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12 items found (0 items not approved), displaying 1 to 10.[First/Prev] 1, 2 [Next/Last]

Instruments / Technical Area	Publication Details	Published
I03-Macromolecular Crystallography I04-Macromolecular Crystallography	Promiscuous recognition of MR1 drives self-reactive mucosal-associated invariant T cell responses Andrew Chancellor , Robert A. Simmons , Rahul C. Khanolkar , Vladimir Nosi , Aisha Beshirova , Giuliano Berloffa , Rodrigo Colombo , Vijaykumar Karuppiah , Johanne M. Pentier , Vanessa Tubb , Hemza Ghadbane , Richard J. Suckling , Keith Page , Rory M. Crean , Alessandro Vacchini , Corinne De Gregorio , Verena Schaefer , Daniel Constantin , Thomas Gligoris , Angharad Lloyd , Miriam Hock , Velupillai Srikannathasan , Ross A. Robinson , Gurdyal S. Besra , Marc W. Van Der Kamp , Lucia Mori , Raffaele Calogero , David K. Cole , Gennaro De Libero , Marco Lepore Journal Of Experimental Medicine 220 DOI: 10.1084/jem.20221939 Diamond Proposal Number(s): [22870, 28224] Show Abstract ▼ Abstract: Mucosal-associated invariant T (MAIT) cells use canonical semi-invariant T cell receptors (TCR) to recognize microbial riboflavin precursors displayed by the antigen-presenting molecule MR1. The extent of MAIT TCR crossreactivity toward physiological, microbially unrelated antigens remains underexplored. We describe MAIT TCRs endowed with MR1-dependent reactivity to tumor and healthy cells in the absence of microbial metabolites. MAIT cells bearing TCRs crossreactive toward self are rare but commonly found within healthy donors and display T-helper-like functions in vitro. Experiments with MR1-tetramers loaded with distinct ligands revealed significant crossreactivity among MAIT TCRs both ex vivo and upon in vitro expansion. A canonical MAIT TCR was selected on the basis of extremely promiscuous MR1 recognition. Structural and molecular dynamic analyses associated promiscuity to unique TCRβ-chain features that were enriched within self-reactive MAIT cells of healthy individuals. Thus, self-reactive recognition of MR1 represents a functionally relevant indication of MAIT TCR crossreactivity, suggesting a potentially broader role of MAIT cells in immune homeostasis and diseases, beyond microbial immunosurveillance.	Sep 2023
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography	Structure‐guided stabilization of pathogen‐derived peptide‐HLA‐E complexes using non‐natural amino acids conserves native TCR recognition Claire Barber , Victoria Arena De Souza , Rachel L. Paterson , Magdalena Martin-Urdiroz , Nitha Charles Mulakkal , Velupillai Srikannathasan , Mary Connolly , Gwilym Phillips , Tein Foong-Leong , Robert Pengelly , Vijaykumar Karuppiah , Tressan Grant , Marcin Dembek , Anil Verma , Dawn Gibbs-Howe , Thomas H. Blicher , Andrew Knox , Ross A. Robinson , David K. Cole , Sarah Leonard European Journal Of Immunology DOI: 10.1002/eji.202149745 Open Access Show Abstract ▼ Abstract: The non-polymorphic class Ib molecule, human leukocyte antigen (HLA)-E, primarily presents peptides from HLA class Ia leader peptides, providing an inhibitory signal to NK cells via CD94/NKG2 interactions. Although peptides of pathogenic origin can also be presented by HLA-E to T cells, the molecular basis underpinning their role in antigen surveillance is largely unknown. Here, we solved a co-complex crystal structure of a T cell receptor (TCR) with an HLA-E presented peptide (pHLA-E) from bacterial (Mycobacterium tuberculosis) origin, and the first TCR-pHLA-E complex with a non-canonically presented peptide from viral (human immuno-deficiency virus; HIV) origin. The structures provided a molecular foundation to develop a novel method to introduce cysteine traps using non-natural amino acid chemistry that stabilized pHLA-E complexes whilst maintaining native interface contacts between the TCRs and different pHLA-E complexes. These pHLA-E monomers could be used to isolate pHLA-E specific T cells, with obvious utility for studying pHLA-E restricted T cells, and for the identification of putative therapeutic TCRs.	Feb 2022
