Ferraldeschi, R., Welti, J., Powers, M.V., Yuan, W., Smyth, T., Seed, G., Riisnaes, R., Hedayat, S., Wang, H., Crespo, M., et al.
(2016). Second-Generation HSP90 Inhibitor Onalespib Blocks mRNA Splicing of Androgen Receptor Variant 7 in Prostate Cancer Cells. Cancer research,
Ang, J.E., Pandher, R., Ang, J.C., Asad, Y.J., Henley, A.T., Valenti, M., Box, G., de Haven Brandon, A., Baird, R.D., Friedman, L., et al.
(2016). Plasma Metabolomic Changes following PI3K Inhibition as Pharmacodynamic Biomarkers: Preclinical Discovery to Phase I Trial Evaluation. Molecular cancer therapeutics,
Mallinger, A., Schiemann, K., Rink, C., Stieber, F., Calderini, M., Crumpler, S., Stubbs, M., Adeniji-Popoola, O., Poeschke, O., Busch, M., et al.
(2016). Discovery of Potent, Selective, and Orally Bioavailable Small-Molecule Modulators of the Mediator Complex-Associated Kinases CDK8 and CDK19. Journal of medicinal chemistry,
Cheeseman, M.D., Westwood, I.M., Barbeau, O., Rowlands, M., Dobson, S., Jones, A.M., Jeganathan, F., Burke, R., Kadi, N., Workman, P., et al.
(2016). Exploiting Protein Conformational Change to Optimize Adenosine-Derived Inhibitors of HSP70. J med chem,
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..
Sarker, D., Ang, J.E., Baird, R., Kristeleit, R., Shah, K., Moreno, V., Clarke, P.A., Raynaud, F.I., Levy, G., Ware, J.A., et al.
(2015). First-in-human phase I study of pictilisib (GDC-0941), a potent pan-class I phosphatidylinositol-3-kinase (PI3K) inhibitor, in patients with advanced solid tumors. Clin cancer res,
PURPOSE: This first-in-human dose-escalation trial evaluated the safety, tolerability, maximal-tolerated dose (MTD), dose-limiting toxicities (DLT), pharmacokinetics, pharmacodynamics, and preliminary clinical activity of pictilisib (GDC-0941), an oral, potent, and selective inhibitor of the class I phosphatidylinositol-3-kinases (PI3K). PATIENTS AND METHODS: Sixty patients with solid tumors received pictilisib at 14 dose levels from 15 to 450 mg once-daily, initially on days 1 to 21 every 28 days and later, using continuous dosing for selected dose levels. Pharmacodynamic studies incorporated (18)F-FDG-PET, and assessment of phosphorylated AKT and S6 ribosomal protein in platelet-rich plasma (PRP) and tumor tissue. RESULTS: Pictilisib was well tolerated. The most common toxicities were grade 1-2 nausea, rash, and fatigue, whereas the DLT was grade 3 maculopapular rash (450 mg, 2 of 3 patients; 330 mg, 1 of 7 patients). The pharmacokinetic profile was dose-proportional and supported once-daily dosing. Levels of phosphorylated serine-473 AKT were suppressed >90% in PRP at 3 hours after dose at the MTD and in tumor at pictilisib doses associated with AUC >20 h·μmol/L. Significant increase in plasma insulin and glucose levels, and >25% decrease in (18)F-FDG uptake by PET in 7 of 32 evaluable patients confirmed target modulation. A patient with V600E BRAF-mutant melanoma and another with platinum-refractory epithelial ovarian cancer exhibiting PTEN loss and PIK3CA amplification demonstrated partial response by RECIST and GCIG-CA125 criteria, respectively. CONCLUSION: Pictilisib was safely administered with a dose-proportional pharmacokinetic profile, on-target pharmacodynamic activity at dose levels ≥100 mg and signs of antitumor activity. The recommended phase II dose was continuous dosing at 330 mg once-daily..
Smith, J.R., de Billy, E., Hobbs, S., Powers, M., Prodromou, C., Pearl, L., Clarke, P.A. & Workman, P.
