Publications by authors named "Claire Colas"

24 Publications

  • Page 1 of 1

Targeting Solute Carrier Transporters through Functional Mapping.

Trends Pharmacol Sci 2021 01 21;42(1):3-6. Epub 2020 Nov 21.

Sorbonne Université, UPMC University Paris 06, CNRS, IBPS, UMR 7238, Laboratoire de Biologie Computationnelle et Quantitative (LCQB), 75005 Paris, France.

Solute carrier (SLC) transporters are emerging drug targets. Identifying the molecular determinants responsible for their specific and selective transport activities and describing key interactions with their ligands are crucial steps towards the design of potential new drugs. A general functional mapping across more than 400 human SLC transporters would pave the way to the rational and systematic design of molecules modulating cellular transport.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.tips.2020.11.005DOI Listing
January 2021

ATP modulates SLC7A5 (LAT1) synergistically with cholesterol.

Sci Rep 2020 10 7;10(1):16738. Epub 2020 Oct 7.

Department of DiBEST (Biologia, Ecologia, Scienze Della Terra) Unit of Biochemistry and Molecular Biotechnology, University of Calabria, via Bucci 4C, 87036, Arcavacata di Rende, Italy.

The plasma membrane transporter hLAT1 is responsible for providing cells with essential amino acids. hLAT1 is over-expressed in virtually all human cancers making the protein a hot-spot in the fields of cancer and pharmacology research. However, regulatory aspects of hLAT1 biology are still poorly understood. A remarkable stimulation of transport activity was observed in the presence of physiological levels of cholesterol together with a selective increase of the affinity for the substrate on the internal site, suggesting a stabilization of the inward open conformation of hLAT1. A synergistic effect by ATP was also observed only in the presence of cholesterol. The same phenomenon was detected with the native protein. Altogether, the biochemical assays suggested that cholesterol and ATP binding sites are close to each other. The computational analysis identified two neighboring regions, one hydrophobic and one hydrophilic, to which cholesterol and ATP were docked, respectively. The computational data predicted interaction of the ϒ-phosphate of ATP with Lys 204, which was confirmed by site-directed mutagenesis. The hLAT1-K204Q mutant showed an impaired function and response to ATP. Interestingly, this residue is conserved in several members of the SLC7 family.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-73757-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7541457PMC
October 2020

Toward a Systematic Structural and Functional Annotation of Solute Carriers Transporters-Example of the SLC6 and SLC7 Families.

Authors:
Claire Colas

Front Pharmacol 2020 19;11:1229. Epub 2020 Aug 19.

University of Vienna, Department of Pharmaceutical Chemistry, Vienna, Austria.

SLC transporters are emerging key drug targets. One important step for drug development is the profound understanding of the structural determinants defining the substrate selectivity of each transporter. Recently, the improvement of computational power and experimental methods such as X-ray and cryo-EM crystallography permitted to conduct structure-based studies on specific transporters having important pharmacological impact. However, a lot remains to be discovered regarding their dynamics, transport modulation and ligand recognition. A detailed functional characterization of transporters would provide opportunities to develop new compounds targeting these key drug targets. Here, we are giving an overview of two major human LeuT-fold families, SLC6 and SLC7, with an emphasis on the most relevant members of each family for drug development. We gather the most recent understanding on the structural determinants of selectivity within and across the two families. We then use this information to discuss the benefits of a more generalized structural and functional annotation of the LeuT fold and the implications of such mapping for drug discovery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fphar.2020.01229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7466448PMC
August 2020

Studies of structural determinants of substrate binding in the Creatine Transporter (CreaT, SLC6A8) using molecular models.

Sci Rep 2020 04 10;10(1):6241. Epub 2020 Apr 10.

University of Vienna, Department of Pharmaceutical Chemistry, Vienna, Austria.

