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Agency: European Commission | Branch: FP7 | Program: CP-FP | Phase: HEALTH.2013.2.3.4-2 | Award Amount: 7.92M | Year: 2013

The trypanosomatid diseases, leishmaniasis, Human African trypanosomiasis (HAT) and Chagas disease (CD), continue to impart a heavy toll on human health. The treatments available are limited and threatened by drug resistance with few newdrugs in the pipeline. The KINDReD consortium integrates five leading academic laboratories in Europe (Portugal, United Kingdom, and Switzerland), the USA (California) and South America (Brazil) with high throughput screening (HTS) facilities equally distributed between all three major kinetoplastid parasites. Intracellular amastigote screening will be employed as the most relevant for Leishmania spp and T cruzi. Compound libraries (focused, diversity oriented or natural) will be screened in these systems, as well as compound series devised through target screening and in silico approaches. For carefully chosen protein targets, all three kinetoplastid parasite homologs will be screened against the closest human homolog to establish selectivity. Promising lead compounds will be optimised for efficacy and tolerability in cell-based and animal disease models. Toxicological markers will be evaluated in human cell lines prior to toxicity (acute,subacute,chronic) testing in lower then higher mammals. In parallel, and in line with the FDAs Critical Path Initiative, several check point controls will be built into the pipeline to flag, identify and allow early correction of potential toxicity/efficacy issues. These will include (i) a systems biology approach to identify drug target and off-target interactions via activity-based chemoproteomics (ii) uptake and metabolismas potential modulators of drug efficacy and/or resistance and (iii) the establishment of a firm set of rules for drug efficacy and safety in kinetoplastid chemotherapy. Our goal is to strengthen the drug development pipeline in order to achieve at least one new Phase I clinical candidate for each trypanosomatid disease at or shortly after the project completion date.

Hoffer L.,University of Strasbourg | Hoffer L.,Novalix Inc. | Horvath D.,University of Strasbourg
Journal of Chemical Information and Modeling | Year: 2013

S4MPLE is a conformational sampling tool, based on a hybrid genetic algorithm, simulating one (conformer enumeration) or more molecules (docking). Energy calculations are based on the AMBER force field [ Cornell et al. J. Am. Chem. Soc. 1995, 117, 5179. ] for biological macromolecules and its generalized version GAFF [ Wang et al. J. Comput. Chem. 2004, 25, 1157. ] for ligands. This paper describes more advanced, specific applications of S4MPLE to problems more complex than classical redocking of drug-like compounds [ Hoffer et al. J. Mol. Graphics Modell. 2012, submitted for publication. ]. Here, simultaneous docking of multiple entities is addressed in two different important contexts. First, simultaneous docking of two fragment-like ligands was attempted, as such ternary complexes are the basis of fragment-based drug design by linkage of the independent binders. As a preliminary, the capacity of S4MPLE to dock fragment-like compounds has been assessed, since this class of small probes used in fragment-based drug design covers a different chemical space than drug-like molecules. Herein reported success rates from fragments redocking are as good as classical benchmarking results on drug-like compounds (Astex Diverse Set [ Hartshorn et al. J. Med. Chem. 2007, 50, 726. ]). Then, S4MPLE is successfully challenged to predict locations of fragments involved in ternary complexes by means of multientity docking. Second, the key problem of predicting water-mediated interaction is addressed by considering explicit water molecules as additional entities to be docked in the presence of the "main" ligand. Blind prediction of solvent molecule positions, reproducing relevant ligand-water-site mediated interactions, is achieved in 76% cases over saved poses. S4MPLE was also successful to predict crystallographic water displacement by a therefore tailored functional group in the optimized ligand. However, water localization is an extremely delicate issue in terms of weighing of electrostatic and desolvation terms and also introduces a significant increase of required sampling efforts. Yet, the herein reported results - not making use of massively parallel deployment of the software - are very encouraging. © 2012 American Chemical Society.

