Ritzefeld, M.
Zhang, L.
Xiao, Z.
Andrei, S.A.
Boyd, O.
Masumoto, N.
Rodgers, U.R.
Artelsmair, M.
Sefer, L.
Hayes, A.
Gavriil, E.-.
Raynaud, F.I.
Burke, R.
Blagg, J.
Rzepa, H.S.
Siebold, C.
Magee, A.I.
Lanyon-Hogg, T.
Tate, E.W.
(2024). Design, Synthesis, and Evaluation of Inhibitors of Hedgehog Acyltransferase. J med chem,
Vol.67
(2),
pp. 1061-1078.
show abstract
Hedgehog signaling is involved in embryonic development and cancer growth. Functional activity of secreted Hedgehog signaling proteins is dependent on N-terminal palmitoylation, making the palmitoyl transferase Hedgehog acyltransferase (HHAT), a potential drug target and a series of 4,5,6,7-tetrahydrothieno[3,2-c]pyridines have been identified as HHAT inhibitors. Based on structural data, we designed and synthesized 37 new analogues which we profiled alongside 13 previously reported analogues in enzymatic and cellular assays. Our results show that a central amide linkage, a secondary amine, and (R)-configuration at the 4-position of the core are three key factors for inhibitory potency. Several potent analogues with low- or sub-μM IC50 against purified HHAT also inhibit Sonic Hedgehog (SHH) palmitoylation in cells and suppress the SHH signaling pathway. This work identifies IMP-1575 as the most potent cell-active chemical probe for HHAT function, alongside an inactive control enantiomer, providing tool compounds for validation of HHAT as a target in cellular assays..
Bouguenina, H.
Scarpino, A.
O'Hanlon, J.A.
Warne, J.
Wang, H.Z.
Wah Hak, L.C.
Sadok, A.
McAndrew, P.C.
Stubbs, M.
Pierrat, O.A.
Hahner, T.
Cabry, M.P.
Le Bihan, Y.-.
Mitsopoulos, C.
Sialana, F.J.
Roumeliotis, T.I.
Burke, R.
van Montfort, R.L.
Choudhari, J.
Chopra, R.
Caldwell, J.J.
Collins, I.
(2023). A Degron Blocking Strategy Towards Improved CRL4CRBN Recruiting PROTAC Selectivity. Chembiochem,
Vol.24
(23),
p. e202300351.
show abstract
Small molecules inducing protein degradation are important pharmacological tools to interrogate complex biology and are rapidly translating into clinical agents. However, to fully realise the potential of these molecules, selectivity remains a limiting challenge. Herein, we addressed the issue of selectivity in the design of CRL4CRBN recruiting PROteolysis TArgeting Chimeras (PROTACs). Thalidomide derivatives used to generate CRL4CRBN recruiting PROTACs have well described intrinsic monovalent degradation profiles by inducing the recruitment of neo-substrates, such as GSPT1, Ikaros and Aiolos. We leveraged structural insights from known CRL4CRBN neo-substrates to attenuate and indeed remove this monovalent degradation function in well-known CRL4CRBN molecular glues degraders, namely CC-885 and Pomalidomide. We then applied these design principles on a previously published BRD9 PROTAC (dBRD9-A) and generated an analogue with improved selectivity profile. Finally, we implemented a computational modelling pipeline to show that our degron blocking design does not impact PROTAC-induced ternary complex formation. We believe that the tools and principles presented in this work will be valuable to support the development of targeted protein degradation..
Liu, M.
Mirza, A.
McAndrew, P.C.
Thapaliya, A.
Pierrat, O.A.
Stubbs, M.
Hahner, T.
Chessum, N.E.
Innocenti, P.
Caldwell, J.
Cheeseman, M.D.
Bellenie, B.R.
van Montfort, R.L.
Newton, G.K.
Burke, R.
Collins, I.
Hoelder, S.
(2023). Determination of Ligand-Binding Affinity (Kd) Using Transverse Relaxation Rate (R2) in the Ligand-Observed 1H NMR Experiment and Applications to Fragment-Based Drug Discovery. J med chem,
Vol.66
(15),
pp. 10617-10627.
show abstract
full text
High hit rates from initial ligand-observed NMR screening can make it challenging to prioritize which hits to follow up, especially in cases where there are no available crystal structures of these hits bound to the target proteins or other strategies to provide affinity ranking. Here, we report a reproducible, accurate, and versatile quantitative ligand-observed NMR assay, which can determine Kd values of fragments in the affinity range of low μM to low mM using transverse relaxation rate R2 as the observable parameter. In this study, we examined the theory and proposed a mathematical formulation to obtain Kd values using non-linear regression analysis. We designed an assay format with automated sample preparation and simplified data analysis. Using tool compounds, we explored the assay reproducibility, accuracy, and detection limits. Finally, we used this assay to triage fragment hits, yielded from fragment screening against the CRBN/DDB1 complex..
Bouguenina, H.
Nicolaou, S.
Le Bihan, Y.-.
Bowling, E.A.
Calderon, C.
Caldwell, J.J.
Harrington, B.
Hayes, A.
McAndrew, P.C.
Mitsopoulos, C.
Sialana, F.J.
Scarpino, A.
Stubbs, M.
Thapaliya, A.
Tyagi, S.
Wang, H.Z.
Wood, F.
Burke, R.
Raynaud, F.
Choudhary, J.
van Montfort, R.L.
Sadok, A.
Westbrook, T.F.
Collins, I.
Chopra, R.
(2023). iTAG an optimized IMiD-induced degron for targeted protein degradation in human and murine cells. Iscience,
Vol.26
(7),
p. 107059.
show abstract
full text
To address the limitation associated with degron based systems, we have developed iTAG, a synthetic tag based on IMiDs/CELMoDs mechanism of action that improves and addresses the limitations of both PROTAC and previous IMiDs/CeLMoDs based tags. Using structural and sequence analysis, we systematically explored native and chimeric degron containing domains (DCDs) and evaluated their ability to induce degradation. We identified the optimal chimeric iTAG(DCD23 60aa) that elicits robust degradation of targets across cell types and subcellular localizations without exhibiting the well documented "hook effect" of PROTAC-based systems. We showed that iTAG can also induce target degradation by murine CRBN and enabled the exploration of natural neo-substrates that can be degraded by murine CRBN. Hence, the iTAG system constitutes a versatile tool to degrade targets across the human and murine proteome..
Saint-Dizier, F.
Matthews, T.P.
Gregson, A.M.
Prevet, H.
McHardy, T.
Colombano, G.
Saville, H.
Rowlands, M.
Ewens, C.
McAndrew, P.C.
Tomlin, K.
Guillotin, D.
Mak, G.W.
Drosopoulos, K.
Poursaitidis, I.
Burke, R.
van Montfort, R.
Linardopoulos, S.
Collins, I.
(2023). Discovery of 2-(3-Benzamidopropanamido)thiazole-5-carboxylate Inhibitors of the Kinesin HSET (KIFC1) and the Development of Cellular Target Engagement Probes. J med chem,
Vol.66
(4),
pp. 2622-2645.
show abstract
full text
The existence of multiple centrosomes in some cancer cells can lead to cell death through the formation of multipolar mitotic spindles and consequent aberrant cell division. Many cancer cells rely on HSET (KIFC1) to cluster the extra centrosomes into two groups to mimic the bipolar spindle formation of non-centrosome-amplified cells and ensure their survival. Here, we report the discovery of a novel 2-(3-benzamidopropanamido)thiazole-5-carboxylate with micromolar in vitro inhibition of HSET (KIFC1) through high-throughput screening and its progression to ATP-competitive compounds with nanomolar biochemical potency and high selectivity against the opposing mitotic kinesin Eg5. Induction of the multipolar phenotype was shown in centrosome-amplified human cancer cells treated with these inhibitors. In addition, a suitable linker position was identified to allow the synthesis of both fluorescent- and trans-cyclooctene (TCO)-tagged probes, which demonstrated direct compound binding to the HSET protein and confirmed target engagement in cells, through a click-chemistry approach..
Harnden, A.C.
Davis, O.A.
Box, G.M.
Hayes, A.
Johnson, L.D.
Henley, A.T.
de Haven Brandon, A.K.
Valenti, M.
Cheung, K.-.
Brennan, A.
Huckvale, R.
Pierrat, O.A.
Talbot, R.
Bright, M.D.
Akpinar, H.A.
Miller, D.S.
Tarantino, D.
Gowan, S.
de Klerk, S.
McAndrew, P.C.
Le Bihan, Y.-.
Meniconi, M.
Burke, R.
Kirkin, V.
van Montfort, R.L.
Raynaud, F.I.
Rossanese, O.W.
Bellenie, B.R.
Hoelder, S.
(2023). Discovery of an In Vivo Chemical Probe for BCL6 Inhibition by Optimization of Tricyclic Quinolinones. J med chem,
Vol.66
(8),
pp. 5892-5906.
show abstract
full text
B-cell lymphoma 6 (BCL6) is a transcriptional repressor and oncogenic driver of diffuse large B-cell lymphoma (DLBCL). Here, we report the optimization of our previously reported tricyclic quinolinone series for the inhibition of BCL6. We sought to improve the cellular potency and in vivo exposure of the non-degrading isomer, CCT373567, of our recently published degrader, CCT373566. The major limitation of our inhibitors was their high topological polar surface areas (TPSA), leading to increased efflux ratios. Reducing the molecular weight allowed us to remove polarity and decrease TPSA without considerably reducing solubility. Careful optimization of these properties, as guided by pharmacokinetic studies, led to the discovery of CCT374705, a potent inhibitor of BCL6 with a good in vivo profile. Modest in vivo efficacy was achieved in a lymphoma xenograft mouse model after oral dosing..
Pierrat, O.A.
Liu, M.
Collie, G.W.
Shetty, K.
Rodrigues, M.J.
Le Bihan, Y.-.
Gunnell, E.A.