Creatine is a crucial metabolite that plays a fundamental role in ATP homeostasis in tissues with high-energy demands. The creatine transporter (CreaT, SLC6A8) belongs to the solute carrier 6 (SLC6) transporters family, and more particularly to the GABA transporters (GATs) subfamily. Understanding the molecular determinants of specificity within the SLC6 transporters in general, and the GATs in particular is very challenging due to the high similarity of these proteins. In the study presented here, our efforts focused on finding key structural features involved in binding selectivity for CreaT using structure-based computational methods. Due to the lack of three-dimensional structures of SLC6A8, our approach was based on the realization of two reliable homology models of CreaT using the structures of two templates, i.e. the human serotonin transporter (hSERT) and the prokaryotic leucine transporter (LeuT). Our models reveal that an optimal complementarity between the shape of the binding site and the size of the ligands is necessary for transport. These findings provide a framework for a deeper understanding of substrate selectivity of the SLC6 family and other LeuT fold transporters.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-63189-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7148354PMC
April 2020

The RESOLUTE consortium: unlocking SLC transporters for drug discovery.

Authors:
Giulio Superti-Furga Daniel Lackner Tabea Wiedmer Alvaro Ingles-Prieto Barbara Barbosa Enrico Girardi Ulrich Goldmann Bettina Gürtl Kristaps Klavins Christoph Klimek Sabrina Lindinger Eva Liñeiro-Retes André C Müller Svenja Onstein Gregor Redinger Daniela Reil Vitaly Sedlyarov Gernot Wolf Matthew Crawford Robert Everley David Hepworth Shenping Liu Stephen Noell Mary Piotrowski Robert Stanton Hui Zhang Salvatore Corallino Andrea Faedo Maria Insidioso Giovanna Maresca Loredana Redaelli Francesca Sassone Lia Scarabottolo Michela Stucchi Paola Tarroni Sara Tremolada Helena Batoulis Andreas Becker Eckhard Bender Yung-Ning Chang Alexander Ehrmann Anke Müller-Fahrnow Vera Pütter Diana Zindel Bradford Hamilton Martin Lenter Diana Santacruz Coralie Viollet Charles Whitehurst Kai Johnsson Philipp Leippe Birgit Baumgarten Lena Chang Yvonne Ibig Martin Pfeifer Jürgen Reinhardt Julian Schönbett Paul Selzer Klaus Seuwen Charles Bettembourg Bruno Biton Jörg Czech Hélène de Foucauld Michel Didier Thomas Licher Vincent Mikol Antje Pommereau Frédéric Puech Veeranagouda Yaligara Aled Edwards Brandon J Bongers Laura H Heitman Ad P IJzerman Huub J Sijben Gerard J P van Westen Justine Grixti Douglas B Kell Farah Mughal Neil Swainston Marina Wright-Muelas Tina Bohstedt Nicola Burgess-Brown Liz Carpenter Katharina Dürr Jesper Hansen Andreea Scacioc Giulia Banci Claire Colas Daniela Digles Gerhard Ecker Barbara Füzi Viktoria Gamsjäger Melanie Grandits Riccardo Martini Florentina Troger Patrick Altermatt Cédric Doucerain Franz Dürrenberger Vania Manolova Anna-Lena Steck Hanna Sundström Maria Wilhelm Claire M Steppan

Nat Rev Drug Discov 2020 07;19(7):429-430

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/d41573-020-00056-6DOI Listing
July 2020

l-Type amino acid transporter 1 activity of 1,2,3-triazolyl analogs of l-histidine and l-tryptophan.

Bioorg Med Chem Lett 2019 08 20;29(16):2254-2258. Epub 2019 Jun 20.

Department of Chemistry, University of Nebraska at Kearney, Kearney, NE 69949, USA. Electronic address:

A series of 1,2,3-triazole analogs of the amino acids l-histidine and l-tryptophan were modeled, synthesized and tested for l-type amino acid transporter 1 (LAT1; SLC7A5) activity to guide the design of amino acid-drug conjugates (prodrugs). These triazoles were conveniently prepared by the highly convergent Huisgen 1,3-dipolar cycloaddition (Click Chemistry). Despite comparable predicted binding modes, triazoles generally demonstrated reduced cell uptake and LAT1 binding potency relative to their natural amino acid counterparts. The structure-activity relationship (SAR) data for these triazoles has important ramifications for treating cancer and brain disorders using amino acid prodrugs or LAT1 inhibitors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bmcl.2019.06.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6690782PMC
August 2019

Homology Modeling Informs Ligand Discovery for the Glutamine Transporter ASCT2.

Front Chem 2018 24;6:279. Epub 2018 Jul 24.

Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States.