(2015). Restricting direct interaction of CDC37 with HSP90 does not compromise chaperoning of client proteins. Oncogene,
The HSP90 molecular chaperone plays a key role in the maturation, stability and activation of its clients, including many oncogenic proteins. Kinases are a substantial and important subset of clients requiring the key cochaperone CDC37. We sought an improved understanding of protein kinase chaperoning by CDC37 in cancer cells. CDC37 overexpression in human colon cancer cells increased CDK4 protein levels, which was negated upon CDC37 knockdown. Overexpressing CDC37 increased CDK4 protein half-life and enhanced binding of HSP90 to CDK4, consistent with CDC37 promoting kinase loading onto chaperone complexes. Against expectation, expression of C-terminus-truncated CDC37 (ΔC-CDC37) that lacks HSP90 binding capacity did not affect kinase client expression or activity; moreover, as with wild-type CDC37 overexpression, it augmented CDK4-HSP90 complex formation. However, although truncation blocked binding to HSP90 in cells, ΔC-CDC37 also showed diminished client protein binding and was relatively unstable. CDC37 mutants with single and double point mutations at residues M164 and L205 showed greatly reduced binding to HSP90, but retained association with client kinases. Surprisingly, these mutants phenocopied wild-type CDC37 overexpression by increasing CDK4-HSP90 association and CDK4 protein levels in cells. Furthermore, expression of the mutants was sufficient to protect kinase clients CDK4, CDK6, CRAF and ERBB2 from depletion induced by silencing endogenous CDC37, indicating that CDC37's client stabilising function cannot be inactivated by substantially reducing its direct interaction with HSP90. However, CDC37 could not compensate for loss of HSP90 function, showing that CDC37 and HSP90 have their own distinct and non-redundant roles in maintaining kinase clients. Our data substantiate the important function of CDC37 in chaperoning protein kinases. Furthermore, we demonstrate that CDC37 can stabilise kinase clients by a mechanism that is not dependent on a substantial direct interaction between CDC37 and HSP90, but nevertheless requires HSP90 activity. These results have significant implications for therapeutic targeting of CDC37. .
Dale, T., Clarke, P.A., Esdar, C., Waalboer, D., Adeniji-Popoola, O., Ortiz-Ruiz, M.-., Mallinger, A., Samant, R.S., Czodrowski, P., Musil, D., et al.
(2015). A selective chemical probe for exploring the role of CDK8 and CDK19 in human disease. Nature chemical biology,
Gurden, M.D., Westwood, I.M., Faisal, A., Naud, S., Cheung, K.-., McAndrew, C., Wood, A., Schmitt, J., Boxall, K., Mak, G., et al.
(2015). Naturally Occurring Mutations in the MPS1 Gene Predispose Cells to Kinase Inhibitor Drug Resistance. Cancer research,
Mallinger, A., Crumpler, S., Pichowicz, M., Waalboer, D., Stubbs, M., Adeniji-Popoola, O., Wood, B., Smith, E., Thai, C., Henley, A.T., et al.
(2015). Discovery of potent, orally bioavailable, small-molecule inhibitors of WNT signaling from a cell-based pathway screen. J med chem,
WNT signaling is frequently deregulated in malignancy, particularly in colon cancer, and plays a key role in the generation and maintenance of cancer stem cells. We report the discovery and optimization of a 3,4,5-trisubstituted pyridine 9 using a high-throughput cell-based reporter assay of WNT pathway activity. We demonstrate a twisted conformation about the pyridine-piperidine bond of 9 by small-molecule X-ray crystallography. Medicinal chemistry optimization to maintain this twisted conformation, cognisant of physicochemical properties likely to maintain good cell permeability, led to 74 (CCT251545), a potent small-molecule inhibitor of WNT signaling with good oral pharmacokinetics. We demonstrate inhibition of WNT pathway activity in a solid human tumor xenograft model with evidence for tumor growth inhibition following oral dosing. This work provides a successful example of hypothesis-driven medicinal chemistry optimization from a singleton hit against a cell-based pathway assay without knowledge of the biochemical target. .
Mitsopoulos, C., Schierz, A.C., Workman, P. & Al-Lazikani, B.
(2015). Distinctive Behaviors of Druggable Proteins in Cellular Networks. Plos computational biology,
Samant, R.S., Clarke, P.A. & Workman, P.