I04-1-Macromolecular Crystallography (fixed wavelength)	Specificity of bispecific T cell receptors and antibodies targeting peptide-HLA Christopher J. Holland , Rory M. Crean , Johanne M. Pentier , Ben De Wet , Angharad Lloyd , Velupillai Srikannathasan , Nikolai Lissin , Katy A. Lloyd , Thomas H. Blicher , Paul J. Conroy , Miriam Hock , Robert J. Pengelly , Thomas E. Spinner , Brian Cameron , Elizabeth A. Potter , Anitha Jeyanthan , Peter E. Molloy , Malkit Sami , Milos Aleksic , Nathaniel Liddy , Ross A. Robinson , Stephen Harper , Marco Lepore , Chris R. Pudney , Marc W. Van Der Kamp , Pierre J. Rizkallah , Bent K. Jakobsen , Annelise Vuidepot , David K. Cole Journal Of Clinical Investigation 130, 2673 - 2688 DOI: 10.1172/JCI130562 Diamond Proposal Number(s): [17077, 14843] Show Abstract ▼ Abstract: Tumor-associated peptide–human leukocyte antigen complexes (pHLAs) represent the largest pool of cell surface–expressed cancer-specific epitopes, making them attractive targets for cancer therapies. Soluble bispecific molecules that incorporate an anti-CD3 effector function are being developed to redirect T cells against these targets using 2 different approaches. The first achieves pHLA recognition via affinity-enhanced versions of natural TCRs (e.g., immune-mobilizing monoclonal T cell receptors against cancer [ImmTAC] molecules), whereas the second harnesses an antibody-based format (TCR-mimic antibodies). For both classes of reagent, target specificity is vital, considering the vast universe of potential pHLA molecules that can be presented on healthy cells. Here, we made use of structural, biochemical, and computational approaches to investigate the molecular rules underpinning the reactivity patterns of pHLA-targeting bispecifics. We demonstrate that affinity-enhanced TCRs engage pHLA using a comparatively broad and balanced energetic footprint, with interactions distributed over several HLA and peptide side chains. As ImmTAC molecules, these TCRs also retained a greater degree of pHLA selectivity, with less off-target activity in cellular assays. Conversely, TCR-mimic antibodies tended to exhibit binding modes focused more toward hot spots on the HLA surface and exhibited a greater degree of crossreactivity. Our findings extend our understanding of the basic principles that underpin pHLA selectivity and exemplify a number of molecular approaches that can be used to probe the specificity of pHLA-targeting molecules, aiding the development of future reagents.	Apr 2020
I02-Macromolecular Crystallography I03-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	A chemical probe for tudor domain protein spindlin1 to investigate chromatin function Vincent Fagan , Catrine Johansson , Carina Gileadi , Octovia Monteiro , James Edward Dunford , Reshma Nibhani , Martin Philpott , Jessica Malzahn , Graham Wells , Ruth Farham , Adam Cribbs , Nadia Halidi , Fengling Li , Irene Chau , Holger Greschik , Srikannathasan Velupillai , Abdellah Allali-Hassani , James M. Bennett , Thomas Christott , Charline Giroud , Andrew M. Lewis , Kilian V. M. Huber , Nick Athanasou , Chas Bountra , Manfred Jung , Roland Schüle , Masoud Vedadi , Cheryl H. Arrowsmith , Yan Xiong , Jian Jin , Oleg Fedorov , Gillian Farnie , Paul E. Brennan , Udo C. T. Oppermann Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.9b00562 Diamond Proposal Number(s): [10619, 15433] Show Abstract ▼ Abstract: Modifications of histone tails, including lysine/arginine methylation, provide the basis of a 'chromatin or histone code'. Proteins that con-tain 'reader' domains can bind to these modifications and form specific effector complexes, which ultimately mediate chromatin function. The spindlin1 (SPIN1) protein