The Alanine-Serine-Cysteine transporter (SLC1A5, ASCT2), is a neutral amino acid exchanger involved in the intracellular homeostasis of amino acids in peripheral tissues. Given its role in supplying glutamine to rapidly proliferating cancer cells in several tumor types such as triple-negative breast cancer and melanoma, ASCT2 has been identified as a key drug target. Here we use a range of computational methods, including homology modeling and ligand docking, in combination with cell-based assays, to develop hypotheses for structure-function relationships in ASCT2. We perform a phylogenetic analysis of the SLC1 family and its prokaryotic homologs to develop a useful multiple sequence alignment for this protein family. We then generate homology models of ASCT2 in two different conformations, based on the human EAAT1 structures. Using ligand enrichment calculations, the ASCT2 models are then compared to crystal structures of various homologs for their utility in discovering ASCT2 inhibitors. We use virtual screening, cellular uptake and electrophysiology experiments to identify a non-amino acid ASCT2 inhibitor that is predicted to interact with the ASCT2 substrate binding site. Our results provide insights into the structural basis of substrate specificity in the SLC1 family, as well as a framework for the design of future selective and potent ASCT2 inhibitors as cancer therapeutics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fchem.2018.00279DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6066518PMC
July 2018

Reevaluating the Substrate Specificity of the L-Type Amino Acid Transporter (LAT1).

J Med Chem 2018 08 10;61(16):7358-7373. Epub 2018 Aug 10.

Department of Chemistry , University of Nebraska at Kearney , Kearney , Nebraska 68849 , United States.

The L-type amino acid transporter 1 (LAT1, SLC7A5) transports essential amino acids across the blood-brain barrier (BBB) and into cancer cells. To utilize LAT1 for drug delivery, potent amino acid promoieties are desired, as prodrugs must compete with millimolar concentrations of endogenous amino acids. To better understand ligand-transporter interactions that could improve potency, we developed structural LAT1 models to guide the design of substituted analogues of phenylalanine and histidine. Furthermore, we evaluated the structure-activity relationship (SAR) for both enantiomers of naturally occurring LAT1 substrates. Analogues were tested in cis-inhibition and trans-stimulation cell assays to determine potency and uptake rate. Surprisingly, LAT1 can transport amino acid-like substrates with wide-ranging polarities including those containing ionizable substituents. Additionally, the rate of LAT1 transport was generally nonstereoselective even though enantiomers likely exhibit different binding modes. Our findings have broad implications to the development of new treatments for brain disorders and cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jmedchem.8b01007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6668346PMC
August 2018

Effects of Mutations and Ligands on the Thermostability of the l-Arginine/Agmatine Antiporter AdiC and Deduced Insights into Ligand-Binding of Human l-Type Amino Acid Transporters.

Int J Mol Sci 2018 Mar 20;19(3). Epub 2018 Mar 20.

Institute of Biochemistry and Molecular Medicine, and Swiss National Centre of Competence in Research (NCCR) TransCure, University of Bern, CH-3012 Bern, Switzerland.

The l-arginine/agmatine transporter AdiC is a prokaryotic member of the SLC7 family, which enables pathogenic enterobacteria to survive the extremely acidic gastric environment. Wild-type AdiC from as well as its previously reported point mutants N22A and S26A, were overexpressed homologously and purified to homogeneity. A size-exclusion chromatography-based thermostability assay was used to determine the melting temperatures (s) of the purified AdiC variants in the absence and presence of the selected ligands l-arginine (Arg), agmatine, l-arginine methyl ester, and l-arginine amide. The resulting s indicated stabilization of AdiC variants upon ligand binding, in which s and ligand binding affinities correlated positively. Considering results from this and previous studies, we revisited the role of AdiC residue S26 in Arg binding and proposed interactions of the α-carboxylate group of Arg exclusively with amide groups of the AdiC backbone. In the context of substrate binding in the human SLC7 family member l-type amino acid transporter-1 (LAT1; SLC7A5), an analogous role of S66 in LAT1 to S26 in AdiC is discussed based on homology modeling and amino acid sequence analysis. Finally, we propose a binding mechanism for l-amino acid substrates to LATs from the SLC7 family.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ijms19030918DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5877779PMC
March 2018

Chemical Modulation of the Human Oligopeptide Transporter 1, hPepT1.

