Wolfson Institute for Biomedical Research

London, United Kingdom

Wolfson Institute for Biomedical Research

London, United Kingdom
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Harikumar K.B.,University of Texas M. D. Anderson Cancer Center | Kunnumakkara A.B.,University of Texas M. D. Anderson Cancer Center | Ochi N.,University of Texas M. D. Anderson Cancer Center | Tong Z.,University of Texas M. D. Anderson Cancer Center | And 19 more authors.
Molecular Cancer Therapeutics | Year: 2010

Protein kinase D (PKD) family members are increasingly implicated in multiple normal and abnormal biological functions, including signaling pathways that promote mitogenesis in pancreatic cancer. However, nothing is known about the effects of targeting PKD in pancreatic cancer. Our PKD inhibitor discovery program identified CRT0066101 as a specific inhibitor of all PKD isoforms. The aim of our study was to determine the effects of CRT0066101 in pancreatic cancer. Initially, we showed that autophosphorylated PKD1 and PKD2 (activated PKD1/2) are significantly upregulated in pancreatic cancer and that PKD1/2 are expressed in multiple pancreatic cancer cell lines. Using Panc-1 as a model system, we showed that CRT0066101 reduced bromodeoxyuridine incorporation; increased apoptosis; blocked neurotensin-induced PKD1/2 activation; reduced neurotensin-induced, PKD-mediated Hsp27 phosphorylation; attenuated PKD1-mediated NF-êB activation; and abrogated the expression of NF-κB-dependent proliferative and prosurvival proteins. We showed that CRT0066101 given orally (80 mg/kg/d) for 24 days significantly abrogated pancreatic cancer growth in Panc-1 subcutaneous xenograft model. Activated PKD1/2 expression in the treated tumor explants was significantly inhibited with peak tumor concentration (12 μmol/L) of CRT0066101 achieved within 2 hours after oral administration. Further, we showed that CRT0066101 given orally (80 mg/kg/d) for 21 days in Panc-1 orthotopic model potently blocked tumor growth in vivo. CRT0066101 significantly reduced Ki-67-positive proliferation index (P < 0.01), increased terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive apoptotic cells (P < 0.05), and abrogated the expression of NF-κB-dependent proteins including cyclin D1, survivin, and cIAP-1. Our results show for the first time that a PKD-specific small-molecule inhibitor CRT0066101 blocks pancreatic cancer growth in vivo and show that PKD is a novel therapeutic target in pancreatic cancer. ©2010 AACR.

Pemberton H.,CRUK London Research Institute | Pemberton H.,Institute of Cancer Research | Anderton E.,CRUK London Research Institute | Anderton E.,Wolfson Institute for Biomedical Research | And 14 more authors.
Genome Biology | Year: 2014

Background: Polycomb group proteins form multicomponent complexes that are important for establishing lineage-specific patterns of gene expression. Mammalian cells encode multiple permutations of the prototypic Polycomb repressive complex 1 (PRC1) with little evidence for functional specialization. An aim of this study is to determine whether the multiple orthologs that are co-expressed in human fibroblasts act on different target genes and whether their genomic location changes during cellular senescence.Results: Deep sequencing of chromatin immunoprecipitated with antibodies against CBX6, CBX7, CBX8, RING1 and RING2 reveals that the orthologs co-localize at multiple sites. PCR-based validation at representative loci suggests that a further six PRC1 proteins have similar binding patterns. Importantly, sequential chromatin immunoprecipitation with antibodies against different orthologs implies that multiple variants of PRC1 associate with the same DNA. At many loci, the binding profiles have a distinctive architecture that is preserved in two different types of fibroblast. Conversely, there are several hundred loci at which PRC1 binding is cell type-specific and, contrary to expectations, the presence of PRC1 does not necessarily equate with transcriptional silencing. Interestingly, the PRC1 binding profiles are preserved in senescent cells despite changes in gene expression.Conclusions: The multiple permutations of PRC1 in human fibroblasts congregate at common rather than specific sites in the genome and with overlapping but distinctive binding profiles in different fibroblasts. The data imply that the effects of PRC1 complexes on gene expression are more subtle than simply repressing the loci at which they bind. © 2014 Pemberton et al.; licensee BioMed Central Ltd.

Caldwell J.J.,Cancer Research UK Research Institute | Caldwell J.J.,University of Sussex | Welsh E.J.,Cancer Research UK Research Institute | Matijssen C.,Cancer Research UK Research Institute | And 13 more authors.
Journal of Medicinal Chemistry | Year: 2011

Structure-based design was applied to the optimization of a series of 2-(quinazolin-2-yl)phenols to generate potent and selective ATP-competitive inhibitors of the DNA damage response signaling enzyme checkpoint kinase 2 (CHK2). Structure-activity relationships for multiple substituent positions were optimized separately and in combination leading to the 2-(quinazolin-2-yl) phenol 46 (IC 50 3 nM) with good selectivity for CHK2 against CHK1 and a wider panel of kinases and with promising in vitro ADMET properties. Off-target activity at hERG ion channels shown by the core scaffold was successfully reduced by the addition of peripheral polar substitution. In addition to showing mechanistic inhibition of CHK2 in HT29 human colon cancer cells, a concentration dependent radioprotective effect in mouse thymocytes was demonstrated for the potent inhibitor 46 (CCT241533). © 2010 American Chemical Society.