contains three Tudor methyl-lysine/arginine reader domains and was identified as a putative onco-gene and transcriptional co-activator. Here we report a SPIN1 chemi-cal probe inhibitor with low nanomolar in vitro activity, exquisite selectivity on a panel of methyl reader and writer proteins, and with submicromolar cellular activity. X-ray crystallography showed that this Tudor domain chemical probe simultaneously engages Tudor domains 1 and 2 via a bidentate binding mode. Small molecule inhibition and siRNA knockdown of SPIN1, as well as chemoproteomic studies, iden-tified genes which are transcriptionally regulated by SPIN1 in squa-mous cell carcinoma and suggest that SPIN1 may have a roll in cancer related inflammation and/or cancer metastasis.	Sep 2019
I02-Macromolecular Crystallography I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	C8-substituted pyrido[3,4-d]pyrimidin-4(3H)-ones: Studies towards the identification of potent, cell penetrant Jumonji C domain containing histone lysine demethylase 4 subfamily (KDM4) inhibitors, compound profiling in cell-based target engagement assays Yann-Vai Le Bihan , Rachel M. Lanigan , Butrus Atrash , Mark G. Mclaughlin , Srikannathasan Velupillai , Andrew G. Malcolm , Katherine S. England , Gian Filippo Ruda , N. Yi Mok , Anthony Tumber , Kathy Tomlin , Harry Saville , Erald Shehu , Craig Mcandrew , Leanne Carmichael , James M. Bennett , Fiona Jeganathan , Paul Eve , Adam Donovan , Angela Hayes , Francesca Wood , Florence I. Raynaud , Oleg Fedorov , Paul Brennan , Rosemary Burke , Rob Van Montfort , Olivia W. Rossanese , Julian Blagg , Vassilios Bavetsias European Journal Of Medicinal Chemistry DOI: 10.1016/j.ejmech.2019.05.041 Diamond Proposal Number(s): [20145] Open Access Show Abstract ▼ Abstract: Residues in the histone substrate binding sites that differ between the KDM4 and KDM5 subfamilies were identified. Subsequently, a C8-substituted pyrido[3,4-d]pyrimidin-4(3H)-one series was designed to rationally exploit these residue differences between the histone substrate binding sites in order to improve affinity for the KDM4-subfamily over KDM5-subfamily enzymes. In particular, residues E169 and V313 (KDM4A numbering) were targeted. Additionally, the conformational restriction of the flexible pyridopyrimidinone C8-substituent was investigated. These approaches yielded potent and cell-penetrant dual KDM4/5-subfamily inhibitors including 19a (KDM4A and KDM5B Ki = 0.004 and 0.007 μM, respectively). Compound cellular profiling in two orthogonal target engagement assays revealed a significant reduction from biochemical to cell-based activity across multiple analogues; this decrease was shown to be consistent with 2OG competition, and suggest that sub-nanomolar biochemical potency will be required with C8-substituted pyrido[3,4-d]pyrimidin-4(3H)-one compounds to achieve sub-micromolar target inhibition in cells.	May 2019
I02-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	Thiophene antibacterials that allosterically stabilize DNA-cleavage complexes with DNA gyrase Pan F. Chan , Thomas Germe , Benjamin Bax , Jianzhong Huang , Reema K. Thalji , Eric Bacqué , Anna Checchia , Dongzhao Chen , Haifeng Cui , Xiao Ding , Karen Ingraham , Lynn Mccloskey , Kaushik Raha , Velupillai Srikannathasan , Anthony Maxwell , Robert A. Stavenger Proceedings Of The National Academy Of Sciences 114, E4492 - E4500 DOI: 10.1073/pnas.1700721114 Diamond Proposal Number(s): [1195] Open Access Show Abstract ▼ Abstract: A paucity of novel acting antibacterials is in development to treat the rising threat of antimicrobial resistance, particularly in Gram-negative hospital pathogens, which has led to renewed efforts in antibiotic drug discovery. Fluoroquinolones are broad-spectrum antibacterials that target DNA gyrase by stabilizing DNA-cleavage complexes, but their clinical utility has been compromised by resistance. We have