Mol Pharm 2017 12 15;14(12):4685-4693. Epub 2017 Nov 15.

Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai , New York, New York 10029, United States.

In humans, peptides derived from dietary proteins and peptide-like drugs are transported via the proton-dependent oligopeptide transporter hPepT1 (SLC15A1). hPepT1 is located across the apical membranes of the small intestine and kidney, where it serves as a high-capacity low-affinity transporter of a broad range of di- and tripeptides. hPepT1 is also overexpressed in the colon of inflammatory bowel disease (IBD) patients, where it mediates the transport of harmful peptides of bacterial origin. Therefore, hPepT1 is a drug target for prodrug substrates interacting with intracellular proteins or inhibitors blocking the transport of toxic bacterial products. In this study, we construct multiple structural models of hPepT1 representing different conformational states that occur during transport and inhibition. We then identify and characterize five ligands of hPepT1 using computational methods, such as virtual screening and QM-polarized ligand docking (QPLD), and experimental testing with uptake kinetic measurements and electrophysiological assays. Our results improve our understanding of the substrate and inhibitor specificity of hPepT1. Furthermore, the newly discovered ligands exhibit unique chemotypes, providing a framework for developing tool compounds with optimal intestinal absorption as well as future IBD therapeutics against this emerging drug target.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.molpharmaceut.7b00775DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5826771PMC
December 2017

Mapping Functionally Important Residues in the Na/Dicarboxylate Cotransporter, NaDC1.

Biochemistry 2017 08 10;56(33):4432-4441. Epub 2017 Aug 10.

Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California-San Diego , La Jolla, California 92130-0714, United States.

Transporters from the SLC13 family couple the transport of two to four Na ions with a di- or tricarboxylate, such as succinate or citrate. We have previously modeled mammalian members of the SLC13 family, including the Na/dicarboxylate cotransporter NaDC1 (SLC13A2), based on a structure of the bacterial homologue VcINDY in an inward-facing conformation with one sodium ion bound at the Na1 site. In the study presented here, we modeled the outward-facing conformation of rabbit and human NaDC1 (rbNaDC1 and hNaDC1, respectively) using an outward-facing model of VcINDY as a template and identified residues in or near the putative Na2 and Na3 cation binding sites. Guided by the structural models in both conformations, we performed site-directed mutagenesis in rbNaDC1 for residues proposed to be in the Na or substrate binding sites. Cysteine substitution of T474 in the predicted Na2 binding site results in an inactive protein. The M539C mutant has a low apparent affinity for both sodium and lithium cations, suggesting that M539 may form part of the putative Na3 binding site. The Y432C and T86C mutants have increased K values for succinate, supporting their proposed location in the outward-facing substrate binding site. In addition, cysteine labeling by MTSEA-biotin shows that Y432C is accessible from the outside of the cell, and the accessibility changes in the presence or absence of Na. The results of this study improve our understanding of substrate and ion recognition in the mammalian members of the SLC13 family and provide a framework for developing conformationally specific inhibitors against these transporters.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.biochem.7b00503DOI Listing
August 2017

Structure activity relationships of benzylproline-derived inhibitors of the glutamine transporter ASCT2.

Bioorg Med Chem Lett 2017 02 27;27(3):398-402. Epub 2016 Dec 27.

Department of Chemistry, Binghamton University, 4400 Vestal Pkwy East, Binghamton, NY 13902, United States. Electronic address:

The glutamine transporter ASCT2 has been identified as a promising target to inhibit rapid growth of cancer cells. However, ASCT2 pharmacology is not well established. In this report, we performed a systematic structure activity analysis of a series of substituted benzylproline derivatives. Substitutions on the phenyl ring resulted in compounds with characteristics of ASCT2 inhibitors. Apparent binding affinity increased with increasing hydrophobicity of the side chain. In contrast, interaction of the ASCT2 binding site with specific positions on the phenyl ring was not observed. The most potent compound inhibits the ASCT2 anion conductance with a K of 3μM, which is in the same range as that of more bulky and higher molecular weight inhibitors recently reported by others. The experimental results are consistent with computational analysis based on docking of the inhibitors against an ASCT2 homology model. The benzylproline scaffold provides a valuable tool for further improving binding potency of future ASCT2 inhibitors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bmcl.2016.12.063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5363072PMC
February 2017

SLC Transporters: Structure, Function, and Drug Discovery.

