I03-Macromolecular Crystallography
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Maria Giulia
Nizi
,
Mirko M.
Maksimainen
,
Sudarshan
Murthy
,
Serena
Massari
,
Juho
Alaviuhkola
,
Barbara E.
Lippok
,
Sven T.
Sowa
,
Albert
Galera-Prat
,
Renata
Prunskaite-Hyyryläinen
,
Bernhard
Lüscher
,
Patricia
Korn
,
Lari
Lehtio
,
Oriana
Tabarrini
Diamond Proposal Number(s):
[19951]
Abstract: While human poly-ADP-ribose chain generating poly-ARTs, PARP1 and 2 and TNKS1 and 2, have been widely characterized, less is known on the pathophysiological roles of the mono-ADP-ribosylating mono-ARTs, partly due to the lack of selective inhibitors. In this context, we have focused on the development of inhibitors for the mono-ART PARP10, whose overexpression is known to induce cell death. Starting from OUL35 (1) and its 4-(benzyloxy)benzamidic derivative (2) we herein report the design and synthesis of new analogues from which the cyclobutyl derivative 3c rescued cells most efficiently from PARP10 induced apoptosis. Most importantly, we also identified 2,3-dihydrophthalazine-1,4-dione as a new suitable nicotinamide mimicking PARP10 inhibitor scaffold. When it was functionalized with cycloalkyl (8a-c), o-fluorophenyl (8h), and thiophene (8l) rings, IC50 values in the 130–160 nM range were obtained, making them the most potent PARP10 inhibitors reported to date. These compounds also inhibited PARP15 with low micromolar IC50s, but none of the other tested poly- and mono-ARTs, thus emerging as dual mono-ART inhibitors. Compounds 8a, 8h and 8l were also able to enter cells and rescue cells from apoptosis. Our work sheds more light on inhibitor development against mono-ARTs and identifies chemical probes to study the cellular roles of PARP10 and PARP15.
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Jul 2022
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Amalyn
Nain-Perez
,
Anders
Foller Füchtbauer
,
Liliana
Håversen
,
Aleksei
Lulla
,
Chunxia
Gao
,
Josipa
Matic
,
Leticia
Monjas
,
Alexandra
Rodríguez
,
Paul
Brear
,
Woonghee
Kim
,
Marko
Hyvonen
,
Jan
Borén
,
Adil
Mardinoglu
,
Mathias
Uhlen
,
Morten
Grøtli
Diamond Proposal Number(s):
[25402]
Abstract: Liver pyruvate kinase (PKL) is a major regulator of metabolic flux and ATP production during liver cell glycolysis and is considered a potential drug target for the treatment of non-alcoholic fatty liver disease (NAFLD). In this study, we report the first ADP-competitive PKL inhibitors and identify several starting points for the further optimization of these inhibitors. Modeling and structural biology guided the optimization of a PKL-specific anthraquinone-based compound. A structure–activity relationship study of 47 novel synthetic derivatives revealed PKL inhibitors with half-maximal inhibitory concentration (IC50) values in the 200 nM range. Despite the difficulty involved in studying a binding site as exposed as the ADP site, these derivatives feature expanded structural diversity and chemical spaces that may be used to improve their inhibitory activities against PKL. The obtained results expand the knowledge of the structural requirements for interactions with the ADP-binding site of PKL.
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Mar 2022
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Martyn
Frederickson
,
Irwin R.
Selvam
,
Dimitrios
Evangelopoulos
,
Kirsty J.
Mclean
,
Mona M.
Katariya
,
Richard B.
Tunnicliffe
,
Bethany
Campbell
,
Madeline E.
Kavanagh
,
Sitthivut
Charoensutthivarakul
,
Richard T.
Blankley
,
Colin W.
Levy
,
Luiz Pedro S.
De Carvalho
,
David
Leys
,
Andrew W.
Munro
,
Anthony G.