(2014). E3 ubiquitin ligase Cullin-5 modulates multiple molecular and cellular responses to heat shock protein 90 inhibition in human cancer cells. Proc natl acad sci u s a,
The molecular chaperone heat shock protein 90 (HSP90) is required for the activity and stability of its client proteins. Pharmacologic inhibition of HSP90 leads to the ubiquitin-mediated degradation of clients, particularly activated or mutant oncogenic protein kinases. Client ubiquitination occurs via the action of one or more E3 ubiquitin ligases. We sought to identify the role of Cullin-RING family E3 ubiquitin ligases in the cellular response to HSP90 inhibition. Through a focused siRNA screen of 28 Cullin-RING ligase family members, we found that CUL5 and RBX2 were required for degradation of several HSP90 clients upon treatment of human cancer cells with the clinical HSP90 inhibitor 17-AAG. Surprisingly, silencing Cullin-5 (CUL5) also delayed the earlier loss of HSP90 client protein activity at the same time as delaying cochaperone dissociation from inhibited HSP90-client complexes. Expression of a dominant-negative CUL5 showed that NEDD8 conjugation of CUL5 is required for client degradation but not for loss of client activity or recruitment of clients and HSP90 to CUL5. Silencing CUL5 reduced cellular sensitivity to three distinct HSP90 inhibitors, across four cancer types driven by different protein kinases. Our results reveal the importance of CUL5 in multiple aspects of the cellular response to HSP90 inhibition. .
(2014). Successes for UK cancer partnership. Nature,
Patel, M.N., Halling-Brown, M.D., Tym, J.E., Workman, P. & Al-Lazikani, B.
(2013). Objective assessment of cancer genes for drug discovery. Nat rev drug discov,
Selecting the best targets is a key challenge for drug discovery, and achieving this effectively, efficiently and systematically is particularly important for prioritizing candidates from the sizeable lists of potential therapeutic targets that are now emerging from large-scale multi-omics initiatives, such as those in oncology. Here, we describe an objective, systematic, multifaceted computational assessment of biological and chemical space that can be applied to any human gene set to prioritize targets for therapeutic exploration. We use this approach to evaluate an exemplar set of 479 cancer-associated genes, reveal the tension between biological relevance and chemical tractability, and describe major gaps in available knowledge that could be addressed to aid objective decision-making. We also propose drug repurposing opportunities and identify potentially druggable cancer-associated proteins that have been poorly explored with regard to the discovery of small-molecule modulators, despite their biological relevance..
Polier, S., Samant, R.S., Clarke, P.A., Workman, P., Prodromou, C. & Pearl, L.H.
(2013). ATP-competitive inhibitors block protein kinase recruitment to the Hsp90-Cdc37 system. Nat chem biol,
Protein kinase clients are recruited to the Hsp90 molecular chaperone system via Cdc37, which simultaneously binds Hsp90 and kinases and regulates the Hsp90 chaperone cycle. Pharmacological inhibition of Hsp90 in vivo results in degradation of kinase clients, with a therapeutic effect in dependent tumors. We show here that Cdc37 directly antagonizes ATP binding to client kinases, suggesting a role for the Hsp90-Cdc37 complex in controlling kinase activity. Unexpectedly, we find that Cdc37 binding to protein kinases is itself antagonized by ATP-competitive kinase inhibitors, including vemurafenib and lapatinib. In cancer cells, these inhibitors deprive oncogenic kinases such as B-Raf and ErbB2 of access to the Hsp90-Cdc37 complex, leading to their degradation. Our results suggest that at least part of the efficacy of ATP-competitive inhibitors of Hsp90-dependent kinases in tumor cells may be due to targeted chaperone deprivation..
Bjerke, L., Mackay, A., Nandhabalan, M., Burford, A., Jury, A., Popov, S., Bax, D.A., Carvalho, D., Taylor, K.R., Vinci, M., et al.
(2013). Histone H3 3 Mutations Drive Pediatric Glioblastoma through Upregulation of MYCN. Cancer discovery,
Workman, P. & Al-Lazikani, B.
(2013). Drugging cancer genomes. Nat rev drug discov,
Al-Lazikani, B., Banerji, U. & Workman, P.
(2012). Combinatorial drug therapy for cancer in the post-genomic era. Nat biotechnol,
Over the past decade, whole genome sequencing and other 'omics' technologies have defined pathogenic driver mutations to which tumor cells are addicted. Such addictions, synthetic lethalities and other tumor vulnerabilities have yielded novel targets for a new generation of cancer drugs to treat discrete, genetically defined patient subgroups. This personalized cancer medicine strategy could eventually replace the conventional one-size-fits-all cytotoxic chemotherapy approach. However, the extraordinary intratumor genetic heterogeneity in cancers revealed by deep sequencing explains why de novo and acquired resistance arise with molecularly targeted drugs and cytotoxic chemotherapy, limiting their utility. One solution to the enduring challenge of polygenic cancer drug resistance is rational combinatorial targeted therapy..