Medchemcomm 2016 Jun 28;7(6):1069-1081. Epub 2016 Mar 28.

Department of Pharmacology and Systems Therapeutics, Icahn School of Medicine at Mount Sinai, New York, NY 10029.; Department of Structural and Chemical Biology, Icahn School of Medicine at Mount Sinai, New York, NY 10029.

The human Solute Carrier (SLC) transporters are important targets for drug development. Structure-based drug discovery for SLC transporters requires the description of their structure, dynamics, and mechanism of interaction with small molecule ligands and ions. The recent determination of atomic structures of human SLC transporters and their homologs, combined with improved computational power and prediction methods have led to an increased applicability of structure-based drug design methods for human SLC members. In this review, we provide an overview of the SLC transporters' structures and transport mechanisms. We then describe computational techniques, such as homology modeling and virtual screening that are emerging as key tools to discover chemical probes for human SLC members. We illustrate the utility of these methods by presenting case studies in which rational integration of computation and experiment was used to characterize SLC members that transport key nutrients and metabolites, including the amino acid transporters LAT-1 and ASCT2, the SLC13 family of citric acid cycle intermediate transporters, and the glucose transporter GLUT1. We conclude with a brief discussion about future directions in structure-based drug discovery for the human SLC superfamily, one of the most structurally and functionally diverse protein families in human.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/C6MD00005CDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034948PMC
June 2016

LAT1 activity of carboxylic acid bioisosteres: Evaluation of hydroxamic acids as substrates.

Bioorg Med Chem Lett 2016 10 3;26(20):5000-5006. Epub 2016 Sep 3.

Department of Chemistry, University of Nebraska Kearney, Kearney, NE 68849, United States. Electronic address:

Large neutral amino acid transporter 1 (LAT1) is a solute carrier protein located primarily in the blood-brain barrier (BBB) that offers the potential to deliver drugs to the brain. It is also up-regulated in cancer cells, as part of a tumor's increased metabolic demands. Previously, amino acid prodrugs have been shown to be transported by LAT1. Carboxylic acid bioisosteres may afford prodrugs with an altered physicochemical and pharmacokinetic profile than those derived from natural amino acids, allowing for higher brain or tumor levels of drug and/or lower toxicity. The effect of replacing phenylalanine's carboxylic acid with a tetrazole, acylsulfonamide and hydroxamic acid (HA) bioisostere was examined. Compounds were tested for their ability to be LAT1 substrates using both cis-inhibition and trans-stimulation cell assays. As HA-Phe demonstrated weak substrate activity, its structure-activity relationship (SAR) was further explored by synthesis and testing of HA derivatives of other LAT1 amino acid substrates (i.e., Tyr, Leu, Ile, and Met). The potential for a false positive in the trans-stimulation assay caused by parent amino acid was evaluated by conducting compound stability experiments for both HA-Leu and the corresponding methyl ester derivative. We concluded that HA's are transported by LAT1. In addition, our results lend support to a recent account that amino acid esters are LAT1 substrates, and that hydrogen bonding may be as important as charge for interaction with the transporter binding site.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5076878PMC
http://dx.doi.org/10.1016/j.bmcl.2016.09.001DOI Listing
October 2016

Mutations in the Na(+)/citrate cotransporter NaCT (SLC13A5) in pediatric patients with epilepsy and developmental delay.

Mol Med 2016 05 26;22. Epub 2016 May 26.

Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California-San Diego, La Jolla, CA 92130-0718.

