral potency. These data suggest that chitosan-ChonS nanoPECs can be promising drug delivery system to improve the antiviral potency of drugs to the viral reservoirs for the treatment of HIV infection.

Zinc(ii) stabilized polyelectrolyte nano-complexes (PECs) of chitosan and hyaluronan (HYA) were designed as safe and efficient drug delivery systems. HIV-1 reverse transcriptase inhibitor tenofovir (TF) was quantitatively encapsulated and the particle interface could be functionalized in PBS with targeting proteins such as anti-α4β7 immunoglobulin A. Chitosan-HYA nanoPECs were non-cytotoxic on human peripheral blood mononuclear cells (PBMCs), within the investigated nanoparticle concentrations. A dose-dependent reduction of the HIV-1 infection of PBMCs co-cultured with the nanocarriers was observed. Even more interestingly, a synergistic effect was evidenced with the nanocarriers by comparing the IC 50 (50% inhibitory concentration) value of the aqueous TF solution (4.35 μmol L -1 ) with that of TF loaded nanoPECs (1.71 μmol L -1 ) and anti-α4β7 IgA functionalized TF/nanoPECs (1.01 μmol L -1 ). This effect could be attributed to the presence of zinc(ii) in the formulation of the colloids. All these data establish that the zinc(ii) stabilized chitosan-HYA nanoPECs can be potentially efficient and safe colloidal delivery system candidates for enhancing antiviral activities in the treatment of HIV infection and AIDS.

Since recent data suggest that nanoparticles and modified vaccinia ankara (MVA) vectors could play a pivotal role in HIV-1 therapeutics and vaccine design, in an ex vivo model of human monocyte-derived dendritic cells (MDDCs), we compared two different loading strategies with HIV-1 vaccine vehicles, either viral or synthetic derived. We used polylactic acid (PLA) colloidal biodegradable particles, coated with HIV Gag antigens (p24), and MVA expressing Gag (rMVA-gag and rMVA-gag/trans membrane) or Tat, Nef and Rev genes (rMVA tat+rev and rMVA nef). PLA-p24 captured by MDDCs from HIV-1 individuals induced a slight degree of MDDC maturation, cytokine and chemokine secretion and migration towards a gradient of CCL19 chemokine and highly increased HIV-specific CD8(+) T-cell proliferation compared with p24 alone. After complete maturation induction of PLA-p24-pulsed MDDCs, maximal migration towards a gradient of CCL19 chemokine and induction of HIV-specific T-cell proliferation (two-fold higher for CD4(+) than CD8(+)) and cytokine secretion (IFN-γ and IL-2) in the co-culture were observed. Upon exposure to MVA-gag, MDDCs produced cytokines and chemokines and maintained their capacity to migrate to a gradient of CCL19. MDDCs infected with MVA-gag and MVA-gag trans-membrane were able to induce HIV-specific CD8(+) proliferation and secretion of IFN-γ, IL-2, IL-6 and TNF-α. We conclude that both HIV antigens loading strategies (PLA-p24 nanoparticles or MVA expressing HIV genes) induce HIV-1-specific T-cell responses, which are able to kill autologous gag-expressing cells. Thus, they are plausible candidates for the development of anti-HIV vaccines., (Copyright © 2014 Elsevier Ltd. All rights reserved.)