McAndrew, P.C.
Stubbs, M.
Rowlands, M.G.
Yahya, N.
Shehu, E.
Talbot, R.
Pickard, L.
Bellenie, B.R.
Cheung, K.-.
Drouin, L.
Innocenti, P.
Woodward, H.
Davis, O.A.
Lloyd, M.G.
Varela, A.
Huckvale, R.
Broccatelli, F.
Carter, M.
Galiwango, D.
Hayes, A.
Raynaud, F.I.
Bryant, C.
Whittaker, S.
Rossanese, O.W.
Hoelder, S.
Burke, R.
van Montfort, R.L.
(2022). Discovering cell-active BCL6 inhibitors: effectively combining biochemical HTS with multiple biophysical techniques, X-ray crystallography and cell-based assays. Sci rep,
Vol.12
(1),
p. 18633.
show abstract
full text
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..
Davis, O.A.
Cheung, K.-.
Brennan, A.
Lloyd, M.G.
Rodrigues, M.J.
Pierrat, O.A.
Collie, G.W.
Le Bihan, Y.-.
Huckvale, R.
Harnden, A.C.
Varela, A.
Bright, M.D.
Eve, P.
Hayes, A.
Henley, A.T.
Carter, M.D.
McAndrew, P.C.
Talbot, R.
Burke, R.
van Montfort, R.L.
Raynaud, F.I.
Rossanese, O.W.
Meniconi, M.
Bellenie, B.R.
Hoelder, S.
(2022). Optimizing Shape Complementarity Enables the Discovery of Potent Tricyclic BCL6 Inhibitors. J med chem,
Vol.65
(12),
pp. 8169-8190.
show abstract
full text
To identify new chemical series with enhanced binding affinity to the BTB domain of B-cell lymphoma 6 protein, we targeted a subpocket adjacent to Val18. With no opportunities for strong polar interactions, we focused on attaining close shape complementarity by ring fusion onto our quinolinone lead series. Following exploration of different sized rings, we identified a conformationally restricted core which optimally filled the available space, leading to potent BCL6 inhibitors. Through X-ray structure-guided design, combined with efficient synthetic chemistry to make the resulting novel core structures, a >300-fold improvement in activity was obtained by the addition of seven heavy atoms..
Huckvale, R.
Harnden, A.C.
Cheung, K.-.
Pierrat, O.A.
Talbot, R.
Box, G.M.
Henley, A.T.
de Haven Brandon, A.K.
Hallsworth, A.E.
Bright, M.D.
Akpinar, H.A.
Miller, D.S.
Tarantino, D.
Gowan, S.
Hayes, A.
Gunnell, E.A.
Brennan, A.
Davis, O.A.
Johnson, L.D.
de Klerk, S.
McAndrew, C.
Le Bihan, Y.-.
Meniconi, M.
Burke, R.
Kirkin, V.
van Montfort, R.L.
Raynaud, F.I.
Rossanese, O.W.
Bellenie, B.R.
Hoelder, S.
(2022). Improved Binding Affinity and Pharmacokinetics Enable Sustained Degradation of BCL6 In Vivo. J med chem,
Vol.65
(12),
pp. 8191-8207.
show abstract
full text
The transcriptional repressor BCL6 is an oncogenic driver found to be deregulated in lymphoid malignancies. Herein, we report the optimization of our previously reported benzimidazolone molecular glue-type degrader CCT369260 to CCT373566, a highly potent probe suitable for sustained depletion of BCL6 in vivo. We observed a sharp degradation SAR, where subtle structural changes conveyed the ability to induce degradation of BCL6. CCT373566 showed modest in vivo efficacy in a lymphoma xenograft mouse model following oral dosing..
Lloyd, M.G.
Huckvale, R.
Cheung, K.-.
Rodrigues, M.J.
Collie, G.W.
Pierrat, O.A.
Gatti Iou, M.
Carter, M.
Davis, O.A.
McAndrew, P.C.
Gunnell, E.
Le Bihan, Y.-.
Talbot, R.
Henley, A.T.
Johnson, L.D.
Hayes, A.
Bright, M.D.
Raynaud, F.I.
Meniconi, M.
Burke, R.
van Montfort, R.L.
Rossanese, O.W.
Bellenie, B.R.
Hoelder, S.
(2021). Into Deep Water: Optimizing BCL6 Inhibitors by Growing into a Solvated Pocket. J med chem,
Vol.64
(23),
pp. 17079-17097.
show abstract
full text
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..
Davidson, K.
Grevitt, P.
Contreras-Gerenas, M.F.
Bridge, K.S.
Hermida, M.
Shah, K.M.
Mardakheh, F.K.
Stubbs, M.
Burke, R.
Casado, P.
Cutillas, P.R.
Martin, S.A.
Sharp, T.V.
(2021). Targeted therapy for LIMD1-deficient non-small cell lung cancer subtypes. Cell death dis,
Vol.12
(11),
p. 1075.
show abstract
full text
An early event in lung oncogenesis is loss of the tumour suppressor gene LIMD1 (LIM domains containing 1); this encodes a scaffold protein, which suppresses tumorigenesis via a number of different mechanisms. Approximately 45% of non-small cell lung cancers (NSCLC) are deficient in LIMD1, yet this subtype of NSCLC has been overlooked in preclinical and clinical investigations. Defining therapeutic targets in these LIMD1 loss-of-function patients is difficult due to a lack of 'druggable' targets, thus alternative approaches are required. To this end, we performed the first drug repurposing screen to identify compounds that confer synthetic lethality with LIMD1 loss in NSCLC cells. PF-477736 was shown to selectively target LIMD1-deficient cells in vitro through inhibition of multiple kinases, inducing cell death via apoptosis. Furthermore, PF-477736 was effective in treating LIMD1-/- tumours in subcutaneous xenograft models, with no significant effect in LIMD1+/+ cells. We have identified a novel drug tool with significant preclinical characterisation that serves as an excellent candidate to explore and define LIMD1-deficient cancers as a new therapeutic subgroup of critical unmet need..
Wilding, B.
Pasqua, A.E.
E A Chessum, N.
Pierrat, O.A.
Hahner, T.
Tomlin, K.
Shehu, E.
Burke, R.
Richards, G.M.
Whitton, B.
Arwert, E.N.
Thapaliya, A.
Salimraj, R.
van Montfort, R.
Skawinska, A.
Hayes, A.
Raynaud, F.
Chopra, R.
Jones, K.
Newton, G.
Cheeseman, M.D.
(2021). Investigating the phosphinic acid tripeptide mimetic DG013A as a tool compound inhibitor of the M1-aminopeptidase ERAP1. Bioorg med chem lett,
Vol.42,
p. 128050.
show abstract
full text
ERAP1 is a zinc-dependent M1-aminopeptidase that trims lipophilic amino acids from the N-terminus of peptides. Owing to its importance in the processing of antigens and regulation of the adaptive immune response, dysregulation of the highly polymorphic ERAP1 has been implicated in autoimmune disease and cancer. To test this hypothesis and establish the role of ERAP1 in these disease areas, high affinity, cell permeable and selective chemical probes are essential. DG013A 1, is a phosphinic acid tripeptide mimetic inhibitor with reported low nanomolar affinity for ERAP1. However, this chemotype is a privileged structure for binding to various metal-dependent peptidases and contains a highly charged phosphinic acid moiety, so it was unclear whether it would display the high selectivity and passive permeability required for a chemical probe. Therefore, we designed a new stereoselective route to synthesize a library of DG013A 1 analogues to determine the suitability of this compound as a cellular chemical probe to validate ERAP1 as a drug discovery target..
Acar, A.
Nichol, D.
Fernandez-Mateos, J.
Cresswell, G.D.
Barozzi, I.
Hong, S.P.
Trahearn, N.
Spiteri, I.
Stubbs, M.
Burke, R.
Stewart, A.
Caravagna, G.
Werner, B.
Vlachogiannis, G.
Maley, C.C.
Magnani, L.
Valeri, N.
Banerji, U.
Sottoriva, A.
(2020). Exploiting evolutionary steering to induce collateral drug sensitivity in cancer. Nat commun,
Vol.11
(1),
p. 1923.
show abstract
full text
Drug resistance mediated by clonal evolution is arguably the biggest problem in cancer therapy today. However, evolving resistance to one drug may come at a cost of decreased fecundity or increased sensitivity to another drug. These evolutionary trade-offs can be exploited using 'evolutionary steering' to control the tumour population and delay resistance. However, recapitulating cancer evolutionary dynamics experimentally remains challenging. Here, we present an approach for evolutionary steering based on a combination of single-cell barcoding, large populations of 108-109 cells grown without re-plating, longitudinal non-destructive monitoring of cancer clones, and mathematical modelling of tumour evolution. We demonstrate evolutionary steering in a lung cancer model, showing that it shifts the clonal composition of the tumour in our favour, leading to collateral sensitivity and proliferative costs. Genomic profiling revealed some of the mechanisms that drive evolved sensitivity. This approach allows modelling evolutionary steering strategies that can potentially control treatment resistance..
Bellenie, B.R.
Cheung, K.-.
Varela, A.
Pierrat, O.A.
Collie, G.W.
Box, G.M.
Bright, M.D.
Gowan, S.
Hayes, A.
Rodrigues, M.J.
Shetty, K.N.
Carter, M.
Davis, O.A.
Henley, A.T.
Innocenti, P.
Johnson, L.D.
Liu, M.
de Klerk, S.
Le Bihan, Y.-.
Lloyd, M.G.
McAndrew, P.C.
Shehu, E.
Talbot, R.
Woodward, H.L.
Burke, R.
Kirkin, V.
van Montfort, R.L.
Raynaud, F.I.
Rossanese, O.W.
Hoelder, S.
(2020). Achieving In Vivo Target Depletion through the Discovery and Optimization of Benzimidazolone BCL6 Degraders. J med chem,
Vol.63
(8),
pp. 4047-4068.
show abstract
full text
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..