Hoffer L.,University of Strasbourg | Hoffer L.,Novalix Inc. | Renaud J.-P.,Novalix Inc. | Horvath D.,University of Strasbourg
Journal of Chemical Information and Modeling | Year: 2013

This paper describes the use and validation of S4MPLE in Fragment-Based Drug Design (FBDD) -a strategy to build drug-like ligands starting from small compounds called fragments. S4MPLE is a conformational sampling tool based on a hybrid genetic algorithm that is able to simulate one (conformer enumeration) or more molecules (docking). The goal of the current paper is to show that due to the judicious design of genetic operators, S4MPLE may be used without any specific adaptation as an in silico FBDD tool. Such fragment-to-lead evolution involves either growing of one or linking of several fragment-like binder(s). The native ability to specifically "dock" a substructure that is covalently anchored to its target (here, some prepositioned fragment formally part of the binding site) enables it to act like dedicated de novo builders and differentiates it from most classical docking tools, which may only cope with non-covalent interactions. Besides, S4MPLE may address growing/linking scenarios involving protein site flexibility, and it might also suggest "growth" moves by bridging the ligand to the site via water-mediated interactions if H2O molecules are simply appended to the input files. Therefore, the only development overhead required to build a virtual fragment→ligand growing/linking strategy based on S4MPLE were two chemoinformatics programs meant to provide a minimalistic management of the linker library. The first creates a duplicate-free library by fragmenting a compound database, whereas the second builds new compounds, attaching chemically compatible linkers to the starting fragments. S4MPLE is subsequently used to probe the optimal placement of the linkers within the binding site, with initial restraints on atoms from initial fragments, followed by an optimization of all kept poses after restraint removal. Ranking is mainly based on two criteria: force-field potential energy and RMSD shifts of the original fragment moieties. This strategy was applied to several examples from the FBDD literature with good results over several monitored criteria: ability to generate the optimized ligand (or close analogs), good ranking of analogs among decoy compounds, and accurate predictions of expected binding modes of reference ligands. Simulations included "classical" covalent growing/linking, more challenging ones involving binding site conformational changes, and growth with optional recognition of putatively favorable water-mediated interactions. © 2013 American Chemical Society.

Ciesielski F.,University of Strasbourg | Ciesielski F.,Novalix Inc. | Sato Y.,University of Strasbourg | Sato Y.,Kyushu University | And 4 more authors.
Journal of Medicinal Chemistry | Year: 2012

Actual use of the active form of vitamin D (calcitriol or 1α,25-dihydroxyvitamin D3) to treat hyperproliferative disorders is hampered by calcemic effects, hence the continuous development of chemically modified analogues with dissociated profiles. Structurally distinct nonsecosteroidal analogues have been developed to mimic calcitriol activity profiles with low calcium serum levels. Here, we report the crystallographic study of vitamin D nuclear receptor (VDR) ligand binding domain in complexes with six nonsecosteroidal analogues harboring two or three phenyl rings. These compounds induce a stimulated transcription in the nanomolar range, similar to calcitriol. Examination of the protein-ligand interactions reveals the mode of binding of these nonsecosteroidal compounds and highlights the role of the various chemical modifications of the ligands to VDR binding and activity, notably (de)solvation effects. The structures with the tris-aromatic ligands exhibit a rearrangement of a novel region of the VDR ligand binding pocket, helix H6. © 2012 American Chemical Society.

Hoffer L.,University of Strasbourg | Hoffer L.,Novalix Inc. | Renaud J.-P.,Novalix Inc. | Horvath D.,University of Strasbourg
Combinatorial Chemistry and High Throughput Screening | Year: 2011

Fragment-based screening is an emerging technology which is used as an alternative to high-throughput screening (HTS), and often in parallel. Fragment screening focuses on very small compounds. Because of their small size and simplicity, fragments exhibit a low to medium binding affinity (mM to uM) and must therefore be screened at high concentration in order to detect binding events. Since some issues are associated with high-concentration screening in biochemical assays, biophysical methods are generally employed in fragment screening campaigns. Moreover, these techniques are very sensitive and some of them can give precise information about the binding mode of fragments, which facilitates the mandatory hit-to-lead optimization. One of the main advantages of fragment-based screening is that fragment hits generally exhibit a strong binding with respect to their size, and their subsequent optimization should lead to compounds with better pharmacokinetic properties compared to molecules evolved from HTS hits. In other words, fragments are interesting starting points for drug discovery projects. Besides, the chemical space of low-complexity compounds is very limited in comparison to that of drug-like molecules, and thus easier to explore with a screening library of limited size. Furthermore, the "combinatorial explosion" effect ensures that the resulting combinations of interlinked binding fragments may cover a significant part of "drug-like" chemical space. In parallel to experimental screening, virtual screening techniques, dedicated to fragments or wider compounds, are gaining momentum in order to further reduce the number of compounds to test. This article is a review of the latest news in both experimental and in silico virtual screening in the fragment-based discovery field. Given the specificity of this journal, special attention will be given to fragment library design. © 2011 Bentham Science Publishers Ltd.

