N-terminal modification of an LAH4-derived peptide increases mRNA delivery in the presence of serum.

The recently developed mRNA-based coronavirus SARS-CoV-2 vaccines highlighted the great therapeutic potential of the mRNA technology. Although the lipid nanoparticles used for the delivery of the mRNA are very efficient, they showed, in some cases, the induction of side effects as well as the production of antibodies directed against particle components. Thus, the development of alternative delivery systems is of great interest in the pursuit of more effective mRNA treatments. In the present work, we evaluated the mRNA transfection capacities of a series of cationic histidine-rich amphipathic peptides derived from LAH4. We found that while the LAH4-A1 peptide was an efficient carrier for mRNA, its activity was highly serum sensitive. Interestingly, modification of this cell penetrating peptide at the N-terminus with two tyrosines or with salicylic acid allowed to confer serum resistance to the carrier.

N-Heterocyclic Carbene-Polyethyleneimine (PEI) Platinum Complexes Inducing Human Cancer Cell Death: Polymer Carrier Impact.

The high interest in N-Heterocyclic platinum carbene complexes in cancer research stems from their high cytotoxicity to human cancer cells, their stability, as well as their ease of functionalization. However, the development of these new molecules as anticancer agents still faces multiple challenges, in particular solubility in aqueous media. Here, we synthesized platinum-NHC bioconjugates that combine water-solubility and cytotoxicity by using polyethyleneimine as polymer carrier. We showed on 8 different types of cells that the activity of these conjugates is modulated by the size of the polymer and the overall density of metal ions onto polymer chains. Using HCT116 cells, the conjugates displayed an effective activity after only 45 min of exposure in vitro correlated with a quick uptake by the cells as shown by the use of various fluorescent-tagged derivatives.

N-Heterocyclic Carbene-Polyethylenimine Platinum Complexes with Potent in Vitro and in Vivo Antitumor Efficacy.

The current interest for platinum N-heterocyclic carbene complexes in cancer research stems from their impressive toxicity reported against a range of different human cancer cells. To date, the demonstration of their in vivo efficacy relative to that of established platinum-based drugs has not been specifically addressed. Here, we introduce an innovative approach to increase the NHC-Pt complex potency whereby multiple NHC-Pt(II) complexes are coordinated along a polyethylenimine polymer (PEI) chain. We show that such NHC-Pt(II)-PEI conjugates induce human cancer cell death in vitro and in vivo in a xenograft mouse model with no observable side effects in contrast to oxaliplatin. Additional studies indicate nucleus and mitochondria targeting and suggest various mechanisms of action compared to classical platinum-based anticancer drugs.

Thioether analogues of disulfide-bridged cyclic peptides targeting death receptor 5: conformational analysis, dimerisation and consequences for receptor activation.

Cyclic peptides containing redox-stable thioether bridges might provide a useful alternative to disulfide-bridged bioactive peptides. We report the effect of replacing the disulfide bridge with a lanthionine linkage in a 16-mer cyclic peptide that binds to death receptor 5 (DR5, TRAIL-R2). Upon covalent oligomerisation, the disulfide-bridged peptide has previously shown similar behaviour to that of TNF-related apoptosis inducing ligand (TRAIL), by selectively triggering the DR5 cell death pathway. The structural and biological properties of the DR5-binding peptide and its desulfurised analogue were compared. Surface plasmon resonance (SPR) data suggest that these peptides bind DR5 with comparable affinities. The same holds true for dimeric versions of these peptides: the thioether is able to induce DR5-mediated apoptosis of BJAB lymphoma and tumorigenic BJELR cells, albeit to a slightly lower extent compared to its disulfide homologue. NMR analysis revealed subtle variation in the conformations of the two peptides and suggests that the thioether peptide is slightly less folded than its disulfide homologue. These observations could account for the different capability of the two dimers to cluster DR5 receptors on the cell surface and to trigger apoptosis. Nevertheless, our results suggest that the thioether peptide is a potential candidate for evaluation in animal models.

Cysteine-rich domain 1 of CD40 mediates receptor self-assembly.

The activation of CD40 on B cells, macrophages, and dendritic cells by its ligand CD154 (CD40L) is essential for the development of humoral and cellular immune responses. CD40L and other TNF superfamily ligands are noncovalent homotrimers, but the form under which CD40 exists in the absence of ligand remains to be elucidated. Here, we show that both cell surface-expressed and soluble CD40 self-assemble, most probably as noncovalent dimers. The cysteine-rich domain 1 (CRD1) of CD40 participated to dimerization and was also required for efficient receptor expression. Modelization of a CD40 dimer allowed the identification of lysine 29 in CRD1, whose mutation decreased CD40 self-interaction without affecting expression or response to ligand. When expressed alone, recombinant CD40-CRD1 bound CD40 with a K(D) of 0.6 µM. This molecule triggered expression of maturation markers on human dendritic cells and potentiated CD40L activity. These results suggest that CD40 self-assembly modulates signaling, possibly by maintaining the receptor in a quiescent state.