Le Bihan, Y.-.
Lanigan, R.M.
Atrash, B.
McLaughlin, M.G.
Velupillai, S.
Malcolm, A.G.
England, K.S.
Ruda, G.F.
Mok, N.Y.
Tumber, A.
Tomlin, K.
Saville, H.
Shehu, E.
McAndrew, C.
Carmichael, L.
Bennett, J.M.
Jeganathan, F.
Eve, P.
Donovan, A.
Hayes, A.
Wood, F.
Raynaud, F.I.
Fedorov, O.
Brennan, P.E.
Burke, R.
van Montfort, R.L.
Rossanese, O.W.
Blagg, J.
Bavetsias, V.
(2019). 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. Eur j med chem,
Vol.177,
pp. 316-337.
show abstract
full text
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, 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 suggests 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..
Clarke, P.A.
Roe, T.
Swabey, K.
Hobbs, S.M.
McAndrew, C.
Tomlin, K.
Westwood, I.
Burke, R.
van Montfort, R.
Workman, P.
(2019). Dissecting mechanisms of resistance to targeted drug combination therapy in human colorectal cancer. Oncogene,
Vol.38
(25),
pp. 5076-5090.
show abstract
full text
Genomic alterations in cancer cells result in vulnerabilities that clinicians can exploit using molecularly targeted drugs, guided by knowledge of the tumour genotype. However, the selective activity of these drugs exerts an evolutionary pressure on cancers that can result in the outgrowth of resistant clones. Use of rational drug combinations can overcome resistance to targeted drugs, but resistance may eventually develop to combinatorial therapies. We selected MAPK- and PI3K-pathway inhibition in colorectal cancer as a model system to dissect out mechanisms of resistance. We focused on these signalling pathways because they are frequently activated in colorectal tumours, have well-characterised mutations and are clinically relevant. By treating a panel of 47 human colorectal cancer cell lines with a combination of MEK- and PI3K-inhibitors, we observe a synergistic inhibition of growth in almost all cell lines. Cells with KRAS mutations are less sensitive to PI3K inhibition, but are particularly sensitive to the combined treatment. Colorectal cancer cell lines with inherent or acquired resistance to monotherapy do not show a synergistic response to the combination treatment. Cells that acquire resistance to an MEK-PI3K inhibitor combination treatment still respond to an ERK-PI3K inhibitor regimen, but subsequently also acquire resistance to this combination treatment. Importantly, the mechanisms of resistance to MEK and PI3K inhibitors observed, MEK1/2 mutation or loss of PTEN, are similar to those detected in the clinic. ERK inhibitors may have clinical utility in overcoming resistance to MEK inhibitor regimes; however, we find a recurrent active site mutation of ERK2 that drives resistance to ERK inhibitors in mono- or combined regimens, suggesting that resistance will remain a hurdle. Importantly, we find that the addition of low concentrations of the BCL2-family inhibitor navitoclax to the MEK-PI3K inhibitor regimen improves the synergistic interaction and blocks the acquisition of resistance..
Anderhub, S.J.
Mak, G.W.
Gurden, M.D.
Faisal, A.
Drosopoulos, K.
Walsh, K.
Woodward, H.L.
Innocenti, P.
Westwood, I.M.
Naud, S.
Hayes, A.
Theofani, E.
Filosto, S.
Saville, H.
Burke, R.
van Montfort, R.L.
Raynaud, F.I.
Blagg, J.
Hoelder, S.
Eccles, S.A.
Linardopoulos, S.
(2019). High Proliferation Rate and a Compromised Spindle Assembly Checkpoint Confers Sensitivity to the MPS1 Inhibitor BOS172722 in Triple-Negative Breast Cancers. Mol cancer ther,
Vol.18
(10),
pp. 1696-1707.
show abstract
full text
BOS172722 (CCT289346) is a highly potent, selective, and orally bioavailable inhibitor of spindle assembly checkpoint kinase MPS1. BOS172722 treatment alone induces significant sensitization to death, particularly in highly proliferative triple-negative breast cancer (TNBC) cell lines with compromised spindle assembly checkpoint activity. BOS172722 synergizes with paclitaxel to induce gross chromosomal segregation defects caused by MPS1 inhibitor-mediated abrogation of the mitotic delay induced by paclitaxel treatment. In in vivo pharmacodynamic experiments, BOS172722 potently inhibits the spindle assembly checkpoint induced by paclitaxel in human tumor xenograft models of TNBC, as measured by inhibition of the phosphorylation of histone H3 and the phosphorylation of the MPS1 substrate, KNL1. This mechanistic synergy results in significant in vivo efficacy, with robust tumor regressions observed for the combination of BOS172722 and paclitaxel versus either agent alone in long-term efficacy studies in multiple human tumor xenograft TNBC models, including a patient-derived xenograft and a systemic metastasis model. The current target indication for BOS172722 is TNBC, based on their high sensitivity to MPS1 inhibition, the well-defined clinical patient population with high unmet need, and the synergy observed with paclitaxel..
Felisberto-Rodrigues, C.
Thomas, J.C.
McAndrew, C.
Le Bihan, Y.-.
Burke, R.
Workman, P.
van Montfort, R.L.
(2019). Structural and functional characterisation of human RNA helicase DHX8 provides insights into the mechanism of RNA-stimulated ADP release. Biochem j,
Vol.476
(18),
pp. 2521-2543.
show abstract
full text
DHX8 is a crucial DEAH-box RNA helicase involved in splicing and required for the release of mature mRNA from the spliceosome. Here, we report the biochemical characterisation of full-length human DHX8 and the catalytically active helicase core DHX8Δ547, alongside crystal structures of DHX8Δ547 bound to ADP and a structure of DHX8Δ547 bound to poly(A)6 single-strand RNA. Our results reveal that DHX8 has an in vitro binding preference for adenine-rich RNA and that RNA binding triggers the release of ADP through significant conformational flexibility in the conserved DEAH-, P-loop and hook-turn motifs. We demonstrate the importance of R620 and both the hook-turn and hook-loop regions for DHX8 helicase activity and propose that the hook-turn acts as a gatekeeper to regulate the directional movement of the 3' end of RNA through the RNA-binding channel. This study provides an in-depth understanding of the activity of DHX8 and contributes insights into the RNA-unwinding mechanisms of the DEAH-box helicase family..
Colombano, G.
Caldwell, J.J.
Matthews, T.P.
Bhatia, C.
Joshi, A.
McHardy, T.
Mok, N.Y.
Newbatt, Y.
Pickard, L.
Strover, J.
Hedayat, S.
Walton, M.I.
Myers, S.M.
Jones, A.M.
Saville, H.
McAndrew, C.
Burke, R.
Eccles, S.A.
Davies, F.E.
Bayliss, R.
Collins, I.
(2019). Binding to an Unusual Inactive Kinase Conformation by Highly Selective Inhibitors of Inositol-Requiring Enzyme 1α Kinase-Endoribonuclease. J med chem,
Vol.62
(5),
pp. 2447-2465.
show abstract
full text
A series of imidazo[1,2- b]pyridazin-8-amine kinase inhibitors were discovered to allosterically inhibit the endoribonuclease function of the dual kinase-endoribonuclease inositol-requiring enzyme 1α (IRE1α), a key component of the unfolded protein response in mammalian cells and a potential drug target in multiple human diseases. Inhibitor optimization gave compounds with high kinome selectivity that prevented endoplasmic reticulum stress-induced IRE1α oligomerization and phosphorylation, and inhibited endoribonuclease activity in human cells. X-ray crystallography showed the inhibitors to bind to a previously unreported and unusually disordered conformation of the IRE1α kinase domain that would be incompatible with back-to-back dimerization of the IRE1α protein and activation of the endoribonuclease function. These findings increase the repertoire of known IRE1α protein conformations and can guide the discovery of highly selective ligands for the IRE1α kinase site that allosterically inhibit the endoribonuclease..
Lampis, A.
Carotenuto, P.
Vlachogiannis, G.
Cascione, L.
Hedayat, S.
Burke, R.
Clarke, P.
Bosma, E.
Simbolo, M.
Scarpa, A.
Yu, S.
Cole, R.
Smyth, E.
Mateos, J.F.
Begum, R.
Hezelova, B.
Eltahir, Z.
Wotherspoon, A.
Fotiadis, N.
Bali, M.A.
Nepal, C.
Khan, K.
Stubbs, M.
Hahne, J.C.
Gasparini, P.
Guzzardo, V.
Croce, C.M.
Eccles, S.
Fassan, M.
Cunningham, D.
Andersen, J.B.
Workman, P.
Valeri, N.
Braconi, C.
(2018). MIR21 Drives Resistance to Heat Shock Protein 90 Inhibition in Cholangiocarcinoma. Gastroenterology,
Vol.154
(4),
pp. 1066-1079.e5.
show abstract
full text
BACKGROUND & AIMS: Cholangiocarcinomas (CCA) are resistant to chemotherapy, so new therapeutic agents are needed. We performed a screen to identify small-molecule compounds that are active against CCAs. Levels of microRNA 21 (MIR21 or miRNA21) are increased in CCAs. We investigated whether miRNA21 mediates resistance of CCA cells and organoids to HSP90 inhibitors. METHODS: We performed a high-throughput screen of 484 small-molecule compounds to identify those that reduced viability of 6 human CCA cell lines. We tested the effects of HSP90 inhibitors on cells with disruption of the MIR21 gene, cells incubated with MIR21 inhibitors, and stable cell lines with inducible expression of MIR21. We obtained CCA biopsies from patients, cultured them as organoids (patient-derived organoids). We assessed their architecture, mutation and gene expression patterns, response to compounds in culture, and when grown as subcutaneous xenograft tumors in mice. RESULTS: Cells with IDH1 and PBRM1 mutations had the highest level of sensitivity to histone deacetylase inhibitors. HSP90 inhibitors were effective in all cell lines, irrespective of mutations. Sensitivity of cells to HSP90 inhibitors correlated inversely with baseline level of MIR21. Disruption of MIR21 increased cell sensitivity to HSP90 inhibitors. CCA cells that expressed transgenic MIR21 were more resistant to HSP90 inhibitors than cells transfected with control vectors; inactivation of MIR21 in these cells restored sensitivity to these agents. MIR21 was shown to target the DnaJ heat shock protein family (Hsp40) member B5 (DNAJB5). Transgenic expression of DNAJB5 in CCA cells that overexpressed MIR21 re-sensitized them to HSP90 inhibitors. Sensitivity of patient-derived organoids to HSP90 inhibitors, in culture and when grown as xenograft tumors in mice, depended on expression of miRNA21. CONCLUSIONS: miRNA21 appears to mediate resistance of CCA cells to HSP90 inhibitors by reducing levels of DNAJB5. HSP90 inhibitors might be developed for the treatment of CCA and miRNA21 might be a marker of sensitivity to these agents..