Mutations in the SLC13A5 gene that codes for the Na(+)/citrate cotransporter, NaCT, are associated with early onset epilepsy, developmental delay and tooth dysplasia in children. In the present study we identify additional SLC13A5 mutations in nine epilepsy patients from six families. To better characterize the syndrome, families with affected children answered questions about the scope of illness and treatment strategies. There are currently no effective treatments, but some anti-epileptic drugs targeting the GABA system reduce seizure frequency. Acetazolamide, a carbonic anhydrase inhibitor and atypical anti-seizure medication decreases seizures in 4 patients. In contrast to previous reports, the ketogenic diet and fasting produce worsening of symptoms. The effects of the mutations on NaCT transport function and protein expression were examined by transient transfections of COS-7 cells. There was no transport activity from any of the mutant transporters, although some of the mutant transporter proteins were present on the plasma membrane. The structural model of NaCT suggests that these mutations can affect helix packing or substrate binding. We tested various treatments, including chemical chaperones and low temperatures, but none improve transport function in the NaCT mutants. Interestingly, coexpression of NaCT and the mutants results in decreased protein expression and activity of the wild-type transporter, indicating functional interaction. In conclusion, our study has identified additional SLC13A5 mutations in patients with chronic epilepsy starting in the neonatal period, with the mutations producing inactive Na(+)/citrate transporters.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2119/molmed.2016.00077DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5023510PMC
May 2016

LAT-1 activity of meta-substituted phenylalanine and tyrosine analogs.

Bioorg Med Chem Lett 2016 06 11;26(11):2616-2621. Epub 2016 Apr 11.

Department of Chemistry, University of Nebraska Kearney, Kearney, NE, 68849.

The transporter protein Large-neutral Amino Acid Transporter 1 (LAT-1, SLC7A5) is responsible for transporting amino acids such as tyrosine and phenylalanine as well as thyroid hormones, and it has been exploited as a drug delivery mechanism. Recently its role in cancer has become increasingly appreciated, as it has been found to be up-regulated in many different tumor types, and its expression levels have been correlated with prognosis. Substitution at the meta position of aromatic amino acids has been reported to increase affinity for LAT-1; however, the SAR for this position has not previously been explored. Guided by newly refined computational models of the binding site, we hypothesized that groups capable of filling a hydrophobic pocket would increase binding to LAT-1, resulting in improved substrates relative to parent amino acid. Tyrosine and phenylalanine analogs substituted at the meta position with halogens, alkyl and aryl groups were synthesized and tested in cis-inhibition and trans-stimulation cell assays to determine activity. Contrary to our initial hypothesis we found that lipophilicity was correlated with diminished substrate activity and increased inhibition of the transporter. The synthesis and SAR of meta-substituted phenylalanine and tyrosine analogs is described.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bmcl.2016.04.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4875506PMC
June 2016

Computing Substrate Selectivity in a Peptide Transporter.

Cell Chem Biol 2016 Feb;23(2):211-213

Departments of Pharmacology and Systems Therapeutics, and Structural and Chemical Biology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA. Electronic address:

The human proton-coupled peptide transporter 1 (PepT1) is responsible for the absorption of di- and tri-peptides from the diet and peptide-like drugs. In this issue of Cell Chemical Biology, Samsudin et al. (2016) use an integrated computational and experimental approach to provide new insights into understanding substrate selectivity of PepTSt, a prokaryotic homolog of the human PepT1.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.chembiol.2016.02.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5457801PMC
February 2016

Ligand Discovery for the Alanine-Serine-Cysteine Transporter (ASCT2, SLC1A5) from Homology Modeling and Virtual Screening.

PLoS Comput Biol 2015 Oct 7;11(10):e1004477. Epub 2015 Oct 7.

Department of Pharmacology and Systems Therapeutics, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.

The Alanine-Serine-Cysteine transporter ASCT2 (SLC1A5) is a membrane protein that transports neutral amino acids into cells in exchange for outward movement of intracellular amino acids. ASCT2 is highly expressed in peripheral tissues such as the lung and intestines where it contributes to the homeostasis of intracellular concentrations of neutral amino acids. ASCT2 also plays an important role in the development of a variety of cancers such as melanoma by transporting amino acid nutrients such as glutamine into the proliferating tumors. Therefore, ASCT2 is a key drug target with potentially great pharmacological importance. Here, we identify seven ASCT2 ligands by computational modeling and experimental testing. In particular, we construct homology models based on crystallographic structures of the aspartate transporter GltPh in two different conformations. Optimization of the models' binding sites for protein-ligand complementarity reveals new putative pockets that can be targeted via structure-based drug design. Virtual screening of drugs, metabolites, fragments-like, and lead-like molecules from the ZINC database, followed by experimental testing of 14 top hits with functional measurements using electrophysiological methods reveals seven ligands, including five activators and two inhibitors. For example, aminooxetane-3-carboxylate is a more efficient activator than any other known ASCT2 natural or unnatural substrate. Furthermore, two of the hits inhibited ASCT2 mediated glutamine uptake and proliferation of a melanoma cancer cell line. Our results improve our understanding of how substrate specificity is determined in amino acid transporters, as well as provide novel scaffolds for developing chemical tools targeting ASCT2, an emerging therapeutic target for cancer and neurological disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pcbi.1004477DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4596572PMC
October 2015

Structure-Based Identification of Inhibitors for the SLC13 Family of Na(+)/Dicarboxylate Cotransporters.

