Label-Free Phenotypic Profiling Identified D-Luciferin as a GPR35 Agonist

Fluorescent and luminescent probes are essential to both in vitro molecular assays and in vivo imaging techniques, and have been extensively used to measure biological function. However, little is known about the biological activity, thus potential interferences with the assay results, of these probe molecules. Here we show that D-luciferin, one of the most widely used bioluminescence substrates, is a partial agonist for G protein-coupled receptor-35 (GPR35). Label-free phenotypic profiling using dynamic mass redistribution (DMR) assays showed that D-luciferin led to a DMR signal in native HT-29 cells, whose characteristics are similar to those induced by known GPR35 agonists including zaprinast and pamoic acid. DMR assays further showed that D-luciferin is a partial agonist competitive to several known GPR35 agonists and antagonists. D-luciferin was found to cause the phosphorylation of ERK that was suppressed by known GPR35 antagonists, and also result in β-arrestin translocation signal but with low efficacy. These results not only suggest that D-luciferin is a partial agonist of GPR35, but also will evoke careful interpretation of biological data obtained using molecular and in vivo imaging assays when these probe molecules are used

Using C. elegans to discover therapeutic compounds for ageing-associated neurodegenerative diseases

Age-associated neurodegenerative disorders such as Alzheimer’s disease are a major public health challenge, due to the demographic increase in the proportion of older individuals in society. However, the relatively few currently approved drugs for these conditions provide only symptomatic relief. A major goal of neurodegeneration research is therefore to identify potential new therapeutic compounds that can slow or even reverse disease progression, either by impacting directly on the neurodegenerative process or by activating endogenous physiological neuroprotective mechanisms that decline with ageing. This requires model systems that can recapitulate key features of human neurodegenerative diseases that are also amenable to compound screening approaches. Mammalian models are very powerful, but are prohibitively expensive for high-throughput drug screens. Given the highly conserved neurological pathways between mammals and invertebrates, Caenorhabditis elegans has emerged as a powerful tool for neuroprotective compound screening. Here we describe how C. elegans has been used to model various human ageing-associated neurodegenerative diseases and provide an extensive list of compounds that have therapeutic activity in these worm models and so may have translational potential

Homo-PROTACs:bivalent small-molecule dimerizers of the VHL E3 ubiquitin ligase to induce self-degradation

E3 ubiquitin ligases are key enzymes within the ubiquitin proteasome system which catalyze the ubiquitination of proteins, targeting them for proteasomal degradation. E3 ligases are gaining importance as targets to small molecules, both for direct inhibition and to be hijacked to induce the degradation of non-native neo-substrates using bivalent compounds known as PROTACs (for 'proteolysis-targeting chimeras'). We describe Homo-PROTACs as an approach to dimerize an E3 ligase to trigger its suicide-type chemical knockdown inside cells. We provide proof-of-concept of Homo-PROTACs using diverse molecules composed of two instances of a ligand for the von Hippel-Lindau (VHL) E3 ligase. The most active compound, CM11, dimerizes VHL with high avidity in vitro and induces potent, rapid and proteasome-dependent self-degradation of VHL in different cell lines, in a highly isoform-selective fashion and without triggering a hypoxic response. This approach offers a novel chemical probe for selective VHL knockdown, and demonstrates the potential for a new modality of chemical intervention on E3 ligases.Targeting the ubiquitin proteasome system to modulate protein homeostasis using small molecules has promising therapeutic potential. Here the authors describe Homo-PROTACS: small molecules that can induce the homo-dimerization of E3 ubiquitin ligases and cause their proteasome-dependent degradation

Metaphors in Nanomedicine: The Case of Targeted Drug Delivery

International audienceThe promises of nanotechnology have been framed by a variety of metaphors, that not only channel the attention of the public, orient the questions asked by researchers, and convey epistemic choices closely linked to ethical preferences. In particular, the image of the 'therapeutic missile' commonly used to present targeted drug delivery devices emphasizes precision, control, surveillance and efficiency. Such values are highly praised in the current context of crisis of pharmaceutical innovation where military metaphors foster a general mobilization of resources from multiple fields of cutting-edge research. The missile metaphor, reminiscent of Paul Ehrlich's 'magic bullet', has framed the problem in simple terms: how to deliver the right dose in the right place at the right moment? Chemists, physicists and engineers who design multi-functional devices operating in vitro can think in such terms, as long as the devices are not actually operating through the messy environment of the body. A close look at what has been done and what remains to be done suggests that the metaphor of the "therapeutic missile" is neither sufficient, nor even necessary. Recent developments in nanomedicine suggest that therapeutic efficacy cannot be obtained without negotiating with the biological milieu and taking advantage of what it affords. An 'oïkological' approach seems more appropriate, more heuristic and more promising than the popular missile. It is based on the view of organism as an oikos that has to be carefully managed. The dispositions of nanocapsules have to be coupled with the affordances of the environment. As it requires dealing with nanoparticles as relational entities (defined by their potential for interactions) rather than as stable substances (defined by intrinsic properties) this metaphor eventually might well change research priorities in nanotechnology in general

High Throughput Screening for Small Molecule Enhancers of the Interferon Signaling Pathway to Drive Next-Generation Antiviral Drug Discovery

Most of current strategies for antiviral therapeutics target the virus specifically and directly, but an alternative approach to drug discovery might be to enhance the immune response to a broad range of viruses. Based on clinical observation in humans and successful genetic strategies in experimental models, we reasoned that an improved interferon (IFN) signaling system might better protect against viral infection. Here we aimed to identify small molecular weight compounds that might mimic this beneficial effect and improve antiviral defense. Accordingly, we developed a cell-based high-throughput screening (HTS) assay to identify small molecules that enhance the IFN signaling pathway components. The assay is based on a phenotypic screen for increased IFN-stimulated response element (ISRE) activity in a fully automated and robust format (Z′>0.7). Application of this assay system to a library of 2240 compounds (including 2160 already approved or approvable drugs) led to the identification of 64 compounds with significant ISRE activity. From these, we chose the anthracycline antibiotic, idarubicin, for further validation and mechanism based on activity in the sub-µM range. We found that idarubicin action to increase ISRE activity was manifest by other members of this drug class and was independent of cytotoxic or topoisomerase inhibitory effects as well as endogenous IFN signaling or production. We also observed that this compound conferred a consequent increase in IFN-stimulated gene (ISG) expression and a significant antiviral effect using a similar dose-range in a cell-culture system inoculated with encephalomyocarditis virus (EMCV). The antiviral effect was also found at compound concentrations below the ones observed for cytotoxicity. Taken together, our results provide proof of concept for using activators of components of the IFN signaling pathway to improve IFN efficacy and antiviral immune defense as well as a validated HTS approach to identify small molecules that might achieve this therapeutic benefit