Accelerating Chemical Tool Discovery by Academic Collaborative Models

The development of chemical tool compounds becomes increasingly important for chemical biology research projects in many disciplines of life sciences. In addition, they form essential parts in both academic and industrial drug discovery efforts. The required expertise and technology platforms for the identification and optimization of these potent and target-selective small molecules often exceed the capabilities of academic groups and smaller companies. Over the years, several initiatives were created all over the world which address this issue by either creating networks or consortia of academic institutes, public-private partnerships with industry, or even dedicated new research infrastructures for chemical biology. Several of these organizations and their different access models will be described. We will focus in particular on the model of EU-OPENSCREEN ERIC, a new European Research Infrastructure which was founded in 2018 and consists of more than 20 partner institutes from eight countries

Pathogen and Toxin Entry - How Pathogens and Toxins Induce and Harness Endocytotic Mechanisms

Humans have been exposed to a plethora of pathogens (bacteria, viruses) ever since. Infectious diseases are among the leading causes of death worldwide. For example, in 2011, 1.34 million people died of tuberculosis, which is caused by an infection with Mycobacterium tuberculosis. Even more died of an infection by the human immunodeficiency virus (HIV; 1.78 million) or lower respiratory tract infection (3.46 million) [1]. In addition, recurring pandemic outbreaks of the influenza A virus, as in 2009, or an epidemic outbreak of enterohemorrhagic E. coli (EHEC) in Germany in 2011, show quite plainly that pathogens in the 21th century still are a severe health problem, not only in developing countries

Lipid Reorganization Induced by Shiga Toxin Clustering on Planar Membranes

The homopentameric B-subunit of bacterial protein Shiga toxin (STxB) binds to the glycolipid Gb3 in plasma membranes, which is the initial step for entering cells by a clathrin-independent mechanism. It has been suggested that protein clustering and lipid reorganization determine toxin uptake into cells. Here, we elucidated the molecular requirements for STxB induced Gb3 clustering and for the proposed lipid reorganization in planar membranes. The influence of binding site III of the B-subunit as well as the Gb3 lipid structure was investigated by means of high resolution methods such as fluorescence and scanning force microscopy. STxB was found to form protein clusters on homogenous 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)/cholesterol/Gb3 (65∶30∶5) bilayers. In contrast, membranes composed of DOPC/cholesterol/sphingomyelin/Gb3 (40∶35∶20∶5) phase separate into a liquid ordered and liquid disordered phase. Dependent on the fatty acid composition of Gb3, STxB-Gb3 complexes organize within the liquid ordered phase upon protein binding. Our findings suggest that STxB is capable of forming a new membrane phase that is characterized by lipid compaction. The significance of this finding is discussed in the context of Shiga toxin-induced formation of endocytic membrane invaginations

Identification et caractérisationde petites molécules qui imbibent l'entrée de toxines par la voie du transport rétrograde

LE KREMLIN-B.- PARIS 11-BU Méd (940432101) / SudocSudocFranceF

Chapter 10 Pathogen and Toxin Entry - How Pathogens and Toxins Induce and Harness Endocytotic Mechanisms

Humans have been exposed to a plethora of pathogens (bacteria, viruses) ever since. Infectious diseases are among the leading causes of death worldwide. For example, in 2011, 1.34 million people died of tuberculosis, which is caused by an infection with Mycobacterium tuberculosis. Even more died of an infection by the human immunodeficiency virus (HIV; 1.78 million) or lower respiratory tract infection (3.46 million) [1]. In addition, recurring pandemic outbreaks of the influenza A virus, as in 2009, or an epidemic outbreak of enterohemorrhagic E. coli (EHEC) in Germany in 2011, show quite plainly that pathogens in the 21th century still are a severe health problem, not only in developing countries

EU-OPENSCREEN-chemical tools for the study of plant biology and resistance mechanisms.