identified a class of antibacterial thiophenes that target DNA gyrase with a unique mechanism of action and have activity against a range of bacterial pathogens, including strains resistant to fluoroquinolones. Although fluoroquinolones stabilize double-stranded DNA breaks, the antibacterial thiophenes stabilize gyrase-mediated DNA-cleavage complexes in either one DNA strand or both DNA strands. X-ray crystallography of DNA gyrase–DNA complexes shows the compounds binding to a protein pocket between the winged helix domain and topoisomerase-primase domain, remote from the DNA. Mutations of conserved residues around this pocket affect activity of the thiophene inhibitors, consistent with allosteric inhibition of DNA gyrase. This druggable pocket provides potentially complementary opportunities for targeting bacterial topoisomerases for antibiotic development.	May 2017
B21-High Throughput SAXS I02-Macromolecular Crystallography I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography	Structural analysis of human KDM5B guides histone demethylase inhibitor development Catrine Johansson , Velupillai Srikannathasan , Anthony Tumber , Aleksandra Szykowska , Edward S. Hookway , Radoslaw Nowak , Claire Strain-Damerell , Carina Gileadi , Martin Philpott , Nicola Burgess-Brown , Na Wu , Jolanta Kopec , Andrea Nuzzi , Holger Steuber , Ursula Egner , Volker Badock , Shonagh Munro , Nicholas B Lathangue , Sue Westaway , Jack Brown , Nick Athanasou , Rab Prinjha , Paul E Brennan , Udo Oppermann Nature Chemical Biology DOI: 10.1038/nchembio.2087 Diamond Proposal Number(s): [10619] Show Abstract ▼ Abstract: Members of the KDM5 (also known as JARID1) family are 2-oxoglutarate- and Fe2+-dependent oxygenases that act as histone H3K4 demethylases, thereby regulating cell proliferation and stem cell self-renewal and differentiation. Here we report crystal structures of the catalytic core of the human KDM5B enzyme in complex with three inhibitor chemotypes. These scaffolds exploit several aspects of the KDM5 active site, and their selectivity profiles reflect their hybrid features with respect to the KDM4 and KDM6 families. Whereas GSK-J1, a previously identified KDM6 inhibitor, showed about sevenfold less inhibitory activity toward KDM5B than toward KDM6 proteins, KDM5-C49 displayed 25–100-fold selectivity between KDM5B and KDM6B. The cell-permeable derivative KDM5-C70 had an antiproliferative effect in myeloma cells, leading to genome-wide elevation of H3K4me3 levels. The selective inhibitor GSK467 exploited unique binding modes, but it lacked cellular potency in the myeloma system. Taken together, these structural leads deliver multiple starting points for further rational and selective inhibitor design.	May 2016
I02-Macromolecular Crystallography I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography	8-substituted pyrido[3,4-d]pyrimidin-4(3H)-one derivatives as potent, cell permeable, KDM4 (JMJD2) and KDM5 (JARID1) histone lysine demethylase inhibitors Vassilios Bavetsias , Rachel M. Lanigan , Gian Filippo Ruda , Butrus Atrash , Mark G. Mclaughlin , Anthony Tumber , N. Yi Mok , Yann-Vaï Le Bihan , Sally Dempster , Katherine J. Boxall , Fiona Jeganathan , Stephanie B. Hatch , Pavel Savitsky , Velupillai Srikannathasan , Tobias Krojer , Katherine S. England , Jimmy Sejberg , Ching Thai , Adam Donovan , Akos Pal Journal Of Medicinal Chemistry 55 (1) DOI: 10.1021/acs.jmedchem.5b01635 PMID: 26741168 Open Access Show Abstract ▼ Abstract: We report the discovery of N-substituted 4-(pyridin-2-yl)thiazole-2-amine derivatives and their subsequent optimization, guided by structure-based design, to give 8-(1H-pyrazol-3-yl)pyrido[3,4-d]pyrimidin-4(3H)-ones, a series of potent JmjC histone N-methyl lysine demethylase (KDM) inhibitors which bind to Fe(II) in the active site. Substitution from C4 of the pyrazole moiety allows access to the histone peptide substrate binding site; incorporation of a conformationally constrained 4-phenylpiperidine linker gives derivatives such as 54j and 54k which demonstrate equipotent activity versus the KDM4 (JMJD2) and KDM5 (JARID1) subfamily demethylases, selectivity over representative exemplars of the KDM2, KDM3, and KDM6 subfamilies, cellular permeability in the Caco-2 assay, and, for 54k, inhibition of H3K9Me3 and H3K4Me3 demethylation in a cell-based assay.	Jan 2016
