I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Iacovos N.
Michaelides
,
Gavin W.
Collie
,
Ulf
Börjesson
,
Christina
Vasalou
,
Omar
Alkhatib
,
Louise
Barlind
,
Tony
Cheung
,
Ian L.
Dale
,
Kevin J.
Embrey
,
Edward J.
Hennessy
,
Puneet
Khurana
,
Cheryl M.
Koh
,
Michelle L.
Lamb
,
Jianming
Liu
,
Thomas A.
Moss
,
Daniel J.
O’neill
,
Christopher
Phillips
,
Joseph
Shaw
,
Arjan
Snijder
,
R. Ian
Storer
,
Christopher J.
Stubbs
,
Fujin
Han
,
Chengzhi
Li
,
Jingchuan
Qiao
,
Dong-Qing
Sun
,
Jingwen
Wang
,
Peng
Wang
,
Wenzhen
Yang
Diamond Proposal Number(s):
[14631, 17180, 20015]
Open Access
Abstract: Recent clinical reports have highlighted the need for wild-type (WT) and mutant dual inhibitors of c-MET kinase for the treatment of cancer. We report herein a novel chemical series of ATP competitive type-III inhibitors of WT and D1228V mutant c-MET. Using a combination of structure-based drug design and computational analyses, ligand 2 was optimized to a highly selective chemical series with nanomolar activities in biochemical and cellular settings. Representatives of the series demonstrate excellent pharmacokinetic profiles in rat in vivo studies with promising free-brain exposures, paving the way for the design of brain permeable drugs for the treatment of c-MET driven cancers.
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Jun 2023
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
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Olivier A.
Pierrat
,
Manjuan
Liu
,
Gavin W.
Collie
,
Kartika N.
Shetty
,
Matthew J.
Rodrigues
,
Yann-Vai
Le Bihan
,
Emma A.
Gunnell
,
P. Craig
Mcandrew
,
Mark
Stubbs
,
Martin G.
Rowlands
,
Norhakim
Yahya
,
Erald
Shehu
,
Rachel
Talbot
,
Lisa
Pickard
,
Benjamin R.
Bellenie
,
Kwai-Ming J.
Cheung
,
Ludovic
Drouin
,
Paolo
Innocenti
,
Hannah
Woodward
,
Owen A.
Davis
,
Matthew G.
Lloyd
,
Ana
Varela
,
Rosemary
Huckvale
,
Fabio
Broccatelli
,
Michael
Carter
,
David
Galiwango
,
Angela
Hayes
,
Florence I.
Raynaud
,
Christopher
Bryant
,
Steven
Whittaker
,
Olivia W.
Rossanese
,
Swen
Hoelder
,
Rosemary
Burke
,
Rob L. M.
Van Montfort
Open Access
Abstract: By suppressing gene transcription through the recruitment of corepressor proteins, B-cell lymphoma 6 (BCL6) protein controls a transcriptional network required for the formation and maintenance of B-cell germinal centres. As BCL6 deregulation is implicated in the development of Diffuse Large B-Cell Lymphoma, we sought to discover novel small molecule inhibitors that disrupt the BCL6-corepressor protein–protein interaction (PPI). Here we report our hit finding and compound optimisation strategies, which provide insight into the multi-faceted orthogonal approaches that are needed to tackle this challenging PPI with small molecule inhibitors. Using a 1536-well plate fluorescence polarisation high throughput screen we identified multiple hit series, which were followed up by hit confirmation using a thermal shift assay, surface plasmon resonance and ligand-observed NMR. We determined X-ray structures of BCL6 bound to compounds from nine different series, enabling a structure-based drug design approach to improve their weak biochemical potency. We developed a time-resolved fluorescence energy transfer biochemical assay and a nano bioluminescence resonance energy transfer cellular assay to monitor cellular activity during compound optimisation. This workflow led to the discovery of novel inhibitors with respective biochemical and cellular potencies (IC50s) in the sub-micromolar and low micromolar range.
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Nov 2022
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
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Matthew G.
Lloyd
,
Rosemary
Huckvale
,
Kwai-Ming J.
Cheung
,
Matthew J.
Rodrigues
,
Gavin W.
Collie
,
Olivier A.
Pierrat
,
Mahad
Gatti Iou
,
Michael
Carter
,
Owen A.
Davis
,
P. Craig
Mcandrew
,
Emma
Gunnell
,
Yann-Vai
Le Bihan
,
Rachel
Talbot
,
Alan T.
Henley
,
Louise D.
Johnson
,
Angela T.
Hayes
,
Michael D.
Bright
,
Florence I.
Raynaud
,
Mirco
Meniconi
,
Rosemary
Burke
,
Rob L. M.
Van Montfort
,
Olivia W.
Rossanese
,
Benjamin R.