Coyne
,
Chris
Abell
Diamond Proposal Number(s):
[17773, 24447]
Abstract: There is a pressing need for new drugs against tuberculosis (TB) to combat the growing resistance to current antituberculars. Herein a novel strategy is described for hit generation against promising TB targets involving X-ray crystallographic screening in combination with phenotypic screening. This combined approach (XP Screen) affords both a validation of target engagement as well as determination of in cellulo activity. The utility of this method is illustrated by way of an XP Screen against CYP121A1, a cytochrome P450 enzyme from Mycobacterium tuberculosis (Mtb) championed as a validated drug discovery target. A focused screening set was synthesized and tested by such means, with several members of the set showing promising activity against Mtb strain H37Rv. One compound was observed as an X-ray hit against CYP121A1 and showed improved activity against Mtb strain H37Rv under multiple assay conditions (pan-assay activity). Data obtained during X-ray crystallographic screening were utilized in a structure-based campaign to design a limited number of analogues (less than twenty), many of which also showed pan-assay activity against Mtb strain H37Rv. These included the benzo[b][1,4]oxazine derivative (MIC90 6.25 μM), a novel hit compound suitable as a starting point for a more involved lead-to-clinical candidate medicinal chemistry campaign.
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Jan 2022
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I04-Macromolecular Crystallography
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Alice
Legru
,
Federica
Verdirosa
,
Jean-François
Hernandez
,
Giusi
Tassone
,
Filomena
Sannio
,
Manuela
Benvenuti
,
Pierre-Alexis
Conde
,
Guillaume
Bossis
,
Caitlyn A.
Thomas
,
Michael W.
Crowder
,
Melissa
Dillenberger
,
Katja
Becker
,
Cecilia
Pozzi
,
Stefano
Mangani
,
Jean-Denis
Docquier
,
Laurent
Gavara
Diamond Proposal Number(s):
[21741]
Abstract: Metallo-β-lactamases (MBLs) are important contributors of Gram-negative bacteria resistance to β-lactam antibiotics. MBLs are highly worrying because of their carbapenemase activity, their rapid spread in major human opportunistic pathogens while no clinically useful inhibitor is available yet. In this context, we are exploring the potential of compounds based on the 1,2,4-triazole-3-thione scaffold as an original ligand of the di-zinc active sites of MBLs, and diversely substituted at its positions 4 and 5. Here, we present a new series of compounds substituted at the 4-position by a thioether-containing alkyl chain with a carboxylic and/or an aryl group at its extremity. Several compounds showed broad-spectrum inhibition with Ki values in the μM to sub-μM range against VIM-type enzymes, NDM-1 and IMP-1. The presence of the sulfur and of the aryl group was important for the inhibitory activity and the binding mode of a few compounds in VIM-2 was revealed by X-ray crystallography. Importantly, in vitro antibacterial susceptibility assays showed that several inhibitors were able to potentiate the activity of meropenem on Klebsiella pneumoniae clinical isolates producing VIM-1 or VIM-4, with a potentiation effect of up to 16-fold. Finally, a selected compound was found to only moderately inhibit the di-zinc human glyoxalase II, and several showed no or only moderate toxicity toward several human cells, thus favourably completing a promising behaviour.
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Oct 2021
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I04-Macromolecular Crystallography
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Mujtaba
Hassan
,
Sjors
Van Klaveren
,
Maria
Håkansson
,
Carl
Diehl
,
Rebeka
Kovačič
,
Floriane
Baussière
,
Anders
Sundin
,
Jaka
Dernovšek
,
Björn
Walse
,
Fredrik
Zetterberg
,
Hakon
Leffler
,
Marko
Anderluh
,
Tihomir
Tomašič
,
Žiga
Jakopin
,
Ulf J.
Nilsson
Diamond Proposal Number(s):
[23282]
Abstract: We have obtained the X-ray crystal structure of the galectin-8 N-terminal domain (galectin-8N) with a previously reported quinoline–galactoside ligand at a resolution of 1.6 Å. Based on this X-ray structure, a collection of galactosides derivatised at O3 with triazole, benzimidazole, benzothiazole, and benzoxazole moieties were designed and synthesised. This led to the discovery of a 3-O-(N-methylbenzimidazolylmethyl)–galactoside with a Kd of 1.8 μM for galectin-8N, the most potent selective synthetic galectin-8N ligand to date. Molecular dynamics simulations showed that benzimidazole–galactoside derivatives bind the non-conserved amino acid Gln47, accounting for the higher selectivity for galectin-8N. Galectin-8 is a carbohydrate-binding protein that plays a key role in pathological lymphangiogenesis, modulation of the immune system, and autophagy. Thus, the benzimidazole-derivatised galactosides represent promising compounds for studies of the pathological implications of galectin-8, as well as a starting point for the development of anti-tumour and anti-inflammatory therapeutics targeting galectin-8.