Biochemistry 2015 Aug 30;54(31):4900-8. Epub 2015 Jul 30.

†Department of Pharmacology and Systems Therapeutics, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, New York 10029, United States.

In mammals, citric acid cycle intermediates play a key role in regulating various metabolic processes, such as fatty acid synthesis and glycolysis. Members of the sodium-dependent SLC13 transporter family mediate the transport of di- and tricarboxylates into cells. SLC13 family members have been implicated in lifespan extension and resistance to high-fat diets; thus, they are emerging drug targets for aging and metabolic disorders. We previously characterized key structural determinants of substrate and cation binding for the human NaDC3/SLC13A3 transporter using a homology model. Here, we combine computational modeling and virtual screening with functional and biochemical testing, to identify nine previously unknown inhibitors for multiple members of the SLC13 family from human and mouse. Our results reveal previously unknown substrate selectivity determinants for the SLC13 family, including key residues that mediate ligand binding and transport, as well as promiscuous and specific SLC13 small molecule ligands. The newly discovered ligands can serve as chemical tools for further characterization of the SLC13 family or as lead molecules for the future development of potent inhibitors for the treatment of metabolic diseases and aging. Our results improve our understanding of the structural components that are important for substrate specificity in this physiologically important family as well as in other structurally related transport systems.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.biochem.5b00388DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4781755PMC
August 2015

Pro-apoptotic meiogynin A derivatives that target Bcl-xL and Mcl-1.

Bioorg Med Chem Lett 2014 Nov 6;24(21):5086-8. Epub 2014 Sep 6.

Institut de Chimie des Substances Naturelles, CNRS UPR 2301, 1 avenue de la terrasse, 91198 Gif-sur-Yvette Cedex, France.

The biological evaluation of a natural sesquiterpene dimer meiogynin A 1, is described as well as that of five non-natural analogues. Although active on a micromolar range on the inhibition of Bcl-xL/Bak and Mcl-1/Bid interaction, meiogynin A 1 is not cytotoxic on three cell lines that overexpress Bcl-xL and Mcl-1. Contrarily, one of its analogues 6 with an inverted configuration on the side chain and an aromatic moiety replacing the cyclohexane ring was active on both target proteins, cytotoxic on a micromolar range and was found to induce apoptosis through a classical pathway.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bmcl.2014.09.004DOI Listing
November 2014

Determinants of substrate and cation transport in the human Na+/dicarboxylate cotransporter NaDC3.

J Biol Chem 2014 Jun 7;289(24):16998-7008. Epub 2014 May 7.

the Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92130-0718

Metabolic intermediates, such as succinate and citrate, regulate important processes ranging from energy metabolism to fatty acid synthesis. Cytosolic concentrations of these metabolites are controlled, in part, by members of the SLC13 gene family. The molecular mechanism underlying Na(+)-coupled di- and tricarboxylate transport by this family is understood poorly. The human Na(+)/dicarboxylate cotransporter NaDC3 (SLC13A3) is found in various tissues, including the kidney, liver, and brain. In addition to citric acid cycle intermediates such as α-ketoglutarate and succinate, NaDC3 transports other compounds into cells, including N-acetyl aspartate, mercaptosuccinate, and glutathione, in keeping with its dual roles in cell nutrition and detoxification. In this study, we construct a homology structural model of NaDC3 on the basis of the structure of the Vibrio cholerae homolog vcINDY. Our computations are followed by experimental testing of the predicted NaDC3 structure and mode of interaction with various substrates. The results of this study show that the substrate and cation binding domains of NaDC3 are composed of residues in the opposing hairpin loops and unwound portions of adjacent helices. Furthermore, these results provide a possible explanation for the differential substrate specificity among dicarboxylate transporters that underpin their diverse biological roles in metabolism and detoxification. The structural model of NaDC3 provides a framework for understanding substrate selectivity and the Na(+)-coupled anion transport mechanism by the human SLC13 family and other key solute carrier transporters.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M114.554790DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4059142PMC
June 2014

Virtual screening of the SAMPL4 blinded HIV integrase inhibitors dataset.