Bellenie
,
Swen
Hoelder
Diamond Proposal Number(s):
[14891, 20145]
Open Access
Abstract: We describe the optimization of modestly active starting points to potent inhibitors of BCL6 by growing into a subpocket, which was occupied by a network of five stably bound water molecules. Identifying potent inhibitors required not only forming new interactions in the subpocket but also perturbing the water network in a productive, potency-increasing fashion while controlling the physicochemical properties. We achieved this goal in a sequential manner by systematically probing the pocket and the water network, ultimately achieving a 100-fold improvement of activity. The most potent compounds displaced three of the five initial water molecules and formed hydrogen bonds with the remaining two. Compound 25 showed a promising profile for a lead compound with submicromolar inhibition of BCL6 in cells and satisfactory pharmacokinetic (PK) properties. Our work highlights the importance of finding productive ways to perturb existing water networks when growing into solvent-filled protein pockets.
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Nov 2021
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I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Gavin W.
Collie
,
Iacovos N.
Michaelides
,
Kevin
Embrey
,
Christopher J.
Stubbs
,
Ulf
Börjesson
,
Ian L.
Dale
,
Arjan
Snijder
,
Louise
Barlind
,
Kun
Song
,
Puneet
Khurana
,
Christopher
Phillips
,
R. Ian
Storer
Open Access
Abstract: We report here a fragment screen directed toward the c-MET kinase from which we discovered a series of inhibitors able to bind to a rare conformation of the protein in which the P-loop adopts a collapsed, or folded, arrangement. Preliminary SAR exploration led to an inhibitor (7) with nanomolar biochemical activity against c-MET and promising cell activity and kinase selectivity. These findings increase our structural understanding of the folded P-loop conformation of c-MET and provide a sound structural and chemical basis for further investigation of this underexplored yet potentially therapeutically exploitable conformational state.
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Dec 2020
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
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Benjamin R.
Bellenie
,
Kwai-Ming J.
Cheung
,
Ana
Varela
,
Olivier A.
Pierrat
,
Gavin W.
Collie
,
Gary M.
Box
,
Michael D.
Bright
,
Sharon
Gowan
,
Angela
Hayes
,
Matthew J.
Rodrigues
,
Kartika N.
Shetty
,
Michael
Carter
,
Owen A.
Davis
,
Alan T.
Henley
,
Paolo
Innocenti
,
Louise D.
Johnson
,
Manjuan
Liu
,
Selby
De Klerk
,
Yann-Vai
Le Bihan
,
Matthew G.
Lloyd
,
P. Craig
Mcandrew
,
Erald
Shehu
,
Rachel
Talbot
,
Hannah L.
Woodward
,
Rosemary
Burke
,
Vladimir
Kirkin
,
Rob L. M.
Van Montfort
,
Florence I.
Raynaud
,
Olivia W.
Rossanese
,
Swen
Hoelder
Diamond Proposal Number(s):
[14891, 20145]
Open Access
Abstract: Deregulation of the transcriptional repressor BCL6 enables tumorigenesis of germinal center B-cells, and hence BCL6 has been proposed as a therapeutic target for the treatment of diffuse large B-cell lymphoma (DLBCL). Herein we report the discovery of a series of benzimidazolone inhibitors of the protein-protein interaction between BCL6 and its co-repressors. A subset of these inhibitors were found to cause rapid degradation of BCL6, and optimization of pharmacokinetic properties led to the discovery of 5-((5-chloro-2-((3R,5S)-4,4-difluoro-3,5-dimethylpiperidin-1-yl)pyrimidin-4-yl)amino)-3-(3-hydroxy-3-methylbutyl)-1-methyl-1,3-dihydro-2H-benzo[d]imidazol-2-one (CCT369260), which reduces BCL6 levels in a lymphoma xenograft mouse model following oral dosing.
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Apr 2020
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Gavin W.
Collie
,
Cheryl M.
Koh
,
Daniel J.
O'Neill
,
Christopher J.
Stubbs
,
Puneet
Khurana
,
Alice
Eddershaw
,
Arjan
Snijder
,
Fredrik
Mauritzson
,
Louise
Barlind
,
Ian L.
Dale
,
Joseph
Shaw
,
Christopher
Phillips
,
Edward J.
Hennessy
,
Tony
Cheung
,
Ana J.
Narvaez
Diamond Proposal Number(s):
[14631]
Open Access
Abstract: Many small molecule inhibitors of the cMET receptor tyrosine kinase have been evaluated in clinical trials for the treatment of cancer and resistance-conferring mutations of cMET are beginning to be reported for a number of such compounds. There is now a need to understand specific cMET mutations at the molecular level, particularly concerning small molecule recognition. Towards this end we report here the first crystal structures of the recent clinically-observed resistance-conferring D1228V cMET mutant in complex with small molecule inhibitors, along with a crystal structure of wild-type cMET bound by the clinical compound savolitinib and supporting cellular, biochemical and biophysical data. Our findings indicate that the D1228V alteration induces conformational changes in the kinase which could have implications for small molecule in-hibitor design. The data we report here increases our molecular understanding of the D1228V cMET mutation and provides insight for future inhibitor design.