Yap, T.A., Walton, M.I., Grimshaw, K.M., Te Poele, R.H., Eve, P.D., Valenti, M.R., de Haven Brandon, A.K., Martins, V., Zetterlund, A., Heaton, S.P., et al.
(2012). AT13148 is a novel, oral multi-AGC kinase inhibitor with potent pharmacodynamic and antitumor activity. Clin cancer res,
PURPOSE: Deregulated phosphatidylinositol 3-kinase pathway signaling through AGC kinases including AKT, p70S6 kinase, PKA, SGK and Rho kinase is a key driver of multiple cancers. The simultaneous inhibition of multiple AGC kinases may increase antitumor activity and minimize clinical resistance compared with a single pathway component. EXPERIMENTAL DESIGN: We investigated the detailed pharmacology and antitumor activity of the novel clinical drug candidate AT13148, an oral ATP-competitive multi-AGC kinase inhibitor. Gene expression microarray studies were undertaken to characterize the molecular mechanisms of action of AT13148. RESULTS: AT13148 caused substantial blockade of AKT, p70S6K, PKA, ROCK, and SGK substrate phosphorylation and induced apoptosis in a concentration and time-dependent manner in cancer cells with clinically relevant genetic defects in vitro and in vivo. Antitumor efficacy in HER2-positive, PIK3CA-mutant BT474 breast, PTEN-deficient PC3 human prostate cancer, and PTEN-deficient MES-SA uterine tumor xenografts was shown. We show for the first time that induction of AKT phosphorylation at serine 473 by AT13148, as reported for other ATP-competitive inhibitors of AKT, is not a therapeutically relevant reactivation step. Gene expression studies showed that AT13148 has a predominant effect on apoptosis genes, whereas the selective AKT inhibitor CCT128930 modulates cell-cycle genes. Induction of upstream regulators including IRS2 and PIK3IP1 as a result of compensatory feedback loops was observed. CONCLUSIONS: The clinical candidate AT13148 is a novel oral multi-AGC kinase inhibitor with potent pharmacodynamic and antitumor activity, which shows a distinct mechanism of action from other AKT inhibitors. AT13148 will now be assessed in a first-in-human phase I trial..
Day, J.E., Sharp, S.Y., Rowlands, M.G., Aherne, W., Hayes, A., Raynaud, F.I., Lewis, W., Roe, S.M., Prodromou, C., Pearl, L.H., et al.
(2011). Targeting the Hsp90 Molecular Chaperone with Novel Macrolactams Synthesis, Structural, Binding, and Cellular Studies. Acs chemical biology,
Rowlands, M., McAndrew, C., Prodromou, C., Pearl, L., Kalusa, A., Jones, K., Workman, P. & Aherne, W.
(2010). Detection of the ATPase activity of the molecular chaperones Hsp90 and Hsp72 using the TranscreenerTM ADP assay kit. J biomol screen,
The molecular chaperone heat shock protein 90 (Hsp90) is required for the correct folding and stability of a number of client proteins that are important for the growth and maintenance of cancer cells. Heat shock protein 72 (Hsp72), a co-chaperone of Hsp90, is also emerging as an attractive cancer drug target. Both proteins bind and hydrolyze adenosine triphosphate (ATP), and ATPase activity is essential for their function. Inhibition of Hsp90 ATPase activity leads to the degradation of client proteins, resulting in cell growth inhibition and apoptosis. Several small-molecule inhibitors of the ATPase activity of Hsp90 have been described and are currently being evaluated clinically for the treatment of cancer. A number of methods for the measurement of ATPase activity have been previously used, but not all of these are ideally suited to screening cascades in drug discovery projects. The authors have evaluated the use of commercial reagents (Transcreener ADP) for the measurement of ATPase activity of both yeast and human Hsp90 (ATP K(m) approximately 500 microM) and human Hsp72 (ATP K(m) ~1 microM). The low ATPase activity of human Hsp90 and its stimulation by the co-chaperone Aha1 was measured with ease using reduced incubation times, generating robust data (Z' = 0.75). The potency of several small-molecule inhibitors of both Hsp90 and Hsp72 was determined using the Transcreener reagents and compared well to that determined using other assay formats..
Gaspar, N., Sharp, S.Y., Eccles, S.A., Gowan, S., Popov, S., Jones, C., Pearson, A., Vassal, G. & Workman, P.