Huet T.,University of Strasbourg | Huet T.,Cbs Therapeutics, Inc. | Laverny G.,University of Strasbourg | Ciesielski F.,University of Strasbourg | And 11 more authors.
Cell Reports | Year: 2015

The bioactive form of vitamin D [1,25(OH)2D3] regulates mineral and bone homeostasis and exerts potent anti-inflammatory and antiproliferative properties through binding to the vitamin D receptor (VDR). The 3D structures of the VDR ligand-binding domain with 1,25(OH)2D3 or gemini analogs unveiled the molecular mechanism underlying ligand recognition. On the basis of structure-function correlations, we generated a point-mutated VDR (VDRgem) that isunresponsive to 1,25(OH)2D3, but the activity of which is efficiently induced by the gemini ligands. Moreover, we show that many VDR target genes are repressed by unliganded VDRgem and that mineral ion and bone homeostasis are more impaired in VDRgem mice than in VDR null mice, demonstrating that mutations abolishing VDR ligand binding result in more severe skeletal defects than VDR null mutations. As gemini ligands induce VDRgem transcriptional activity in mice and normalize their serum calcium levels, VDRgem is a powerful tool to further unravel both liganded and unliganded VDR signaling. © 2015 The Authors.

Mendoza A.,National Autonomous University of Mexico | Navarrete-Ramirez P.,National Autonomous University of Mexico | Hernandez-Puga G.,National Autonomous University of Mexico | Villalobos P.,National Autonomous University of Mexico | And 4 more authors.
Endocrinology | Year: 2013

Several liganded nuclear receptors have alternative ligands acting in a tissue-specific fashion and playing important biological roles. We present evidence that 3,5-diiodothyronine (T2), a naturally occurring iodothyronine that results from T3 outer-ring deiodination, is an alternative ligand for thyroid hormone receptor β1 (TRβ1). In tilapia, 2 TRβ isoforms differing by 9 amino acids in the ligand-binding domain were cloned. Binding and transactivation studies showed that T2 activates the human and the long tilapia TRβ1 isoform, but not the short one. A chimeric human TRβ1 (hTRβ1) that contained the 9-amino-acid insert showed no response to T2, suggesting that the conformation of the hTRβ1 naturally allows T2 binding and that other regions of the receptor are implicated in TR activation by T2. Indeed, further analysis showed that the N terminus is essential for T2-mediated transactivation but not for that by T3 in the long and hTRβ1, suggesting a functional interaction between the N-terminal domain and the insertion in the ligand-binding domain. To establish the functional relevance of T2-mediated TRβ1 binding and activation, mRNA expression and its regulation by T2 and T3 was evaluated for both isoforms. Our data show that long TRβ1expression is 106-fold higher than that of the short isoform, and T3 and T2 differentially regulate the expression of these 2 TRβ1 isoforms in vivo. Taken together, our results prompted a reevaluation of the role and mechanism of action of thyroid hormone metabolites previously believed to be inactive. More generally, we propose that classical liganded receptors are only partially locked to very specific ligands and that alternative ligands may play a role in the tissuespecific action of receptors. © 2013 by The Endocrine Society.

Reverdy C.,Hybrigenics | Conrath S.,Hybrigenics | Lopez R.,Hybrigenics | Planquette C.,Hybrigenics | And 7 more authors.
Chemistry and Biology | Year: 2012

The human USP7 deubiquitinating enzyme was shown to regulate many proteins involved in the cell cycle, as well as tumor suppressors and oncogenes. Thus, USP7 offers a promising, strategic target for cancer therapy. Using biochemical assays and activity-based protein profiling in living systems, we identified small-molecule antagonists of USP7 and demonstrated USP7 inhibitor occupancy and selectivity in cancer cell lines. These compounds bind USP7 in the active site through a covalent mechanism. In cancer cells, these active-site-targeting inhibitors were shown to regulate the level of several USP7 substrates and thus recapitulated the USP7 knockdown phenotype that leads to G1 arrest in colon cancer cells. The data presented in this report provide proof of principle that USP7 inhibitors may be a valuable therapeutic for cancer. In addition, the discovery of such molecules offers interesting tools for studying deubiquitination. © 2012 Elsevier Ltd.

Barre B.,Paris West University Nanterre La Défense | Gonnard L.,Paris West University Nanterre La Défense | Campagne R.,Paris West University Nanterre La Défense | Reymond S.,Paris West University Nanterre La Défense | And 5 more authors.
Organic Letters | Year: 2014

Iron- and cobalt-catalyzed cross-couplings between iodo-azetidines, -pyrrolidines, -piperidines, and Grignard reagents are disclosed. The reaction is efficient, cheap, chemoselective and tolerates a large variety of (hetero)aryl Grignard reagents. © 2014 American Chemical Society.

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