EU-OPENSCREEN is an academic research infrastructure initiative in Europe for enabling researchers in all life sciences to take advantage of chemical biology approaches to their projects. In a collaborative effort of national networks in 16 European countries, EU-OPENSCREEN will develop novel chemical compounds with external users to address questions in, among other fields, systems and network biology (directed and selective perturbation of signalling pathways), structural biology (compound-target interactions at atomic resolution), pharmacology (early drug discovery and toxicology) and plant biology (response of wild or crop plants to environmental and agricultural substances). EU-OPENSCREEN supports all stages of a tool development project, including assay adaptation, high-throughput screening and chemical optimisation of the 'hit' compounds. All tool compounds and data will be made available to the scientific community. EU-OPENSCREEN integrates high-capacity screening platforms throughout Europe, which share a rationally selected compound collection comprising up to 300,000 (commercial and proprietary compounds collected from European chemists). By testing systematically this chemical collection in hundreds of assays originating from very different biological themes, the screening process generates enormous amounts of information about the biological activities of the substances and thereby steadily enriches our understanding of how and where they act

Academic collaborative models fostering the translation of physiological in vitro systems from basic research into drug discovery.

The success of preclinical drug discovery strongly relies on the ability of experimental models to resemble human pathophysiology. The number of compounds receiving approval for clinical use is limited, and this has led to the development of more physiologically relevant cellular models aimed at making preclinical results more prone to be successfully translated into clinical use. In this review, we summarize the technologies available in the field of high-throughput screening (HTS) using complex cellular models, and describe collaborative initiatives, such as EU-OPENSCREEN, which can efficiently support researchers to easily access state-of-the-art chemical biology platforms for improving the drug discovery process

Inhibition of retrograde transport protects mice from lethal ricin challenge.

International audienceBacterial Shiga-like toxins are virulence factors that constitute a significant public health threat worldwide, and the plant toxin ricin is a potential bioterror weapon. To gain access to their cytosolic target, ribosomal RNA, these toxins follow the retrograde transport route from the plasma membrane to the endoplasmic reticulum, via endosomes and the Golgi apparatus. Here, we used high-throughput screening to identify small molecule inhibitors that protect cells from ricin and Shiga-like toxins. We identified two compounds that selectively block retrograde toxin trafficking at the early endosome-TGN interface, without affecting compartment morphology, endogenous retrograde cargos, or other trafficking steps, demonstrating an unexpected degree of selectivity and lack of toxicity. In mice, one compound clearly protects from lethal nasal exposure to ricin. Our work discovers the first small molecule that shows efficacy against ricin in animal experiments and identifies the retrograde route as a potential therapeutic target

Retrograde transport is not required for cytosolic translocation of the B-subunit of Shiga toxin.

Antigen-presenting cells have the remarkable capacity to transfer exogenous antigens to the cytosol for processing by proteasomes and subsequent presentation on major histocompatibility complex class-I (MHC-I) molecules, a process termed cross-presentation. This is the target of biomedical approaches that aim to trigger a therapeutic immune response. The receptor-binding B-subunit of Shiga toxin (STxB) has been developed as an antigen delivery tool for such immunotherapy applications. In this study, we have analyzed pathways and trafficking factors that are involved in this process. A covalent conjugate between STxB and saporin was generated to quantitatively sample the membrane translocation step to the cytosol in differentiated monocyte-derived THP-1 cells. We have found that retrograde trafficking to the Golgi complex was not required for STxB-saporin translocation to the cytosol or for STxB-dependent antigen cross-presentation. Depletion of endosomal Rab7 inhibited, and lowering membrane cholesterol levels favored STxB-saporin translocation. Interestingly, experiments with reducible and non-reducible linker-arm-STxB conjugates led to the conclusion that after translocation, STxB remains associated with the cytosolic membrane leaflet. In summary, we report new facets of the endosomal escape process bearing relevance to antigen cross-presentation