Chessum, N.E.
Sharp, S.Y.
Caldwell, J.J.
Pasqua, A.E.
Wilding, B.
Colombano, G.
Collins, I.
Ozer, B.
Richards, M.
Rowlands, M.
Stubbs, M.
Burke, R.
McAndrew, P.C.
Clarke, P.A.
Workman, P.
Cheeseman, M.D.
Jones, K.
(2018). Demonstrating In-Cell Target Engagement Using a Pirin Protein Degradation Probe (CCT367766). J med chem,
Vol.61
(3),
pp. 918-933.
show abstract
full text
Demonstrating intracellular protein target engagement is an essential step in the development and progression of new chemical probes and potential small molecule therapeutics. However, this can be particularly challenging for poorly studied and noncatalytic proteins, as robust proximal biomarkers are rarely known. To confirm that our recently discovered chemical probe 1 (CCT251236) binds the putative transcription factor regulator pirin in living cells, we developed a heterobifunctional protein degradation probe. Focusing on linker design and physicochemical properties, we generated a highly active probe 16 (CCT367766) in only three iterations, validating our efficient strategy for degradation probe design against nonvalidated protein targets..
Meyers, J.
Chessum, N.E.
Ali, S.
Mok, N.Y.
Wilding, B.
Pasqua, A.E.
Rowlands, M.
Tucker, M.J.
Evans, L.E.
Rye, C.S.
O'Fee, L.
Le Bihan, Y.-.
Burke, R.
Carter, M.
Workman, P.
Blagg, J.
Brown, N.
van Montfort, R.L.
Jones, K.
Cheeseman, M.D.
(2018). Privileged Structures and Polypharmacology within and between Protein Families. Acs med chem lett,
Vol.9
(12),
pp. 1199-1204.
show abstract
full text
Polypharmacology is often a key contributor to the efficacy of a drug, but is also a potential risk. We investigated two hits discovered via a cell-based phenotypic screen, the CDK9 inhibitor CCT250006 (1) and the pirin ligand CCT245232 (2), to establish methodology to elucidate their secondary protein targets. Using computational pocket-based analysis, we discovered intrafamily polypharmacology for our kinase inhibitor, despite little overall sequence identity. The interfamily polypharmacology of 2 with B-Raf was used to discover a novel pirin ligand from a very small but privileged compound library despite no apparent ligand or binding site similarity. Our data demonstrates that in areas of drug discovery where intrafamily polypharmacology is often an issue, ligand dissimilarity cannot necessarily be used to assume different off-target profiles and that understanding interfamily polypharmacology will be important in the future to reduce the risk of idiopathic toxicity and in the design of screening libraries..
Liu, M.
Mallinger, A.
Tortorici, M.
Newbatt, Y.
Richards, M.
Mirza, A.
van Montfort, R.L.
Burke, R.
Blagg, J.
Kaserer, T.
(2018). Evaluation of APOBEC3B Recognition Motifs by NMR Reveals Preferred Substrates. Acs chem biol,
Vol.13
(9),
pp. 2427-2432.
show abstract
full text
APOBEC3B (A3B) deamination activity on ssDNA is considered a contributing factor to tumor heterogeneity and drug resistance in a number of human cancers. Despite its clinical impact, little is known about A3B ssDNA substrate preference. We have used nuclear magnetic resonance to monitor the catalytic turnover of A3B substrates in real-time. This study reports preferred nucleotide sequences for A3B substrates, including optimized 4-mer oligonucleotides, and reveals a breadth of substrate recognition that includes DNA sequences known to be mutated in drug-resistant cancer clones. Our results are consistent with available clinical and structural data and may inform the design of substrate-based A3B inhibitors..
Vlachogiannis, G.
Hedayat, S.
Vatsiou, A.
Jamin, Y.
Fernández-Mateos, J.
Khan, K.
Lampis, A.
Eason, K.
Huntingford, I.
Burke, R.
Rata, M.
Koh, D.-.
Tunariu, N.
Collins, D.
Hulkki-Wilson, S.
Ragulan, C.
Spiteri, I.
Moorcraft, S.Y.
Chau, I.
Rao, S.
Watkins, D.
Fotiadis, N.
Bali, M.
Darvish-Damavandi, M.
Lote, H.
Eltahir, Z.
Smyth, E.C.
Begum, R.
Clarke, P.A.
Hahne, J.C.
Dowsett, M.
de Bono, J.
Workman, P.
Sadanandam, A.
Fassan, M.
Sansom, O.J.
Eccles, S.
Starling, N.
Braconi, C.
Sottoriva, A.
Robinson, S.P.
Cunningham, D.
Valeri, N.
(2018). Patient-derived organoids model treatment response of metastatic gastrointestinal cancers. Science,
Vol.359
(6378),
pp. 920-926.
show abstract
full text
Patient-derived organoids (PDOs) have recently emerged as robust preclinical models; however, their potential to predict clinical outcomes in patients has remained unclear. We report on a living biobank of PDOs from metastatic, heavily pretreated colorectal and gastroesophageal cancer patients recruited in phase 1/2 clinical trials. Phenotypic and genotypic profiling of PDOs showed a high degree of similarity to the original patient tumors. Molecular profiling of tumor organoids was matched to drug-screening results, suggesting that PDOs could complement existing approaches in defining cancer vulnerabilities and improving treatment responses. We compared responses to anticancer agents ex vivo in organoids and PDO-based orthotopic mouse tumor xenograft models with the responses of the patients in clinical trials. Our data suggest that PDOs can recapitulate patient responses in the clinic and could be implemented in personalized medicine programs..
Woodward, H.L.
Innocenti, P.
Cheung, K.-.
Hayes, A.
Roberts, J.
Henley, A.T.
Faisal, A.
Mak, G.W.
Box, G.
Westwood, I.M.
Cronin, N.
Carter, M.
Valenti, M.
De Haven Brandon, A.
O'Fee, L.
Saville, H.
Schmitt, J.
Burke, R.
Broccatelli, F.
van Montfort, R.L.
Raynaud, F.I.
Eccles, S.A.
Linardopoulos, S.
Blagg, J.
Hoelder, S.
(2018). Introduction of a Methyl Group Curbs Metabolism of Pyrido[3,4- d]pyrimidine Monopolar Spindle 1 (MPS1) Inhibitors and Enables the Discovery of the Phase 1 Clinical Candidate N2-(2-Ethoxy-4-(4-methyl-4 H-1,2,4-triazol-3-yl)phenyl)-6-methyl- N8-neopentylpyrido[3,4- d]pyrimidine-2,8-diamine (BOS172722). J med chem,
Vol.61
(18),
pp. 8226-8240.
show abstract
Monopolar spindle 1 (MPS1) occupies a central role in mitosis and is one of the main components of the spindle assembly checkpoint. The MPS1 kinase is an attractive cancer target, and herein, we report the discovery of the clinical candidate BOS172722. The starting point for our work was a series of pyrido[3,4- d]pyrimidine inhibitors that demonstrated excellent potency and kinase selectivity but suffered from rapid turnover in human liver microsomes (HLM). Optimizing HLM stability proved challenging since it was not possible to identify a consistent site of metabolism and lowering lipophilicity proved unsuccessful. Key to overcoming this problem was the finding that introduction of a methyl group at the 6-position of the pyrido[3,4- d]pyrimidine core significantly improved HLM stability. Met ID studies suggested that the methyl group suppressed metabolism at the distant aniline portion of the molecule, likely by blocking the preferred pharmacophore through which P450 recognized the compound. This work ultimately led to the discovery of BOS172722 as a Phase 1 clinical candidate..
Cheeseman, M.D.
Chessum, N.E.
Rye, C.S.
Pasqua, A.E.
Tucker, M.J.
Wilding, B.
Evans, L.E.
Lepri, S.
Richards, M.
Sharp, S.Y.
Ali, S.
Rowlands, M.
O'Fee, L.
Miah, A.
Hayes, A.
Henley, A.T.
Powers, M.
Te Poele, R.
De Billy, E.
Pellegrino, L.
Raynaud, F.
Burke, R.
van Montfort, R.L.
Eccles, S.A.
Workman, P.
Jones, K.
(2017). Discovery of a Chemical Probe Bisamide (CCT251236): An Orally Bioavailable Efficacious Pirin Ligand from a Heat Shock Transcription Factor 1 (HSF1) Phenotypic Screen. J med chem,
Vol.60
(1),
pp. 180-201.
show abstract
full text
Phenotypic screens, which focus on measuring and quantifying discrete cellular changes rather than affinity for individual recombinant proteins, have recently attracted renewed interest as an efficient strategy for drug discovery. In this article, we describe the discovery of a new chemical probe, bisamide (CCT251236), identified using an unbiased phenotypic screen to detect inhibitors of the HSF1 stress pathway. The chemical probe is orally bioavailable and displays efficacy in a human ovarian carcinoma xenograft model. By developing cell-based SAR and using chemical proteomics, we identified pirin as a high affinity molecular target, which was confirmed by SPR and crystallography..