J Comput Aided Mol Des 2014 Apr 24;28(4):455-62. Epub 2014 Jan 24.

Institut de Chimie des Substances Naturelles, CNRS UPR 2301, Centre de Recherche de Gif-sur-Yvette, Labex LERMIT, 1 Avenue de la Terrasse, 91198, Gif-sur-Yvette, France.

Several combinations of docking software and scoring functions were evaluated for their ability to predict the binding of a dataset of potential HIV integrase inhibitors. We found that different docking software were appropriate for each one of the three binding sites considered (LEDGF, Y3 and fragment sites), and the most suitable two docking protocols, involving Glide SP and Gold ChemScore, were selected using a training set of compounds identified from the structural data available. These protocols could successfully predict respectively 20.0 and 23.6 % of the HIV integrase binders, all of them being present in the LEDGF site. When a different analysis of the results was carried out by removing all alternate isomers of binders from the set, our predictions were dramatically improved, with an overall ROC AUC of 0.73 and enrichment factor at 10 % of 2.89 for the prediction obtained using Gold ChemScore. This study highlighted the ability of the selected docking protocols to correctly position in most cases the ortho-alkoxy-carboxylate core functional group of the ligands in the corresponding binding site, but also their difficulties to correctly rank the docking poses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10822-014-9707-5DOI Listing
April 2014

Identification of cinnamic acid derivatives as novel antagonists of the prokaryotic proton-gated ion channel GLIC.

J Med Chem 2013 Jun 4;56(11):4619-30. Epub 2013 Jun 4.

Institut Pasteur, Unité Récepteurs-Canaux, Paris, France.

Pentameric ligand gated ion channels (pLGICs) mediate signal transduction. The binding of an extracellular ligand is coupled to the transmembrane channel opening. So far, all known agonists bind at the interface between subunits in a topologically conserved "orthosteric site" whose amino acid composition defines the pharmacological specificity of pLGIC subtypes. A striking exception is the bacterial proton-activated GLIC protein, exhibiting an uncommon orthosteric binding site in terms of sequence and local architecture. Among a library of Gloeobacter violaceus metabolites, we identified a series of cinnamic acid derivatives, which antagonize the GLIC proton-elicited response. Structure-activity analysis shows a key contribution of the carboxylate moiety to GLIC inhibition. Molecular docking coupled to site-directed mutagenesis support that the binding pocket is located below the classical orthosteric site. These antagonists provide new tools to modulate conformation of GLIC, currently used as a prototypic pLGIC, and opens new avenues to study the signal transduction mechanism.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/jm400374qDOI Listing
June 2013

Discrimination of agonists versus antagonists of nicotinic ligands based on docking onto AChBP structures.

J Mol Graph Model 2011 Sep 29;30:100-9. Epub 2011 Jun 29.

Laboratoire de Conception et Application de Molécules Bioactives, UMR 7199 CNRS-Université de Strasbourg, 74 Route du Rhin-BP 60024, 67401 Illkirch Cedex, France.

Numerous high-resolution crystallographic structures of the acetylcholine binding protein (AChBP), a molluscan cholinergic protein, homologous to the extracellular domain of nicotinic acetylcholine receptors, are available. This offers opportunities to model the interaction between various ligands and the acetylcholine binding site. Herein we present a study of the interplay between ligand binding and motions of the C-loop capping the binding site. Nicotinic agonists and antagonists were docked on AChBP X-ray structures. It is shown that the studied agonists and antagonists can be discriminated according to their higher affinities for structures respectively obtained in the presence of agonists or antagonists, highlighting the fact that AChBP structures retain a pharmacological footprint of the compound used in crystallography experiments. A detailed analysis of the binding site cavities suggests that this property is mainly related to the shape of the cavities.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jmgm.2011.06.008DOI Listing
September 2011