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Aug 2019
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I04-Macromolecular Crystallography
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Abstract: Tetra-substituted naphthalene diimide (ND) derivatives with positively charged termini are potent stabilizers of human telomeric and gene promoter DNA quadruplexes and inhibit the growth of human cancer cells in vitro and in vivo. The present study reports the enhancement of the pharmacological properties of earlier ND compounds using structure-based design. Crystal structures of three complexes with human telomeric intramolecular quadruplexes demonstrate that two of the four strongly basic N-methyl-piperazine groups can be replaced by less basic morpholine groups with no loss of intermolecular interactions in the grooves of the quadruplex. The new compounds retain high affinity to human telomeric quadruplex DNA but are 10-fold more potent against the MIA PaCa-2 pancreatic cancer cell line, with IC50 values of ?10 nM. The lead compound induces cellular senescence but does not inhibit telomerase activity at the nanomolar dosage levels required for inhibition of cellular proliferation. Gene array qPCR analysis of MIA PaCa-2 cells treated with the lead compound revealed significant dose-dependent modulation of a distinct subset of genes, including strong induction of DNA damage responsive genes CDKN1A, DDIT3, GADD45A/G, and PPM1D, and repression of genes involved in telomere maintenance, including hPOT1 and PARP1.
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Apr 2013
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I04-Macromolecular Crystallography
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Edwin
Nkansah
,
Rahi
Shah
,
Gary N.
Parkinson
,
Gavin W.
Collie
,
Jonathan
Palmer
,
Khondaker M.
Rahman
,
Tam T.
Bui
,
Alex F.
Drake
,
Jarmila
Husby
,
Stephen
Neidle
,
Giovanna
Zinzalla
,
David E.
Thurston
,
Andrew F.
Wilderspin
Abstract: The STAT3 transcription factor plays a central role in a wide range of cancer types where it is over-expressed. Previously, phosphorylation of this protein was thought to be a prerequisite for direct binding to DNA. However, we have now shown complete binding of a purified unphosphorylated STAT3 (uSTAT3) core directly to M67 DNA, the high affinity STAT3 target DNA sequence, by a protein electrophoretic mobility shift assay (PEMSA). Binding to M67 DNA was inhibited by addition of increasing concentrations of a phosphotyrosyl peptide. X-ray crystallography demonstrates one mode of binding that is similar to that known for the STAT3 core phosphorylated at Y705.
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Feb 2013
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I24-Microfocus Macromolecular Crystallography
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Abstract: The folding of the single-stranded 3? end of the human telomere into G-quadruplex arrangements inhibits the overhang from hybridizing with the RNA template of telomerase and halts telomere maintenance in cancer cells. The ability to thermally stabilize human telomeric DNA as a four-stranded G-quadruplex structure by developing selective small molecule compounds is a therapeutic path to regulating telomerase activity and thereby selectively inhibit cancer cell growth. The development of compounds with the necessary selectivity and affinity to target parallel-stranded G-quadruplex structures has proved particularly challenging to date, relying heavily upon limited structural data. We report here on a structure-based approach to the design of quadruplex-binding ligands to enhance affinity and selectivity for human telomeric DNA. Crystal structures have been determined of complexes between a 22-mer intramolecular human telomeric quadruplex and two potent tetra-substituted naphthalene diimide compounds, functionalized with positively charged N-methyl-piperazine side-chains. These compounds promote parallel-stranded quadruplex topology, binding exclusively to the 3? surface of each quadruplex. There are significant differences between the complexes in terms of ligand mobility and in the interactions with quadruplex grooves. One of the two ligands is markedly less mobile in the crystal complex and is more quadruplex-stabilizing, forming multiple electrostatic/hydrogen bond contacts with quadruplex phosphate groups. The data presented here provides a structural rationale for the biophysical (effects on quadruplex thermal stabilization) and biological data (inhibition of proliferation in cancer cell lines and evidence of in vivo antitumor activity) on compounds in this series and, thus, for the concept of telomere targeting with DNA quadruplex-binding small molecules.
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Jan 2012
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I04-Macromolecular Crystallography
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Abstract: Human telomeric DNA is now known to be transcribed into noncoding RNA sequences, termed TERRA. These sequences, which are believed to play roles in the regulation of telomere function, can form higher-order quadruplex structures and may themselves be the target of therapeutic intervention. The crystal structure of a TERRA quadruplex?acridine small-molecule complex at a resolution of 2.60 , is reported here and contrasts remarkably with the structure of the analogous DNA quadruplex complex. The bimolecular RNA complex has a parallel-stranded topology with propeller-like UUA loops. These loops are held in particular conformations by multiple hydrogen bonds involving the O2? hydroxyl groups of the ribonucleotide sugars and play an active role in binding the acridine molecules to the RNA quadruplex. By contrast, the analogous DNA quadruplex complex has simpler 1:1 acridine binding, with no loop involvement. There are significant loop conformational changes in the RNA quadruplex compared to the native TERRA quadruplex (Collie, G. W.; Haider, S. M.; Neidle, S.; Parkinson, G. N. Nucleic Acids Res. 2010, 38,, 5569?5580), which have implications for the future design of small molecules targeting TERRA quadruplexes, and RNA quadruplexes more generally.
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Feb 2011
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