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Jun 2021
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I04-Macromolecular Crystallography
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Martina
Durcik
,
Ákos
Nyerges
,
Žiga
Skok
,
Darja Gramec
Skledar
,
Jurij
Trontelj
,
Nace
Zidar
,
Janez
Ilaš
,
Anamarija
Zega
,
Cristina D.
Cruz
,
Päivi
Tammela
,
Martin
Welin
,
Yengo R.
Kimbung
,
Dorota
Focht
,
Ondřej
Benek
,
Tamás
Révész
,
Gábor
Draskovits
,
Petra Éva
Szili
,
Lejla
Daruka
,
Csaba
Pál
,
Danijel
Kikelj
,
Lucija Peterlin
Mašič
,
Tihomir
Tomašič
Diamond Proposal Number(s):
[20028]
Abstract: The rise in multidrug-resistant bacteria defines the need for identification of new antibacterial agents that are less prone to resistance acquisition. Compounds that simultaneously inhibit multiple bacterial targets are more likely to suppress the evolution of target-based resistance than monotargeting compounds. The structurally similar ATP binding sites of DNA gyrase and topoisomerase Ⅳ offer an opportunity to accomplish this goal. Here we present the design and structure-activity relationship analysis of balanced, low nanomolar inhibitors of bacterial DNA gyrase and topoisomerase IV that show potent antibacterial activities against the ESKAPE pathogens. For inhibitor 31c, a crystal structure in complex with Staphylococcus aureus DNA gyrase B was obtained that confirms the mode of action of these compounds. The best inhibitor, 31h, does not show any in vitro cytotoxicity and has excellent potency against Gram-positive (MICs: range, 0.0078–0.0625 μg/mL) and Gram-negative pathogens (MICs: range, 1–2 μg/mL). Furthermore, 31h inhibits GyrB mutants that can develop resistance to other drugs. Based on these data, we expect that structural derivatives of 31h will represent a step toward clinically efficacious multitargeting antimicrobials that are not impacted by existing antimicrobial resistance.
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Mar 2021
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Anka
Lucic
,
Philip
Hinchliffe
,
Tika R.
Malla
,
Catherine L.
Tooke
,
Jurgen
Brem
,
Karina
Calvopina
,
Christopher T.
Lohans
,
Patrick
Rabe
,
Michael A.
Mcdonough
,
Timothy
Armistead
,
Allen M.
Orville
,
James
Spencer
,
Christopher J.
Schofield
Diamond Proposal Number(s):
[17212, 23269, 18069]
Abstract: Penems have demonstrated potential as antibacterials and β-lactamase inhibitors; however, their clinical use has been limited, especially in comparison with the structurally related carbapenems. Faropenem is an orally active antibiotic with a C2 tetrahydrofuran (THF) ring, which is resistant to hydrolysis by some β-lactamases. We report studies on the reactions of faropenem with carbapenem-hydrolysing β-lactamases, focusing on the class A serine β-lactamase KPC-2 and the metallo β-lactamases (MBLs) VIM-2 (a subclass B1 MBL) and L1 (a B3 MBL). Kinetic studies show that faropenem is a substrate for all three β-lactamases, though it is less efficiently hydrolysed by KPC-2. Crystallographic analyses on faropenem-derived complexes reveal the opening of the β-lactam ring with formation of an imine with KPC-2, VIM-2, and L1. In the cases of the KPC-2 and VIM-2 structures, the THF ring is opened to give an alkene, but with L1 the THF ring remains intact. Solution state studies, employing NMR, were performed on L1, KPC-2, VIM-2, VIM-1, NDM-1, OXA-23, OXA-10, and OXA-48. The solution results reveal, in all cases, formation of imine products in which the THF ring is opened; formation of a THF ring-closed imine product was only observed with VIM-1 and VIM-2. An enamine product with a closed THF ring was also observed in all cases, at varying levels. Combined with previous reports, the results exemplify the potential for different outcomes in the reactions of penems with MBLs and SBLs and imply further structure-activity relationship studies are worthwhile to optimise the interactions of penems with β-lactamases. They also exemplify how crystal structures of β-lactamase substrate/inhibitor complexes do not always reflect reaction outcomes in solution.
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Feb 2021
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I03-Macromolecular Crystallography
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Sandra
Röhm
,
Martin
Schroeder
,
Jessica E.