Faisal, A.
Mak, G.W.
Gurden, M.D.
Xavier, C.P.
Anderhub, S.J.
Innocenti, P.
Westwood, I.M.
Naud, S.
Hayes, A.
Box, G.
Valenti, M.R.
De Haven Brandon, A.K.
O'Fee, L.
Schmitt, J.
Woodward, H.L.
Burke, R.
vanMontfort, R.L.
Blagg, J.
Raynaud, F.I.
Eccles, S.A.
Hoelder, S.
Linardopoulos, S.
(2017). Characterisation of CCT271850, a selective, oral and potent MPS1 inhibitor, used to directly measure in vivo MPS1 inhibition vs therapeutic efficacy. Br j cancer,
Vol.116
(9),
pp. 1166-1176.
show abstract
full text
BACKGROUND: The main role of the cell cycle is to enable error-free DNA replication, chromosome segregation and cytokinesis. One of the best characterised checkpoint pathways is the spindle assembly checkpoint, which prevents anaphase onset until the appropriate attachment and tension across kinetochores is achieved. MPS1 kinase activity is essential for the activation of the spindle assembly checkpoint and has been shown to be deregulated in human tumours with chromosomal instability and aneuploidy. Therefore, MPS1 inhibition represents an attractive strategy to target cancers. METHODS: To evaluate CCT271850 cellular potency, two specific antibodies that recognise the activation sites of MPS1 were used and its antiproliferative activity was determined in 91 human cancer cell lines. DLD1 cells with induced GFP-MPS1 and HCT116 cells were used in in vivo studies to directly measure MPS1 inhibition and efficacy of CCT271850 treatment. RESULTS: CCT271850 selectively and potently inhibits MPS1 kinase activity in biochemical and cellular assays and in in vivo models. Mechanistically, tumour cells treated with CCT271850 acquire aberrant numbers of chromosomes and the majority of cells divide their chromosomes without proper alignment because of abrogation of the mitotic checkpoint, leading to cell death. We demonstrated a moderate level of efficacy of CCT271850 as a single agent in a human colorectal carcinoma xenograft model. CONCLUSIONS: CCT271850 is a potent, selective and orally bioavailable MPS1 kinase inhibitor. On the basis of in vivo pharmacodynamic vs efficacy relationships, we predict that more than 80% inhibition of MPS1 activity for at least 24 h is required to achieve tumour stasis or regression by CCT271850..
Hatch, S.B.
Yapp, C.
Montenegro, R.C.
Savitsky, P.
Gamble, V.
Tumber, A.
Ruda, G.F.
Bavetsias, V.
Fedorov, O.
Atrash, B.
Raynaud, F.
Lanigan, R.
Carmichael, L.
Tomlin, K.
Burke, R.
Westaway, S.M.
Brown, J.A.
Prinjha, R.K.
Martinez, E.D.
Oppermann, U.
Schofield, C.J.
Bountra, C.
Kawamura, A.
Blagg, J.
Brennan, P.E.
Rossanese, O.
Müller, S.
(2017). Assessing histone demethylase inhibitors in cells: lessons learned. Epigenetics chromatin,
Vol.10,
p. 9.
show abstract
full text
BACKGROUND: Histone lysine demethylases (KDMs) are of interest as drug targets due to their regulatory roles in chromatin organization and their tight associations with diseases including cancer and mental disorders. The first KDM inhibitors for KDM1 have entered clinical trials, and efforts are ongoing to develop potent, selective and cell-active 'probe' molecules for this target class. Robust cellular assays to assess the specific engagement of KDM inhibitors in cells as well as their cellular selectivity are a prerequisite for the development of high-quality inhibitors. Here we describe the use of a high-content cellular immunofluorescence assay as a method for demonstrating target engagement in cells. RESULTS: A panel of assays for the Jumonji C subfamily of KDMs was developed to encompass all major branches of the JmjC phylogenetic tree. These assays compare compound activity against wild-type KDM proteins to a catalytically inactive version of the KDM, in which residues involved in the active-site iron coordination are mutated to inactivate the enzyme activity. These mutants are critical for assessing the specific effect of KDM inhibitors and for revealing indirect effects on histone methylation status. The reported assays make use of ectopically expressed demethylases, and we demonstrate their use to profile several recently identified classes of KDM inhibitors and their structurally matched inactive controls. The generated data correlate well with assay results assessing endogenous KDM inhibition and confirm the selectivity observed in biochemical assays with isolated enzymes. We find that both cellular permeability and competition with 2-oxoglutarate affect the translation of biochemical activity to cellular inhibition. CONCLUSIONS: High-content-based immunofluorescence assays have been established for eight KDM members of the 2-oxoglutarate-dependent oxygenases covering all major branches of the JmjC-KDM phylogenetic tree. The usage of both full-length, wild-type and catalytically inactive mutant ectopically expressed protein, as well as structure-matched inactive control compounds, allowed for detection of nonspecific effects causing changes in histone methylation as a result of compound toxicity. The developed assays offer a histone lysine demethylase family-wide tool for assessing KDM inhibitors for cell activity and on-target efficacy. In addition, the presented data may inform further studies to assess the cell-based activity of histone lysine methylation inhibitors..
Vaughan, L.
Clarke, P.A.
Barker, K.
Chanthery, Y.
Gustafson, C.W.
Tucker, E.
Renshaw, J.
Raynaud, F.
Li, X.
Burke, R.
Jamin, Y.
Robinson, S.P.
Pearson, A.
Maira, M.
Weiss, W.A.
Workman, P.
Chesler, L.
(2016). Inhibition of mTOR-kinase destabilizes MYCN and is a potential therapy for MYCN-dependent tumors. Oncotarget,
Vol.7
(36),
pp. 57525-57544.
show abstract
full text
MYC oncoproteins deliver a potent oncogenic stimulus in several human cancers, making them major targets for drug development, but efforts to deliver clinically practical therapeutics have not yet been realized. In childhood cancer, aberrant expression of MYC and MYCN genes delineates a group of aggressive tumours responsible for a major proportion of pediatric cancer deaths. We designed a chemical-genetic screen that identifies compounds capable of enhancing proteasomal elimination of MYCN oncoprotein. We isolated several classes of compound that selectively kill MYCN expressing cells and we focus on inhibitors of PI3K/mTOR pathway in this study. We show that PI3K/mTOR inhibitors selectively killed MYCN-expressing neuroblastoma tumor cells, and induced significant apoptosis of transgenic MYCN-driven neuroblastoma tumors concomitant with elimination of MYCN protein in vivo. Mechanistically, the ability of these compounds to degrade MYCN requires complete blockade of mTOR but not PI3 kinase activity and we highlight NVP-BEZ235 as a PI3K/mTOR inhibitor with an ideal activity profile. These data establish that MYCN expression is a marker indicative of likely clinical sensitivity to mTOR inhibition, and provide a rationale for the selection of clinical candidate MYCN-destabilizers likely to be useful for the treatment of MYCN-driven cancers..
Jones, A.M.
Westwood, I.M.
Osborne, J.D.
Matthews, T.P.
Cheeseman, M.D.
Rowlands, M.G.
Jeganathan, F.
Burke, R.
Lee, D.
Kadi, N.
Liu, M.
Richards, M.
McAndrew, C.
Yahya, N.
Dobson, S.E.
Jones, K.
Workman, P.
Collins, I.
van Montfort, R.L.
(2016). A fragment-based approach applied to a highly flexible target: Insights and challenges towards the inhibition of HSP70 isoforms. Sci rep,
Vol.6,
p. 34701.
show abstract
full text
The heat shock protein 70s (HSP70s) are molecular chaperones implicated in many cancers and of significant interest as targets for novel cancer therapies. Several HSP70 inhibitors have been reported, but because the majority have poor physicochemical properties and for many the exact mode of action is poorly understood, more detailed mechanistic and structural insight into ligand-binding to HSP70s is urgently needed. Here we describe the first comprehensive fragment-based inhibitor exploration of an HSP70 enzyme, which yielded an amino-quinazoline fragment that was elaborated to a novel ATP binding site ligand with different physicochemical properties to known adenosine-based HSP70 inhibitors. Crystal structures of amino-quinazoline ligands bound to the different conformational states of the HSP70 nucleotide binding domain highlighted the challenges of a fragment-based approach when applied to this particular flexible enzyme class with an ATP-binding site that changes shape and size during its catalytic cycle. In these studies we showed that Ser275 is a key residue in the selective binding of ATP. Additionally, the structural data revealed a potential functional role for the ATP ribose moiety in priming the protein for the formation of the ATP-bound pre-hydrolysis complex by influencing the conformation of one of the phosphate binding loops..
Mallinger, A.
Schiemann, K.
Rink, C.
Sejberg, J.
Honey, M.A.
Czodrowski, P.
Stubbs, M.
Poeschke, O.
Busch, M.
Schneider, R.
Schwarz, D.
Musil, D.
Burke, R.
Urbahns, K.
Workman, P.
Wienke, D.
Clarke, P.A.
Raynaud, F.I.
Eccles, S.A.
Esdar, C.
Rohdich, F.
Blagg, J.
(2016). 2,8-Disubstituted-1,6-Naphthyridines and 4,6-Disubstituted-Isoquinolines with Potent, Selective Affinity for CDK8/19. Acs med chem lett,
Vol.7
(6),
pp. 573-578.
show abstract
full text
We demonstrate a designed scaffold-hop approach to the discovery of 2,8-disubstituted-1,6-naphthyridine- and 4,6-disubstituted-isoquinoline-based dual CDK8/19 ligands. Optimized compounds in both series exhibited rapid aldehyde oxidase-mediated metabolism, which could be abrogated by introduction of an amino substituent at C5 of the 1,6-naphthyridine scaffold or at C1 of the isoquinoline scaffold. Compounds 51 and 59 were progressed to in vivo pharmacokinetic studies, and 51 also demonstrated sustained inhibition of STAT1(SER727) phosphorylation, a biomarker of CDK8 inhibition, in an SW620 colorectal carcinoma human tumor xenograft model following oral dosing..