Baud M.G.J.,Imperial College London | Leiser T.,FH Darmstadt | Haus P.,FH Darmstadt | Samlal S.,Wolfson Institute for Biomedical Research | And 11 more authors.
Journal of Medicinal Chemistry | Year: 2012

Psammaplin A (11c) is a marine metabolite previously reported to be a potent inhibitor of two classes of epigenetic enzymes: histone deacetylases and DNA methyltransferases. The design and synthesis of a focused library based on the psammaplin A core has been carried out to probe the molecular features of this molecule responsible for its activity. By direct in vitro assay of the free thiol generated upon reduction of the dimeric psammaplin scaffold, we have unambiguously demonstrated that 11c functions as a natural prodrug, with the reduced form being highly potent against HDAC1 in vitro (IC50 0.9 nM). Furthermore, we have shown it to have high isoform selectivity, being 360-fold selective for HDAC1 over HDAC6 and more than 1000-fold less potent against HDAC7 and HDAC8. SAR around our focused library revealed a number of features, most notably the oxime functionality to be important to this selectivity. Many of the compounds show significant cytotoxicity in A549, MCF7, and W138 cells, with the SAR of cytotoxicity correlating to HDAC inhibition. Furthermore, compound treatment causes upregulation of histone acetylation but little effect on tubulin acetylation. Finally, we have found no evidence for 11c functioning as a DNMT inhibitor. © 2012 American Chemical Society.

Heikkila T.,Wolfson Institute for Biomedical Research | Wheatley E.,Wolfson Institute for Biomedical Research | Crighton D.,Beatson Institute for Cancer Research | Schroder E.,Wolfson Institute for Biomedical Research | And 7 more authors.
PLoS ONE | Year: 2011

MRCKα and MRCKβ (myotonic dystrophy kinase-related Cdc42-binding kinases) belong to a subfamily of Rho GTPase activated serine/threonine kinases within the AGC-family that regulate the actomyosin cytoskeleton. Reflecting their roles in myosin light chain (MLC) phosphorylation, MRCKα and MRCKβ influence cell shape and motility. We report further evidence for MRCKα and MRCKβ contributions to the invasion of cancer cells in 3-dimensional matrix invasion assays. In particular, our results indicate that the combined inhibition of MRCKα and MRCKβ together with inhibition of ROCK kinases results in significantly greater effects on reducing cancer cell invasion than blocking either MRCK or ROCK kinases alone. To probe the kinase ligand pocket, we screened 159 kinase inhibitors in an in vitro MRCKβ kinase assay and found 11 compounds that inhibited enzyme activity >80% at 3 μM. Further analysis of three hits, Y-27632, Fasudil and TPCA-1, revealed low micromolar IC 50 values for MRCKα and MRCKβ. We also describe the crystal structure of MRCKβ in complex with inhibitors Fasudil and TPCA-1 bound to the active site of the kinase. These high-resolution structures reveal a highly conserved AGC kinase fold in a typical dimeric arrangement. The kinase domain is in an active conformation with a fully-ordered and correctly positioned αC helix and catalytic residues in a conformation competent for catalysis. Together, these results provide further validation for MRCK involvement in regulation of cancer cell invasion and present a valuable starting point for future structure-based drug discovery efforts. © 2011 Heikkila et al.

News Article | November 11, 2016
Site: www.eurekalert.org

Many pain-sensing nerves in the body are thought to respond to all types of 'painful events', but new research reveals that in fact most are specialized to respond to specific types such as heat, cold or mechanical pain Many pain-sensing nerves in the body are thought to respond to all types of 'painful events', but new UCL research in mice reveals that in fact most are specialised to respond to specific types such as heat, cold or mechanical pain. The study, published in Science Advances and funded by Wellcome and Arthritis Research UK, found that over 85% of pain-sensing neurons in whole organisms are sensitive to one specific type of painful stimulus. It was previously thought that most pain-sensing neurons were very similar, so the new finding could enable scientists to develop new specific painkillers for different pain conditions. Previous research using electrodes to monitor pain-sensing neurons had suggested that they respond to all types of pain, but the new study suggests that this recording technique may have altered the neuron's properties. "While the majority of neurons are specific to one type of pain, they can become universal pain sensors when the tissue is damaged," explains lead author Dr Edward Emery (UCL Wolfson Institute for Biomedical Research). "This may explain the discrepancies between our findings and those from other studies where more invasive approaches have been used." The team used a form of fluorescent activity-dependent imaging, where pain-sensing neurons in mice were genetically marked to emit a fluorescent glow when activated. The mice were briefly exposed to either a small pinch, cold water or hot water stimulus on one of their paws to see which neurons were activated. The results showed that over 85% of pain-sensing neurons were specific to one type of pain and did not react to others. "Our next step is to look at animal models for specific chronic pain conditions to see which neurons cells are activated," says senior author Professor John Wood (UCL Wolfson Institute for Biomedical Research). "We hope to identify the different neurons through which chronic pain can develop, so that focussed treatments can be developed. We use 'chronic pain' to describe all sorts of pain conditions with different causes, but we now need to differentiate them so that we can develop new specific treatments."