Reduction of Pro-Inflammatory Markers in RAW264.7 Macrophages by Polyethylenimines.

The physiological problem of chronic inflammation and its associated pathologies attract ongoing attention with regard to methods for their control. Current systemic pharmacological treatments present problematic side effects. Thus, the possibility of new anti-inflammatory compounds with differing mechanisms of action or biophysical properties is enticing. Cationic polymers, with their ability to act as carriers for other molecules or to form bio-compatible materials, present one such possibility. Although not well described, several polycations such as chitosan and polyarginine, have displayed anti-inflammatory properties. The present work shows the ubiquitous laboratory transfection reagent, polyethylenimine (PEI) and more specifically low molecular weight branched PEI (B-PEI) as also possessing such properties. Using a RAW264.7 murine cell line macrophage as an inflammation model, it is found the B-PEI 700 Da as being capable of reducing the production of several pro-inflammatory molecules induced by the endotoxin lipopolysaccharide. Although further studies are required for elucidation of its mechanisms, the revelation that such a common lab reagent may present these effects has wide-ranging implications, as well as an abundance of possibilities.

Antidepressants suppress neuropathic pain by a peripheral ß2-adrenoceptor mediated anti-TNFα mechanism.

Neuropathic pain is pain arising as a direct consequence of a lesion or disease affecting the somatosensory system. It is usually chronic and challenging to treat. Some antidepressants are first-line pharmacological treatments for neuropathic pain. The noradrenaline that is recruited by the action of the antidepressants on reuptake transporters has been proposed to act through ß2-adrenoceptors (ß2-ARs) to lead to the observed therapeutic effect. However, the complex downstream mechanism mediating this action remained to be identified. In this study, we demonstrate in a mouse model of neuropathic pain that an antidepressant's effect on neuropathic allodynia involves the peripheral nervous system and the inhibition of cytokine tumor necrosis factor α (TNFα) production. The antiallodynic action of nortriptyline is indeed lost after peripheral sympathectomy, but not after lesion of central descending noradrenergic pathways. More particularly, we report that antidepressant-recruited noradrenaline acts, within dorsal root ganglia, on ß2-ARs expressed by non-neuronal satellite cells. This stimulation of ß2-ARs decreases the neuropathy-induced production of membrane-bound TNFα, resulting in relief of neuropathic allodynia. This indirect anti-TNFα action was observed with the tricyclic antidepressant nortriptyline, the selective serotonin and noradrenaline reuptake inhibitor venlafaxine and the ß2-AR agonist terbutaline. Our data revealed an original therapeutic mechanism that may open novel research avenues for the management of painful peripheral neuropathies.

Multimerization of an apoptogenic TRAIL-mimicking peptide by using adamantane-based dendrons.

We have developed a straightforward strategy to multimerize an apoptogenic peptide that mimics the natural tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) by using adamantane-based dendrons as multivalent scaffolds. The selective binding affinity of the ligands to TRAIL receptor 2 (TR2) was studied by surface plasmon resonance, thus demonstrating that the trimeric and hexameric forms of the peptide exert an increased affinity of about 1500- and 20,000-fold, respectively, relative to the monomer. Moreover, only the trimeric and hexameric ligands were able to induce cell death in TR2 expressing cells (BJAB), thus confirming that a multivalent form of the peptide is necessary to trigger a substantial TR2-dependent apoptotic response in vitro. These results provide interesting insight into the multivalency effect on biological ligand/receptor interactions for future therapeutic applications.

Triphenylphosphonium-functionalized N-heterocyclic carbene platinum complexes [(NHC-TPP+)Pt] induce cell death of human glioblastoma cancer stem cells.

A growing body of experimental and clinical evidence suggests that rare cell populations, known as cancer stem cells (CSCs), play an important role in the development and therapeutic resistance of several cancers, including glioblastoma. Elimination of these cells is therefore of paramount importance. Interestingly, recent results have shown that the use of drugs that specifically disrupt mitochondria or induce mitochondria-dependent apoptosis can efficiently kill cancer stem cells. In this context, a novel series of platinum(II) complexes bearing N-heterocyclic carbene (NHC) of the type [(NHC)PtI2(L)] modified with the mitochondria targeting group triphenylphosphonium were synthesized. After a complete characterization of the platinum complexes, the cytotoxicity against two different cancer cell lines, including a cancer stem cell line, was investigated. The best compound reduced the cell viability of both cell lines by 50% in the low µM range, with an approximately 300-fold higher anticancer activity on the CSC line compared to oxaliplatin. Finally, mechanistic studies showed that the triphenylphosphonium functionalized platinum complexes significantly altered mitochondrial function and also induced atypical cell death.