Mallinger, A.
Schiemann, K.
Rink, C.
Stieber, F.
Calderini, M.
Crumpler, S.
Stubbs, M.
Adeniji-Popoola, O.
Poeschke, O.
Busch, M.
Czodrowski, P.
Musil, D.
Schwarz, D.
Ortiz-Ruiz, M.-.
Schneider, R.
Thai, C.
Valenti, M.
de Haven Brandon, A.
Burke, R.
Workman, P.
Dale, T.
Wienke, D.
Clarke, P.A.
Esdar, C.
Raynaud, F.I.
Eccles, S.A.
Rohdich, F.
Blagg, J.
(2016). Discovery of Potent, Selective, and Orally Bioavailable Small-Molecule Modulators of the Mediator Complex-Associated Kinases CDK8 and CDK19. J med chem,
Vol.59
(3),
pp. 1078-1101.
show abstract
full text
The Mediator complex-associated cyclin-dependent kinase CDK8 has been implicated in human disease, particularly in colorectal cancer where it has been reported as a putative oncogene. Here we report the discovery of 109 (CCT251921), a potent, selective, and orally bioavailable inhibitor of CDK8 with equipotent affinity for CDK19. We describe a structure-based design approach leading to the discovery of a 3,4,5-trisubstituted-2-aminopyridine series and present the application of physicochemical property analyses to successfully reduce in vivo metabolic clearance, minimize transporter-mediated biliary elimination while maintaining acceptable aqueous solubility. Compound 109 affords the optimal compromise of in vitro biochemical, pharmacokinetic, and physicochemical properties and is suitable for progression to animal models of cancer..
Rodgers, U.R.
Lanyon-Hogg, T.
Masumoto, N.
Ritzefeld, M.
Burke, R.
Blagg, J.
Magee, A.I.
Tate, E.W.
(2016). Characterization of Hedgehog Acyltransferase Inhibitors Identifies a Small Molecule Probe for Hedgehog Signaling by Cancer Cells. Acs chem biol,
Vol.11
(12),
pp. 3256-3262.
show abstract
full text
The Sonic Hedgehog (Shh) signaling pathway plays a critical role during embryonic development and cancer progression. N-terminal palmitoylation of Shh by Hedgehog acyltransferase (Hhat) is essential for efficient signaling, raising interest in Hhat as a novel drug target. A recently identified series of dihydrothienopyridines has been proposed to function via this mode of action; however, the lead compound in this series (RUSKI-43) was subsequently shown to possess cytotoxic activity unrelated to canonical Shh signaling. To identify a selective chemical probe for cellular studies, we profiled three RUSKI compounds in orthogonal cell-based assays. We found that RUSKI-43 exhibits off-target cytotoxicity, masking its effect on Hhat-dependent signaling, hence results obtained with this compound in cells should be treated with caution. In contrast, RUSKI-201 showed no off-target cytotoxicity, and quantitative whole-proteome palmitoylation profiling with a bioorthogonal alkyne-palmitate reporter demonstrated specific inhibition of Hhat in cells. RUSKI-201 is the first selective Hhat chemical probe in cells and should be used in future studies of Hhat catalytic function..
Tisi, D.
Chiarparin, E.
Tamanini, E.
Pathuri, P.
Coyle, J.E.
Hold, A.
Holding, F.P.
Amin, N.
Martin, A.C.
Rich, S.J.
Berdini, V.
Yon, J.
Acklam, P.
Burke, R.
Drouin, L.
Harmer, J.E.
Jeganathan, F.
van Montfort, R.L.
Newbatt, Y.
Tortorici, M.
Westlake, M.
Wood, A.
Hoelder, S.
Heightman, T.D.
(2016). Structure of the Epigenetic Oncogene MMSET and Inhibition by N-Alkyl Sinefungin Derivatives. Acs chem biol,
Vol.11
(11),
pp. 3093-3105.
show abstract
The members of the NSD subfamily of lysine methyl transferases are compelling oncology targets due to the recent characterization of gain-of-function mutations and translocations in several hematological cancers. To date, these proteins have proven intractable to small molecule inhibition. Here, we present initial efforts to identify inhibitors of MMSET (aka NSD2 or WHSC1) using solution phase and crystal structural methods. On the basis of 2D NMR experiments comparing NSD1 and MMSET structural mobility, we designed an MMSET construct with five point mutations in the N-terminal helix of its SET domain for crystallization experiments and elucidated the structure of the mutant MMSET SET domain at 2.1 Å resolution. Both NSD1 and MMSET crystal systems proved resistant to soaking or cocrystallography with inhibitors. However, use of the close homologue SETD2 as a structural surrogate supported the design and characterization of N-alkyl sinefungin derivatives, which showed low micromolar inhibition against both SETD2 and MMSET..
Bavetsias, V.
Lanigan, R.M.
Ruda, G.F.
Atrash, B.
McLaughlin, M.G.
Tumber, A.
Mok, N.Y.
Le Bihan, Y.-.
Dempster, S.
Boxall, K.J.
Jeganathan, F.
Hatch, S.B.
Savitsky, P.
Velupillai, S.
Krojer, T.
England, K.S.
Sejberg, J.
Thai, C.
Donovan, A.
Pal, A.
Scozzafava, G.
Bennett, J.M.
Kawamura, A.
Johansson, C.
Szykowska, A.
Gileadi, C.
Burgess-Brown, N.A.
von Delft, F.
Oppermann, U.
Walters, Z.
Shipley, J.
Raynaud, F.I.
Westaway, S.M.
Prinjha, R.K.
Fedorov, O.
Burke, R.
Schofield, C.J.
Westwood, I.M.
Bountra, C.
Müller, S.
van Montfort, R.L.
Brennan, P.E.
Blagg, J.
(2016). 8-Substituted Pyrido[3,4-d]pyrimidin-4(3H)-one Derivatives As Potent, Cell Permeable, KDM4 (JMJD2) and KDM5 (JARID1) Histone Lysine Demethylase Inhibitors. J med chem,
Vol.59
(4),
pp. 1388-1409.
show abstract
full text
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..
Cheeseman, M.D.
Westwood, I.M.
Barbeau, O.
Rowlands, M.
Dobson, S.
Jones, A.M.
Jeganathan, F.
Burke, R.
Kadi, N.
Workman, P.
Collins, I.
van Montfort, R.L.
Jones, K.
(2016). Exploiting Protein Conformational Change to Optimize Adenosine-Derived Inhibitors of HSP70. J med chem,
Vol.59
(10),
pp. 4625-4636.
show abstract
full text
HSP70 is a molecular chaperone and a key component of the heat-shock response. Because of its proposed importance in oncology, this protein has become a popular target for drug discovery, efforts which have as yet brought little success. This study demonstrates that adenosine-derived HSP70 inhibitors potentially bind to the protein with a novel mechanism of action, the stabilization by desolvation of an intramolecular salt-bridge which induces a conformational change in the protein, leading to high affinity ligands. We also demonstrate that through the application of this mechanism, adenosine-derived HSP70 inhibitors can be optimized in a rational manner..
Innocenti, P.
Woodward, H.L.
Solanki, S.
Naud, S.
Westwood, I.M.
Cronin, N.
Hayes, A.
Roberts, J.
Henley, A.T.
Baker, R.
Faisal, A.
Mak, G.W.
Box, G.
Valenti, M.
De Haven Brandon, A.
O'Fee, L.
Saville, H.
Schmitt, J.
Matijssen, B.
Burke, R.
van Montfort, R.L.
Raynaud, F.I.
Eccles, S.A.
Linardopoulos, S.
Blagg, J.
Hoelder, S.
(2016). Rapid Discovery of Pyrido[3,4-d]pyrimidine Inhibitors of Monopolar Spindle Kinase 1 (MPS1) Using a Structure-Based Hybridization Approach. J med chem,
Vol.59
(8),
pp. 3671-3688.
show abstract
full text
Monopolar spindle 1 (MPS1) plays a central role in the transition of cells from metaphase to anaphase and is one of the main components of the spindle assembly checkpoint. Chromosomally unstable cancer cells rely heavily on MPS1 to cope with the stress arising from abnormal numbers of chromosomes and centrosomes and are thus more sensitive to MPS1 inhibition than normal cells. We report the discovery and optimization of a series of new pyrido[3,4-d]pyrimidine based inhibitors via a structure-based hybridization approach from our previously reported inhibitor CCT251455 and a modestly potent screening hit. Compounds in this novel series display excellent potency and selectivity for MPS1, which translates into biomarker modulation in an in vivo human tumor xenograft model..
Bavetsias, V.
Pérez-Fuertes, Y.
McIntyre, P.J.
Atrash, B.
Kosmopoulou, M.
O'Fee, L.
Burke, R.
Sun, C.
Faisal, A.
Bush, K.
Avery, S.
Henley, A.
Raynaud, F.I.
Linardopoulos, S.
Bayliss, R.
Blagg, J.
(2015). 7-(Pyrazol-4-yl)-3H-imidazo[4,5-b]pyridine-based derivatives for kinase inhibition: Co-crystallisation studies with Aurora-A reveal distinct differences in the orientation of the pyrazole N1-substituent. Bioorg med chem lett,
Vol.25
(19),
pp. 4203-4209.
show abstract
full text
Introduction of a 1-benzyl-1H-pyrazol-4-yl moiety at C7 of the imidazo[4,5-b]pyridine scaffold provided 7a which inhibited a range of kinases including Aurora-A. Modification of the benzyl group in 7a, and subsequent co-crystallisation of the resulting analogues with Aurora-A indicated distinct differences in binding mode dependent upon the pyrazole N-substituent. Compounds 7a and 14d interact with the P-loop whereas 14a and 14b engage with Thr217 in the post-hinge region. These crystallographic insights provide options for the design of compounds interacting with the DFG motif or with Thr217..