(2010). Mechanistic evaluation of the novel HSP90 inhibitor NVP-AUY922 in adult and pediatric glioblastoma. Mol cancer ther,
The dismal prognosis of glioblastoma (GB) indicates the urgent need for new therapies for these tumors. Heat shock protein 90 (HSP90) inhibitors induce the proteasome-mediated degradation of many oncogenic client proteins involved in all of the hallmark characteristics of cancer. Here, we explored the mechanistic potential of the potent synthetic diarylisoxazole amide resorcinol HSP90 inhibitor, NVP-AUY922, in adult and pediatric GB. In vitro antiproliferative potency (nanomolar range) was seen in both adult and pediatric human GB cell lines with different molecular pathologies. A cytostatic effect was observed in all GB lines; more apoptosis was observed at lower concentrations in the SF188 pediatric GB line and at 144 hours in the slower growing KNS42 pediatric GB line, as compared with the adult GB lines U87MG and SF268. In vitro combination studies with inhibitors of phosphoinositide 3-kinase/mammalian target of rapamycin (PI-103) or mitogen-activated protein/extracellular signal-regulated kinase (ERK) kinase (PD-0325901) supported the hypothesis that sustained inhibition of ERK up to 72 hours and at least temporary inhibition of AKT were necessary to induce apoptosis in GB lines. In athymic mice bearing established s.c U87MG GB xenografts, NVP-AUY922 (50 mg/kg i.p x 3 days) caused the inhibition of ERK1/2 and AKT phosphorylation and induced apoptosis, whereas 17-AAG used at maximum tolerated dose was less effective. NVP-AUY922 antitumor activity with objective tumor regression resulted from antiproliferative, proapoptotic, and antiangiogenic effects, the latter shown by decreased microvessel density and HIF1alpha levels. Our results have established a mechanistic proof of concept for the potential of novel synthetic HSP90 inhibitors in adult and pediatric GB, alone or in combination with phosphoinositide 3-kinase/mammalian target of rapamycin and mitogen-activated protein/ERK kinase inhibitors..
Pacey, S., Gore, M., Chao, D., Banerji, U., Larkin, J., Sarker, S., Owen, K., Asad, Y., Raynaud, F., Walton, M., et al.
(2010). A Phase II trial of 17-allylamino, 17-demethoxygeldanamycin (17-AAG, tanespimycin) in patients with metastatic melanoma. Invest new drugs,
Purpose A Phase II study to screen for anti-melanoma activity of the heat shock protein 90 (HSP90) inhibitor, 17-AAG (17-allylamino-17-demethoxygeldanamycin) was performed. The primary endpoint was the rate of disease stabilisation in patients with progressive, metastatic melanoma treated with 17-AAG. Secondary endpoints were to determine: the toxicity of 17-AAG, the duration of response(s), median survival and further study the pharmacokinetics and pharmacodynamics of 17-AAG. Patients and Methods Patients with metastatic melanoma (progressive disease documented =6 months of entering study) were treated with weekly, intravenous 17-AAG. A Simon one sample two stage minimax design was used. A stable disease rate of >/=25% at 6 months was considered compatible with 17-AAG having activity. Results Fourteen patients (8 male: 6 female) were entered, eleven received 17-AAG (performance status 0 or 1). Median age was 60 (range 29-81) years. The majority (93%) received prior chemotherapy and had stage M1c disease (71%). Toxicity was rarely >/= Grade 2 in severity and commonly included fatigue, headache and gastrointestinal disturbances. One of eleven patients treated with 17-AAG had stable disease for 6 months and median survival for all patients was 173 days. The study was closed prematurely prior to completion of the first stage of recruitment and limited planned pharmacokinetic and pharmacodynamic analyses. Conclusion Some evidence of 17-AAG activity was observed although early study termination meant study endpoints were not reached. Stable disease rates can be incorporated into trials screening for anti-melanoma activity and further study of HSP90 inhibitors in melanoma should be considered..
Grimshaw, K.M., Hunter, L.J., Yap, T.A., Heaton, S.P., Walton, M.I., Woodhead, S.J., Fazal, L., Reule, M., Davies, T.G., Seavers, L.C., et al.