Anti-inflammatory effects of free and liposome-encapsulated Algerian thermal waters in RAW 264.7 macrophages.

The main objectives of this work were to formulate liposomes encapsulating highly mineralized thermal waters (TWs) and to study anti-inflammatory effect of free and encapsulated thermal waters on RAW 264.7 macrophage cells stimulated with lipopolysaccharide (LPS). TWs-loaded conventional and deformable liposomes (TWs-Lip and TWs-DLip) were prepared by sonication and extrusion, respectively. They were considered for their vesicle size, zeta potential, entrapment efficiency, physical stability and in vitro anti-inflammatory effect. Formulated liposome suspensions have a low polydispersity and nanometric size range with zeta potential values close to zero. The vesicle size was stable for 30 days. Entrapment efficiency of TWs was above 90% in conventional liposomes and 70% in deformable liposomes. Pretreatment of LPS-stimulated murine macrophages, with free and liposome-encapsulated TWs, resulted in a significant reduction in nitric oxide (NO) production and modulated tumor necrosis factor-α (TNF-α) production suggesting an anti-inflammatory effect which was even more striking with TWs-Lip and TWs-DLip. Liposome formulations may offer a suitable approach for transdermal delivery of TWs, indicated in inflammatory skin diseases.

An N-heterocyclic carbene iridium(III) complex as a potent anti-cancer stem cell therapeutic.

Cancer stem cells (CSCs) represent a difficult to treat cellular niche within tumours due to their unique characteristics, which give them a high propensity for resistance to classical anti-cancer treatments and the ability to repopulate the tumour mass. An attribute that may be implicated in the high rates of recurrence of certain tumours. However, other characteristics specific to these cells, such as their high dependence on mitochondria, may be exploited for the development of new therapeutic agents that are effective against the niche. As such, a previously described phosphorescent N-heterocyclic carbene iridium(III) compound which showed a high level of cytotoxicity against classical tumour cell lines with mitochondria-specific effects was studied for its potential against CSCs. The results showed a significantly higher level of activity against several CSC lines compared to non-CSCs. Mitochondrial localisation and superoxide production were confirmed. Although the cell death involved caspase activation, their role in cell death was not definitive, with a potential implication of other, non-apoptotic pathways shown. A cytostatic effect of the compound was also displayed at low mortality doses. This study thus provides important insights into the mechanisms and the potential for this class of molecule in the domain of anti-CSC therapeutics.

Polyarginine as a Simultaneous Antimicrobial, Immunomodulatory, and miRNA Delivery Agent within Polyanionic Hydrogel.

Implantation of biomedical devices is followed by immune response to the implant, as well as occasionally bacterial, yeast, and/or fungal infections. In this context, new implant materials and coatings that deal with medical device-associated complications are required. Antibacterial and anti-inflammatory materials are also required for wound healing applications, especially in diabetic patients with chronic wounds. In this work, hyaluronic acid (HA) hydrogels with triple activity: antimicrobial, immunomodulatory, and miRNA delivery agent, are presented. It is demonstrated that polyarginine with a degree of polymerization of 30 (PAR30), which is previously shown to have a prolonged antibacterial activity, decreases inflammatory response of lipopolysaccharide-stimulated macrophages. In addition, PAR30 accelerates fibroblast migration in macrophage/fibroblast coculture system, suggesting a positive effect on wound healing. Furthermore, PAR30 allows to load miRNA into HA hydrogels, and then to deliver them into the cells. To the authors knowledge, this study is the first describing miRNA-loaded hydrogels with antibacterial effect and anti-inflammatory features. Such system can become a tool for the treatment of infected wounds, e.g., diabetic ulcers, as well as for foreign body response modulation.

Past mastering of metal transformation enabled physicians to increase their therapeutic potential.