Dwyer
,
Caroline S.
Widdowson
,
Apirat
Chaikuad
,
Benedict-Tilman
Berger
,
Andreas C.
Joerger
,
Andreas
Krämer
,
Jule
Harbig
,
Daniel
Dauch
,
Mark
Kudolo
,
Stefan
Laufer
,
Mark C.
Bagley
,
Stefan
Knapp
Diamond Proposal Number(s):
[10619]
Abstract: The p38 MAPK cascade is a key signaling pathway linked to a multitude of physiological functions and of central importance in inflammatory and autoimmune diseases. Although studied extensively, little is known about how conformation-specific inhibitors alter signaling outcomes. Here, we have explored the highly dynamic back pocket of p38 MAPK with allosteric urea fragments. However, screening against known off-targets showed that these fragments maintained the selectivity issues of their parent compound BIRB-796, while combination with the hinge-binding motif of VPC-00628 greatly enhanced inhibitor selectivity. Further efforts focused therefore on the exploration of the αC-out pocket of p38 MAPK, yielding compound 137 as a highly selective type-II inhibitor. Even though 137 is structurally related to a recent p38 type-II chemical probe, SR-318, the data presented here provide valuable insights into back-pocket interactions that are not addressed in SR-318 and it provides an alternative chemical tool with good cellular activity targeting also the p38 back pocket.
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Dec 2020
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I04-1-Macromolecular Crystallography (fixed wavelength)
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Lucía
Serran Aguilera
,
Elena
Mariotto
,
Gianluca
Rubbini
,
Francisco Fermín
Castro Navas
,
Carmen
Marco
,
María Paz
Carrasco-Jiménez
,
Marco
Ballarotto
,
Antonio
Macchiarulo
,
Ramon
Hurtado-Guerrero
,
Giampietro
Viola
,
Luisa Carlota
Lopez-Cara
Diamond Proposal Number(s):
[8035]
Abstract: Seeking for new anticancer drugs with strong antiproliferative activity and simple molecular structure, we designed a novel series of compounds based on our previous reported pharmacophore model composed of five moieties. Antiproliferative assays on four tumoral cell lines and evaluation of Human Choline Kinase CKα1 enzymatic activity was performed for these compounds. Among tested molecules, those ones with biphenyl spacer showed betters enzymatic and antiproliferative activities (n-v). Docking and crystallization studies validate the hypothesis and confirm the results. The most active compound (t) induces a significant arrest of the cell cycle in G0/G1 phase that ultimately lead to apoptosis, following the mitochondrial pathway, as demonstrated for other choline kinase inhibitors. However additional assays reveal that the inhibition of choline uptake could also be involved in the antiproliferative outcome of this class of compounds.
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Dec 2020
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I04-Macromolecular Crystallography
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Sonia
Martínez-González
,
Ana
Belén García
,
M. Isabel
Albarrán
,
Antonio
Cebriá
,
Adrián
Amezquita-Alves
,
Francisco Javier
García-Campos
,
Jaime
Martínez-Gago
,
Jorge
Martínez-Torrecuadrada
,
Ines
Munoz
,
Carmen
Blanco-Aparicio
,
Joaquín
Pastor
Diamond Proposal Number(s):
[16252]
Abstract: CDK8 is a cyclin-dependent kinase that forms part of the mediator complex, and modulates the transcriptional output from distinct transcription factors involved in oncogenic control. Overexpression of CDK8 has been observed in various cancers, representing a potential target for developing novel CDK8 inhibitors in cancer therapeutics. In the course of our investigations to discover new CDK8 inhibitors, we designed and synthesized tricyclic pyrido[2,3-b][1,5]benzoxazepin-5(6H)-one derivatives, by introduction of chemical complexity in the multi-kinase inhibitor Sorafenib taking into account the flexibility of the P-loop motif of CDK8 protein observed after analysis of structural information of co-crystallized CDK8 inhibitors. In vitro evaluation of the inhibitory activity of the prepared compounds against CDK8 led us to identify compound 2 as the most potent inhibitor of the series (IC50 = 8.25 nM). Co-crystal studies and the remarkable selectivity profile of compound 2 are presented. Compound 2 showed moderate reduction of phosphorylation of CDK8 substrate STAT1 in cells, in line with other reported Type II CDK8 inhibitors. We propose herein an alternative to find a potential therapeutic use for this chemical series.
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Sep 2020
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