Hughes S.,Cancer Research UK Research Institute | Elustondo F.,Wolfson Institute for Biomedical Research | Di Fonzo A.,Cancer Research UK Research Institute | Leroux F.G.,Wolfson Institute for Biomedical Research | And 4 more authors.
Nature Structural and Molecular Biology | Year: 2012

CDC7 is a serine/threonine kinase that is essential for the initiation of eukaryotic DNA replication. CDC7 activity is controlled by its activator, DBF4. Here we present crystal structures of human CDC7-DBF4 in complex with a nucleotide or ATP-competing small molecules, revealing the active and inhibited forms of the kinase, respectively. DBF4 wraps around CDC7, burying approximately 6,000 Å 2 of hydrophobic molecular surface in a bipartite interface. The effector domain of DBF4, containing conserved motif C, is essential and sufficient to support CDC7 kinase activity by binding to the kinase N-terminal lobe and stabilizing its canonical αC helix. DBF4 motif M latches onto the C-terminal lobe of the kinase, acting as a tethering domain. Our results elucidate the structural basis for binding to and activation of CDC7 by DBF4 and provide a framework for the design of more potent and specific CDC7 inhibitors. © 2012 Nature America, Inc. All rights reserved.

Pachitariu M.,Gatsby Unit | Packer A.,Wolfson Institute for Biomedical Research | Pettit N.,Wolfson Institute for Biomedical Research | Dagleish H.,Wolfson Institute for Biomedical Research | And 2 more authors.
Advances in Neural Information Processing Systems | Year: 2013

Biological tissue is often composed of cells with similar morphologies replicated throughout large volumes and many biological applications rely on the accurate identification of these cells and their locations from image data. Here we develop a generative model that captures the regularities present in images composed of repeating elements of a few different types. Formally, the model can be described as convolutional sparse block coding. For inference we use a variant of convolutional matching pursuit adapted to block-based representations. We extend the KSVD learning algorithm to subspaces by retaining several principal vectors from the SVD decomposition instead of just one. Good models with little cross-talk between subspaces can be obtained by learning the blocks incrementally. We perform extensive experiments on simulated images and the inference algorithm consistently recovers a large proportion of the cells with a small number of false positives. We fit the convolutional model to noisy GCaMP6 two-photon images of spiking neurons and to Nissl-stained slices of cortical tissue and show that it recovers cell body locations without supervision. The flexibility of the block-based representation is reflected in the variability of the recovered cell shapes.

Melander R.J.,North Carolina State University | Selwood D.L.,Wolfson Institute for Biomedical Research
Chemical Biology and Drug Design | Year: 2015

Governments, academics and industry are beginning to listen to the medical communities call for new anti-bacterials. This special issue brings together diverse review articles on topics from economics and pricing to new discovery methods. © 2015 John Wiley & Sons A/S.

Flanagan J.U.,University of Auckland | Yosaatmadja Y.,University of Auckland | Teague R.M.,University of Auckland | Chai M.Z.L.,University of Auckland | And 2 more authors.
PLoS ONE | Year: 2012

Aldo-keto reductase 1C3 (AKR1C3) catalyses the NADPH dependent reduction of carbonyl groups in a number of important steroid and prostanoid molecules. The enzyme is also over-expressed in prostate and breast cancer and its expression is correlated with the aggressiveness of the disease. The steroid products of AKR1C3 catalysis are important in proliferative signalling of hormone-responsive cells, while the prostanoid products promote prostaglandin-dependent proliferative pathways. In these ways, AKR1C3 contributes to tumour development and maintenance, and suggest that inhibition of AKR1C3 activity is an attractive target for the development of new anti-cancer therapies. Non-steroidal anti-inflammatory drugs (NSAIDs) are one well-known class of compounds that inhibits AKR1C3, yet crystal structures have only been determined for this enzyme with flufenamic acid, indomethacin, and closely related analogues bound. While the flufenamic acid and indomethacin structures have been used to design novel inhibitors, they provide only limited coverage of the NSAIDs that inhibit AKR1C3 and that may be used for the development of new AKR1C3 targeted drugs. To understand how other NSAIDs bind to AKR1C3, we have determined ten crystal structures of AKR1C3 complexes that cover three different classes of NSAID, N-phenylanthranilic acids (meclofenamic acid, mefenamic acid), arylpropionic acids (flurbiprofen, ibuprofen, naproxen), and indomethacin analogues (indomethacin, sulindac, zomepirac). The N-phenylanthranilic and arylpropionic acids bind to common sites including the enzyme catalytic centre and a constitutive active site pocket, with the arylpropionic acids probing the constitutive pocket more effectively. By contrast, indomethacin and the indomethacin analogues sulindac and zomepirac, display three distinctly different binding modes that explain their relative inhibition of the AKR1C family members. This new data from ten crystal structures greatly broadens the base of structures available for future structure-guided drug discovery efforts. © 2012 Flanagan et al.

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