Gurden, M.D.
Westwood, I.M.
Faisal, A.
Naud, S.
Cheung, K.-.
McAndrew, C.
Wood, A.
Schmitt, J.
Boxall, K.
Mak, G.
Workman, P.
Burke, R.
Hoelder, S.
Blagg, J.
Van Montfort, R.L.
Linardopoulos, S.
(2015). Naturally Occurring Mutations in the MPS1 Gene Predispose Cells to Kinase Inhibitor Drug Resistance. Cancer res,
Vol.75
(16),
pp. 3340-3354.
show abstract
Acquired resistance to therapy is perhaps the greatest challenge to effective clinical management of cancer. With several inhibitors of the mitotic checkpoint kinase MPS1 in preclinical development, we sought to investigate how resistance against these inhibitors may arise so that mitigation or bypass strategies could be addressed as early as possible. Toward this end, we modeled acquired resistance to the MPS1 inhibitors AZ3146, NMS-P715, and CCT251455, identifying five point mutations in the kinase domain of MPS1 that confer resistance against multiple inhibitors. Structural studies showed how the MPS1 mutants conferred resistance by causing steric hindrance to inhibitor binding. Notably, we show that these mutations occur in nontreated cancer cell lines and primary tumor specimens, and that they also preexist in normal lymphoblast and breast tissues. In a parallel piece of work, we also show that the EGFR p.T790M mutation, the most common mutation conferring resistance to the EGFR inhibitor gefitinib, also preexists in cancer cells and normal tissue. Our results therefore suggest that mutations conferring resistance to targeted therapy occur naturally in normal and malignant cells and these mutations do not arise as a result of the increased mutagenic plasticity of cancer cells..
Joshi, A.
Newbatt, Y.
McAndrew, P.C.
Stubbs, M.
Burke, R.
Richards, M.W.
Bhatia, C.
Caldwell, J.J.
McHardy, T.
Collins, I.
Bayliss, R.
(2015). Molecular mechanisms of human IRE1 activation through dimerization and ligand binding. Oncotarget,
Vol.6
(15),
pp. 13019-13035.
show abstract
full text
IRE1 transduces the unfolded protein response by splicing XBP1 through its C-terminal cytoplasmic kinase-RNase region. IRE1 autophosphorylation is coupled to RNase activity through formation of a back-to-back dimer, although the conservation of the underlying molecular mechanism is not clear from existing structures. We have crystallized human IRE1 in a back-to-back conformation only previously seen for the yeast homologue. In our structure the kinase domain appears primed for catalysis but the RNase domains are disengaged. Structure-function analysis reveals that IRE1 is autoinhibited through a Tyr-down mechanism related to that found in the unrelated Ser/Thr protein kinase Nek7. We have developed a compound that potently inhibits human IRE1 kinase activity while stimulating XBP1 splicing. A crystal structure of the inhibitor bound to IRE1 shows an increased ordering of the kinase activation loop. The structures of hIRE in apo and ligand-bound forms are consistent with a previously proposed model of IRE1 regulation in which formation of a back-to-back dimer coupled to adoption of a kinase-active conformation drive RNase activation. The structures provide opportunities for structure-guided design of IRE1 inhibitors..
Cheeseman, M.D.
Faisal, A.
Rayter, S.
Barbeau, O.R.
Kalusa, A.
Westlake, M.
Burke, R.
Swan, M.
van Montfort, R.
Linardopoulos, S.
Jones, K.
(2014). Targeting the PPM1D phenotype; 2,4-bisarylthiazoles cause highly selective apoptosis in PPM1D amplified cell-lines. Bioorg med chem lett,
Vol.24
(15),
pp. 3469-3474.
show abstract
The metal-dependent phosphatase PPM1D (WIP1) is an important oncogene in cancer, with over-expression of the protein being associated with significantly worse clinical outcomes. In this communication we describe the discovery and optimization of novel 2,4-bisarylthiazoles that phenocopy the knockdown of PPM1D, without inhibiting its phosphatase activity. These compounds cause growth inhibition at nanomolar concentrations, induce apoptosis, activate p53 and display impressive cell-line selectivity. The results demonstrate the potential for targeting phenotypes in drug discovery when tackling challenging targets or unknown mechanisms..
Newbatt, Y.
Hardcastle, A.
McAndrew, P.C.
Strover, J.A.
Mirza, A.
Morgan, G.J.
Burke, R.
Davies, F.E.
Collins, I.
van Montfort, R.L.
(2013). Identification of autophosphorylation inhibitors of the inositol-requiring enzyme 1 alpha (IRE1α) by high-throughput screening using a DELFIA assay. J biomol screen,
Vol.18
(3),
pp. 298-308.
show abstract
Inositol-requiring enzyme 1 alpha (IRE1α) is a transmembrane sensor protein with both kinase and ribonuclease activity, which plays a crucial role in the unfolded protein response (UPR). Protein misfolding in the endoplasmic reticulum (ER) lumen triggers dimerization and subsequent trans-autophosphorylation of IRE1α. This leads to the activation of its endoribonuclease (RNase) domain and splicing of the mRNA of the transcriptional activator XBP1, ultimately generating an active XBP1 (XBP1s) implicated in multiple myeloma survival. Previously, we have identified human IRE1α as a target for the development of kinase inhibitors that could modulate the UPR in human cells, which has particular relevance for multiple myeloma and other secretory malignancies. Here we describe the development and validation of a 384-well high-throughput screening assay using DELFIA technology that is specific for IRE1α autophosphorylation. Using this format, a focused library of 2312 potential kinase inhibitors was screened, and several novel IRE1α kinase inhibitor scaffolds were identified that could potentially be developed toward new therapies to treat multiple myeloma..
Hitchin, J.R.
Blagg, J.
Burke, R.
Burns, S.
Cockerill, M.J.
Fairweather, E.E.
Hutton, C.
Jordan, A.M.
McAndrew, C.
Mirza, A.
Mould, D.
Thomson, G.J.
Waddell, I.
Ogilvie, D.J.
(2013). Development and evaluation of selective, reversible LSD1 inhibitors derived from fragments. Medchemcomm,
Vol.4
(11),
pp. 1513-1522.
Stowell, A.
Hamilton, N.
Hitchin, J.
Blagg, J.
Burke, R.
Burns, S.
Cockerill, M.J.
Fairweather, E.
Hutton, C.
Jordan, A.
Mould, D.
Thomson, G.
Waddell, I.
Ogilvie, D.
(2013). Development and evaluation of selective, reversible LSD1 inhibitors from fragment startpoints. Molecular cancer therapeutics,
Vol.12
(11).
Bavetsias, V.
Faisal, A.
Crumpler, S.
Brown, N.
Kosmopoulou, M.
Joshi, A.
Atrash, B.
Pérez-Fuertes, Y.
Schmitt, J.A.
Boxall, K.J.
Burke, R.
Sun, C.
Avery, S.
Bush, K.
Henley, A.
Raynaud, F.I.
Workman, P.
Bayliss, R.
Linardopoulos, S.
Blagg, J.
(2013). Aurora isoform selectivity: design and synthesis of imidazo[4,5-b]pyridine derivatives as highly selective inhibitors of Aurora-A kinase in cells. J med chem,
Vol.56
(22),
pp. 9122-9135.
show abstract
full text
Aurora-A differs from Aurora-B/C at three positions in the ATP-binding pocket (L215, T217, and R220). Exploiting these differences, crystal structures of ligand-Aurora protein interactions formed the basis of a design principle for imidazo[4,5-b]pyridine-derived Aurora-A-selective inhibitors. Guided by a computational modeling approach, appropriate C7-imidazo[4,5-b]pyridine derivatization led to the discovery of highly selective inhibitors, such as compound 28c, of Aurora-A over Aurora-B. In HCT116 human colon carcinoma cells, 28c and 40f inhibited the Aurora-A L215R and R220K mutants with IC50 values similar to those seen for the Aurora-A wild type. However, the Aurora-A T217E mutant was significantly less sensitive to inhibition by 28c and 40f compared to the Aurora-A wild type, suggesting that the T217 residue plays a critical role in governing the observed isoform selectivity for Aurora-A inhibition. These compounds are useful small-molecule chemical tools to further explore the function of Aurora-A in cells..
Silva-Santisteban, M.C.
Westwood, I.M.
Boxall, K.
Brown, N.
Peacock, S.
McAndrew, C.
Barrie, E.
Richards, M.
Mirza, A.
Oliver, A.W.
Burke, R.
Hoelder, S.
Jones, K.
Aherne, G.W.
Blagg, J.
Collins, I.
Garrett, M.D.
van Montfort, R.L.
(2013). Fragment-based screening maps inhibitor interactions in the ATP-binding site of checkpoint kinase 2. Plos one,
Vol.8
(6),
p. e65689.
show abstract
Checkpoint kinase 2 (CHK2) is an important serine/threonine kinase in the cellular response to DNA damage. A fragment-based screening campaign using a combination of a high-concentration AlphaScreen™ kinase assay and a biophysical thermal shift assay, followed by X-ray crystallography, identified a number of chemically different ligand-efficient CHK2 hinge-binding scaffolds that have not been exploited in known CHK2 inhibitors. In addition, it showed that the use of these orthogonal techniques allowed efficient discrimination between genuine hit matter and false positives from each individual assay technology. Furthermore, the CHK2 crystal structures with a quinoxaline-based fragment and its follow-up compound highlight a hydrophobic area above the hinge region not previously explored in rational CHK2 inhibitor design, but which might be exploited to enhance both potency and selectivity of CHK2 inhibitors..
Naud, S.
Westwood, I.M.
Faisal, A.
Sheldrake, P.
Bavetsias, V.
Atrash, B.