BACKGROUND: Metals are trace elements, vital in some instances or toxic in others. Due to this toxicity, they have been used since ancient time as antimicrobials, and prescribed when plant-only remedies were not efficient enough. These remedies could still contain secrets that may lead to the discovery of new therapeutically interesting combinations. The objective of this study was to give a proof of concept that such remedies combining metals and plants are worth studying again. METHODS: We exploited 4 medical formularies (aqrabadhin), from three Arab authors from the 9-12th century. We reproduced a remedy, and analyzed the role of each ingredient. We further looked for the minimum inhibitory concentration against three pathogenic bacteria, and we analyzed toxic and inflammatory effects of this remedy on macrophages. RESULTS: Even if plants were extensively used (almost 80 % of all ingredients), more than 36 different minerals have been found in these 4 aqrabadhin. When it came to remedies against infections that could be applied externally, the use of metals grew to 70 %. We focused on a remedy, containing mainly metals. We have been able to attribute a role for each ingredient, to show that this skin remedy helped to combat the infection and to resorb the wound, and to highlight the mastering of metal transformation by these physicians. CONCLUSIONS: With a very simple recipe, mainly composed of metals, these past physicians designed a complete and synergistic remedy to combat abscesses, while restricting the toxic effect of metals to the site of infection. It is a first example showing that different metal manufactures were evolved to improve their therapeutic potentials. The knowledge acquired by these physician should deserve more attention, and unexpected features, original organo-metallic compounds or therapeutic synergy could still be found from such research.

Polyethylenimine, an Autophagy-Inducing Platinum-Carbene-Based Drug Carrier with Potent Toxicity towards Glioblastoma Cancer Stem Cells.

The difficulty involved in the treatment of many tumours due to their recurrence and resistance to chemotherapy is tightly linked to the presence of cancer stem cells (CSCs). This CSC sub-population is distinct from the majority of cancer cells of the tumour bulk. Indeed, CSCs have increased mitochondrial mass that has been linked to increased sensitivity to mitochondrial targeting compounds. Thus, a platinum-based polyethylenimine (PEI) polymer-drug conjugate (PDC) was assessed as a potential anti-CSC therapeutic since it has previously displayed mitochondrial accumulation. Our results show that CSCs have increased specific sensitivity to the PEI carrier and to the PDC. The mechanism of cell death seems to be necrotic in nature, with an absence of apoptotic markers. Cell death is accompanied by the induction of a protective autophagy. The interference in the balance of this pathway, which is highly important for CSCs, may be responsible for a partial reversion of the stem-like phenotype observed with prolonged PEI and PDC treatment. Several markers also indicate the cell death mode to be capable of inducing an anti-cancer immune response. This study thus indicates the potential therapeutic perspectives of polycations against CSCs.

Evaluation of the Cytotoxicity of Cationic Polymers on Glioblastoma Cancer Stem Cells.

Cationic polymers such as polyethylenimine (PEI) have found a pervasive place in laboratories across the world as gene delivery agents. However, their applications are not limited to this role, having found a place as delivery agents for drugs, in complexes known as polymer-drug conjugates (PDCs). Yet a potentially underexplored domain of research is in their inherent potential as anti-cancer therapeutic agents, which has been indicated by several studies. Even more interesting is the recent observation that certain polycations may present a significantly greater toxicity towards the clinically important cancer stem cell (CSC) niche than towards more differentiated bulk tumour cells. These cells, which possess the stem-like characteristics of self-renewal and differentiation, are highly implicated in cancer drug resistance, tumour recurrence and poor clinical prognosis. The search for compounds which may target and eliminate these cells is thus of great research interest. As such, the observation in our previous study on a PEI-based PDC which showed a considerably higher toxicity of PEI towards glioblastoma CSCs (GSCs) than on more differentiated glioma (U87) cells led us to investigate other cationic polymers for a similar effect. The evaluation of the toxicity of a range of different types of polycations, and an investigation into the potential source of GSC's sensitivity to such compounds is thus described.

Autoantigen microarrays for multiplex characterization of autoantibody responses.

We constructed miniaturized autoantigen arrays to perform large-scale multiplex characterization of autoantibody responses directed against structurally diverse autoantigens, using submicroliter quantities of clinical samples. Autoantigen microarrays were produced by attaching hundreds of proteins, peptides and other biomolecules to the surface of derivatized glass slides using a robotic arrayer. Arrays were incubated with patient serum, and spectrally resolvable fluorescent labels were used to detect autoantibody binding to specific autoantigens on the array. We describe and characterize arrays containing the major autoantigens in eight distinct human autoimmune diseases, including systemic lupus erythematosus and rheumatoid arthritis. This represents the first report of application of such technology to multiple human disease sera, and will enable validated detection of antibodies recognizing autoantigens including proteins, peptides, enzyme complexes, ribonucleoprotein complexes, DNA and post-translationally modified antigens. Autoantigen microarrays represent a powerful tool to study the specificity and pathogenesis of autoantibody responses, and to identify and define relevant autoantigens in human autoimmune diseases.