I02-Macromolecular Crystallography I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	Structural basis of DNA gyrase inhibition by antibacterial QPT-1, anticancer drug etoposide and moxifloxacin Pan F. Chan , Velupillai Srikannathasan , Jianzhong Huang , Haifeng Cui , Andrew P. Fosberry , Minghua Gu , Michael M. Hann , Martin Hibbs , Paul Homes , Karen Ingraham , Jason Pizzollo , Carol Shen , Anthony J. Shillings , Claus E. Spitzfaden , Robert Tanner , Andrew J. Theobald , Robert A. Stavenger , Benjamin D. Bax , Michael N. Gwynn Nature Communications 6 DOI: 10.1038/ncomms10048 PMID: 26640131 Open Access Show Abstract ▼ Abstract: New antibacterials are needed to tackle antibiotic-resistant bacteria. Type IIA topoisomerases (topo2As), the targets of fluoroquinolones, regulate DNA topology by creating transient double-strand DNA breaks. Here we report the first co-crystal structures of the antibacterial QPT-1 and the anticancer drug etoposide with Staphylococcus aureus DNA gyrase, showing binding at the same sites in the cleaved DNA as the fluoroquinolone moxifloxacin. Unlike moxifloxacin, QPT-1 and etoposide interact with conserved GyrB TOPRIM residues rationalizing why QPT-1 can overcome fluoroquinolone resistance. Our data show etoposide’s antibacterial activity is due to DNA gyrase inhibition and suggests other anticancer agents act similarly. Analysis of multiple DNA gyrase co-crystal structures, including asymmetric cleavage complexes, led to a ‘pair of swing-doors’ hypothesis in which the movement of one DNA segment regulates cleavage and religation of the second DNA duplex. This mechanism can explain QPT-1’s bacterial specificity. Structure-based strategies for developing topo2A antibacterials are suggested.	Dec 2015
I02-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography	Docking and linking of fragments to discover Jumonji histone demethylase inhibitors Magdalena Korczynska , Daniel D. Le , Noah Younger , Elisabet Gregori-Puigjané , Anthony Tumber , Tobias Krojer , Velupillai Srikannathasan , Carina Gileadi , Radoslaw Nowak , Eriko Iwasa , Samuel B. Pollock , Idelisse Ortiz Torres , Udo Oppermann , Brian K. Shoichet , Danica Galonić Fujimori Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.5b01527 PMID: 26699912 Show Abstract ▼ Abstract: Development of tool molecules that inhibit Jumonji demethylases allows for the investigation of cancer-associated transcription. While scaffolds such as 2,4-pyridinedicarboxylic acid (2,4-PDCA) are potent inhibitors, they exhibit limited selectivity. To discover new inhibitors for the KDM4 demethylases, enzymes overexpressed in several cancers, we docked a library of 600 000 fragments into the high-resolution structure of KDM4A. Among the most interesting chemotypes were the 5-aminosalicylates, which docked in two distinct but overlapping orientations. Docking poses informed the design of covalently linked fragment compounds, which were further derivatized. This combined approach improved affinity by ∼3 log-orders to yield compound 35 (Ki = 43 nM). Several hybrid inhibitors were selective for KDM4C over the related enzymes FIH, KDM2A, and KDM6B while lacking selectivity against the KDM3 and KDM5 subfamilies. Cocrystal structures corroborated the docking predictions. This study extends the use of structure-based docking from fragment discovery to fragment linking optimization, yielding novel KDM4 inhibitors.	Dec 2015

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