Cheung, K.M.
Liu, M.
Hayes, A.
Schmitt, J.
Wood, A.
Choi, V.
Boxall, K.
Mak, G.
Gurden, M.
Valenti, M.
de Haven Brandon, A.
Henley, A.
Baker, R.
McAndrew, C.
Matijssen, B.
Burke, R.
Hoelder, S.
Eccles, S.A.
Raynaud, F.I.
Linardopoulos, S.
Van Montfort, R.L.
Blagg, J.
(2013). Structure-based design of orally bioavailable 1H-pyrrolo[3,2-c]pyridine inhibitors of the mitotic kinase monopolar spindle 1 (MPS1). J med chem,
Vol.56,
pp. 10045-10065.
full text
Faisal, A.
Naud, S.
Schmitt, J.
Westwood, I.
Hayes, A.
Gurden, M.
Bavetsias, V.
Berry, T.
Mak, G.
Innocenti, P.
Cheung, J.
Sheldrake, P.
Atrash, B.
Sun, C.
Matijssen, B.
Burke, R.
Baker, R.
McAndrew, C.
Rowlands, M.
Workman, P.
Eccles, S.A.
Hoelder, S.
Raynaud, F.I.
vanMontfort, R.
Biagg, J.
Linardopoulos, S.
(2012). Characterisation of CCT251455, a novel, selective and highly potent Mps1 kinase inhibitor. Cancer research,
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Sime, M.
Allan, A.C.
Chapman, P.
Fieldhouse, C.
Giblin, G.M.
Healy, M.P.
Lambert, M.H.
Leesnitzer, L.M.
Lewis, A.
Merrihew, R.V.
Rutter, R.A.
Sasse, R.
Shearer, B.G.
Willson, T.M.
Xu, R.X.
Virley, D.J.
(2011). Discovery of GSK1997132B a novel centrally penetrant benzimidazole PPARγ partial agonist. Bioorg med chem lett,
Vol.21
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pp. 5568-5572.
show abstract
The peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-activated nuclear receptor, thought to play a role in energy metabolism, glucose homeostasis and microglia-mediated neuroinflammation. A novel benzimidazole series of centrally penetrant PPARγ partial agonists has been identified. The optimization of PPARγ activity and in vivo pharmacokinetics leading to the identification of GSK1997132B a potent, metabolically stable and centrally penetrant PPARγ partial agonist, is described..
Rimland, J.
Dunne, A.
Hunjan, S.
Sasse, R.
Uings, I.
Montanari, D.
Caivano, M.
Shah, P.
Standing, D.
Gray, D.
Brown, D.
Cairns, W.
Trump, R.
Smith, P.W.
Bertheleme, N.
D'Alessandro, P.
Gul, S.
Vimal, M.
Smith, D.W.
Watson, S.P.
(2010). The identification a novel, selective, non-steroidal, functional glucocorticoid receptor antagonist. Bioorg med chem lett,
Vol.20
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pp. 2340-2343.
show abstract
The identification of novel, potent, non-steroidal/small molecule functional GR antagonist GSK1564023A selective over PR is described. Associated structure–activity relationships and the process of optimisation of an initial HTS hit are also described.
Hall, A.
Elliott, R.L.
Giblin, G.M.
Hussain, I.
Musgrave, J.
Naylor, A.
Sasse, R.
Smith, B.
(2010). Piperidine-derived gamma-secretase modulators. Bioorg med chem lett,
Vol.20
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pp. 1306-1311.
show abstract
This Letter details the SAR of a novel series of piperidine-derived gamma-secretase modulators. Compound 10h was found to be a potent modulator in vitro, which on further profiling, was found to decrease Abeta42, increase Abeta38 and have no effect on Abeta40 levels. Furthermore, 10h demonstrated excellent pharmacokinetic parameters in the mouse, rat and dog in addition to good CNS penetration in the mouse..
Yates, C.M.
Brown, P.J.
Stewart, E.L.
Patten, C.
Austin, R.J.
Holt, J.A.
Maglich, J.M.
Angell, D.C.
Sasse, R.Z.
Taylor, S.J.
Uings, I.J.
Trump, R.P.
(2010). Structure guided design of 5-arylindazole glucocorticoid receptor agonists and antagonists. J med chem,
Vol.53
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pp. 4531-4544.
show abstract
Glucocorticoid receptor (GR) agonists have been used for more than half a century as the most effective treatment of acute and chronic inflammatory conditions despite serious side effects that accompany their extended use that include glucose intolerance, muscle wasting, skin thinning, and osteoporosis. As a starting point for the identification of GR ligands with an improved therapeutic index, we wished to discover selective nonsteroidal GR agonists and antagonists with simplified structure compared to known GR ligands to serve as starting points for the optimization of dissociated GR modulators. To do so, we selected multiple chemical series by structure guided docking studies and evaluated GR agonist activity. From these efforts we identified 5-arylindazole compounds that showed moderate binding to the glucocorticoid receptor (GR) with clear opportunities for further development. Structure guided optimization was used to design arrays that led to potent GR agonists and antagonists. Several in vitro and in vivo experiments were utilized to demonstrate that GR agonist 23a (GSK9027) had a profile similar to that of a classical steroidal GR agonist..
Barnett, H.A.
Coe, D.M.
Cooper, T.W.
Jack, T.I.
Jones, H.T.
Macdonald, S.J.
McLay, I.M.
Rayner, N.
Sasse, R.Z.
Shipley, T.J.
Skone, P.A.
Somers, G.I.
Taylor, S.
Uings, I.J.
Woolven, J.M.
Weingarten, G.G.
(2009). Aryl aminopyrazole benzamides as oral non-steroidal selective glucocorticoid receptor agonists. Bioorg med chem lett,
Vol.19
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pp. 158-162.
show abstract
Aryl aminopyrazole amides capped with N-alkylbenzamides 13-16 are selective glucocorticoid receptor agonists. 2,6-Disubstituted benzamides have prednisolone-like potency or better in vitro. Good oral exposure was demonstrated in the rat, with compounds with lower lipophilicity, for example N-hydroxyethyl benzamides (e.g., 16e)..
Procopiou, P.A.
Barrett, V.J.
Bevan, N.J.
Biggadike, K.
Butchers, P.R.
Coe, D.M.
Conroy, R.
Edney, D.D.
Field, R.N.
Ford, A.J.
Guntrip, S.B.
Looker, B.E.
McLay, I.M.
Monteith, M.J.
Morrison, V.S.
Mutch, P.J.
Richards, S.A.
Sasse, R.
Smith, C.E.
(2009). Synthesis and structure-activity relationships of long-acting beta2 adrenergic receptor agonists incorporating arylsulfonamide groups. J med chem,
Vol.52
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pp. 2280-2288.
show abstract
A series of saligenin alkoxyalkylphenylsulfonamide beta(2) adrenoceptor agonists were prepared by reacting a protected saligenin oxazolidinone with alkynyloxyalkyl bromides, followed by Sonogashira reaction, hydrogenation, and deprotection. The meta-substituted primary sulfonamide was more potent than the para- and the ortho-analogues. Primary sulfonamides were more potent than the secondary and tertiary analogues. The onset and duration of action in vitro of selected compounds was assessed on isolated superfused guinea pig trachea. Sulfonamide 29b had the best profile of potency, selectivity, onset, and duration of action on both guinea pig trachea and human bronchus. Furthermore, 29b was found to have low oral bioavailability in rat and dog and also to have long duration of action in an in vivo model of bronchodilation. Crystalline salts of 29b were identified that had suitable properties for inhaled administration. A proposed binding mode for 29b to the beta(2)-receptor is presented..
HOLMES, F.E.
SASSE, R.
WATTS, S.
BAINES, A.J.
(1992). EVALUATION OF ENDOGENOUS TUBULIN ACETYL TRANSFERASE-ACTIVITY IN MAMMALIAN BRAIN MICROTUBULE PROTEIN. Molecular biology of the cell,
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Woods, A.
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MacRae, T.H.
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Gull, K.
(1989). Definition of individual components within the cytoskeleton of Trypanosoma brucei by a library of monoclonal antibodies. J cell sci,
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show abstract
The detergent-insoluble T. brucei cytoskeleton consists of several morphologically distinct regions and organelles, many of which are detectable only by electron microscopy. We have produced a set of monoclonal antibodies that define each structural component of this highly ordered cytoskeleton. The monoclonal antibodies were selected by cloning of hybridomas produced from mice injected with complex mixtures of proteins of either the cytoskeleton itself or salt extracts thereof. Four antibodies define particular tubulin isotypes and locate the microtubules of the axoneme and sub-pellicular array; two antibodies recognize the flagellum attachment zone; one recognizes the paraflagellar rod and another the basal bodies. Finally, one antibody defines a detergent-insoluble component of the nucleus. The antigens detected by each monoclonal antibody have been analysed by immunofluorescence microscopy, immunogold electron microscopy and Western blotting..
Sasse, R.
Gull, K.
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(1987). SUBPELLICULAR AND FLAGELLAR MICROTUBULES OF TRYPANOSOMA-BRUCEI-BRUCEI CONTAIN THE SAME ALPHA-TUBULIN ISOFORMS. Journal of cell biology,
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SHERWIN, T.
SCHNEIDER, A.
SASSE, R.
SEEBECK, T.
GULL, K.
(1987). DISTINCT LOCALIZATION AND CELL-CYCLE DEPENDENCE OF COOH TERMINALLY TYROSINOLATED ALPHA-TUBULIN IN THE MICROTUBULES OF TRYPANOSOMA-BRUCEI-BRUCEI. Journal of cell biology,
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GULL, K.
HUSSEY, P.J.
SASSE, R.
SCHNEIDER, A.
SEEBECK, T.
SHERWIN, T.
(1986). TUBULIN ISOTYPES - GENERATION OF DIVERSITY IN CELLS AND MICROTUBULAR ORGANELLES. Journal of cell science,
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