Recent Advances in Antiinflammatory Material Design.

Implants and prostheses are widely used to replace damaged tissues or to treat various diseases. However, besides the risk of bacterial or fungal infection, an inflammatory response usually occurs. Here, recent progress in the field of anti-inflammatory biomaterials is described. Different materials and approaches are used to decrease the inflammatory response, including hydrogels, nanoparticles, implant surface coating by polymers, and a variety of systems for anti-inflammatory drug delivery. Complex multifunctional systems dealing with inflammation, microbial infection, bone regeneration, or angiogenesis are also described. New promising stimuli-responsive systems, such as pH- and temperature-responsive materials, are also being developed that would enable an "intelligent" antiinflammatory response when the inflammation occurs. Together, different approaches hold promise for creation of novel multifunctional smart materials allowing better implant integration and tissue regeneration.

Novel powerful water-soluble lipid immunoadjuvants inducing mouse dendritic cell maturation and B cell proliferation using TLR2 pathway.

Four novel water-soluble lipid immunoadjuvants were designed, synthesized and characterized by MS and NMR. They all induce mouse dendritic cell maturation and B cell proliferation. We demonstrate that in spite of the chemical modification, the four compounds remain TLR2 agonists.

Liposomes as tunable platform to decipher the antitumor immune response triggered by TLR and NLR agonists.

Liposomes are powerful tools for the optimization of peptides and adjuvant composition in cancer vaccines. Here, we take advantage of a liposomal platform versatility to develop three vaccine candidates associating a peptide from HA influenza virus protein as CD4 epitope, a peptide from HPV16 E7 oncoprotein as CD8 epitope and TLR4, TLR2/6 or NOD1 agonists as adjuvant. Liposomal vaccine containing MPLA (TLR4 liposomes), are the most effective treatment against the HPV-transformed orthotopic lung tumor mouse model, TC-1. This vaccine induces a potent Th1-oriented antitumor immunity, which leads to a significant reduction in tumor growth and a prolonged survival of mice, even when injected after tumor appearance. This efficacy is dependent on CD8+ T cells. Subcutaneous injection of this treatment induces the migration of skin DCs to draining lymph nodes. Interestingly, TLR2/6 liposomes trigger a weaker Th1-immune response which is not sufficient for the induction of a prolonged antitumor activity. Although NOD1 liposome treatment results in the control of early tumor growth, it does not extend mice survival. Surprisingly, the antitumor activity of NOD1 vaccine is not associated with a specific adaptive immune response. This study shows that our modulable platform can be used for the strategical development of vaccines.

La Société de Biologie de Strasbourg : 100 ans au service de la science et de la société.

Founded in 1919, the Society of Biology of Strasbourg (SBS) is a learned society whose purpose is the dissemination and promotion of scientific knowledge in biology. Subsidiary of the Society of Biology, the SBS celebrated its Centenary on Wednesday, the 16th of October 2019 on the Strasbourg University campus and at the Strasbourg City Hall. This day allowed retracing the various milestones of the SBS, through its main strengths, its difficulties and its permanent goal to meet scientific and societal challenges. The common thread of this day was the transmission of knowledge related to the past, the present, but also the future. At the start of the 21st century, the SBS must continue to reinvent itself to pursue its objective of transmitting scientific knowledge in biology and beyond. Scientific talks performed by senior scientists and former SBS thesis prizes awardees, a round table, and informal discussions reflected the history and the dynamism of the SBS association. All SBS Centennial participants have set the first milestone for the SBS Bicentennial.

TITLE: La Société de Biologie de Strasbourg : 100 ans au service de la science et de la société. ABSTRACT: Filiale de la Société de Biologie, la Société de Biologie de Strasbourg (SBS) est une société savante qui a pour objet la diffusion et la promotion du savoir scientifique en biologie et en médecine. Fondée en 1919, La SBS a célébré son Centenaire le mercredi 16 octobre 2019. Cette journée a permis de retracer les différents jalons de la SBS, à travers ses lignes de forces, ses difficultés et sa volonté permanente de mettre en exergue les défis scientifiques et sociétaux auxquels participent les recherches strasbourgeoises. Le fil rouge de cette journée a été la transmission d'un savoir en lien avec le passé, le présent, mais également le futur. En ce début du 21e siècle, la SBS se doit de continuer de se réinventer pour poursuivre son objectif de transmission des connaissances scientifiques en biologie et au-delà. L'ensemble des participants du Centenaire de la SBS a ainsi posé la première pierre du Bicentenaire de la SBS.