(2010). AT7867 Is a Potent and Oral Inhibitor of AKT and p70 S6 Kinase That Induces Pharmacodynamic Changes and Inhibits Human Tumor Xenograft Growth. Mol cancer ther,
The serine/threonine kinase AKT plays a pivotal role in signal transduction events involved in malignant transformation and chemoresistance and is an attractive target for the development of cancer therapeutics. Fragment-based lead discovery, combined with structure-based drug design, has recently identified AT7867 as a novel and potent inhibitor of both AKT and the downstream kinase p70 S6 kinase (p70S6K) and also of protein kinase A. This ATP-competitive small molecule potently inhibits both AKT and p70S6K activity at the cellular level, as measured by inhibition of GSK3 beta and S6 ribosomal protein phosphorylation, and also causes growth inhibition in a range of human cancer cell lines as a single agent. Induction of apoptosis was detected by multiple methods in tumor cells following AT7867 treatment. Administration of AT7867 (90 mg/kg p.o. or 20 mg/kg i.p.) to athymic mice implanted with the PTEN-deficient U87MG human glioblastoma xenograft model caused inhibition of phosphorylation of downstream substrates of both AKT and p70S6K and induction of apoptosis, confirming the observations made in vitro. These doses of AT7867 also resulted in inhibition of human tumor growth in PTEN-deficient xenograft models. These data suggest that the novel strategy of AKT and p70S6K blockade may have therapeutic value and supports further evaluation of AT7867 as a single-agent anticancer strategy. Mol Cancer Ther; 9(5); 1100-10. (C) 2010 AACR..
Yap, T.A., Sandhu, S.K., Workman, P. & de Bono, J.S.
(2010). Envisioning the future of early anticancer drug development. Nat rev cancer,
The development of novel molecularly targeted cancer therapeutics remains slow and expensive with many late-stage failures. There is an urgent need to accelerate this process by improving early clinical anticancer drug evaluation through modern and rational trial designs that incorporate predictive, pharmacokinetic, pharmacodynamic, pharmacogenomic and intermediate end-point biomarkers. In this article, we discuss current approaches and propose strategies that will potentially maximize benefit to patients and expedite the regulatory approvals of new anticancer drugs..
Hayward, D.G., Newbatt, Y., Pickard, L., Byrne, E., Mao, G.J., Burns, S., Sahota, N.K., Workman, P., Collins, I., Aherne, W., et al.
(2010). Identification by High-Throughput Screening of Viridin Analogs as Biochemical and Cell-Based Inhibitors of the Cell Cycle-Regulated Nek2 Kinase. J biomol screen,
Nek2 is a serine/threonine protein kinase that localizes to the centrosome and is implicated in mitotic regulation. Overexpression of Nek2 induces premature centrosome separation and nuclear defects indicative of mitotic errors, whereas depletion of Nek2 interferes with cell growth. As Nek2 expression is upregulated in a range of cancer cell lines and primary human tumors, inhibitors of Nek2 may have therapeutic value in cancer treatment. The authors used a radiometric proximity assay in a high-throughput screen to identify small-molecule inhibitors of Nek2 kinase activity. The assay was based on the measurement of the radiolabeled phosphorylated product of the kinase reaction brought into contact with the surface of wells of solid scintillant-coated microplates. Seventy nonaggregating hits were identified from approximately 73,000 compounds screened and included a number of toxoflavins and a series of viridin/wortmannin-like compounds. The viridin-like compounds were >70-fold selective for Nek2 over Nek6 and Nek7 and inhibited the growth of human tumor cell lines at concentrations consistent with their biochemical potencies. An automated mechanism-based microscopy assay in which centrosomes were visualized using pericentrin antibodies confirmed that 2 of the viridin inhibitors reduced centrosome separation in a human tumor cell line. The data presented show that pharmacological inhibition of Nek2 kinase results in the expected phenotype of disruption to centrosome function associated with growth inhibition and further supports Nek2 as a target for cancer drug discovery. (Journal of Biomolecular Screening 2010:918-927).
Ewan, K., Pajak, B., Stubbs, M., Todd, H., Barbeau, O., Quevedo, C., Botfield, H., Young, R., Ruddle, R., Samuel, L., et al.
(2010). A Useful Approach to Identify Novel Small-Molecule Inhibitors of Wnt-Dependent Transcription. Cancer res,
The Wnt signaling pathway is frequently deregulated in cancer due to mutations in genes encoding APC, beta-catenin, and axin. To identify small-molecule inhibitors of Wnt signaling as potential therapeutics, a diverse chemical library was screened using a transcription factor reporter cell line in which the activity of the pathway was induced at the level of Disheveled protein. A series of deconvolution studies was used to focus on three compound series that selectively killed cancer cell lines with constitutive Wnt signaling. Activities of the compounds included the ability to induce degradation of beta-catenin that had been stabilized by a glycogen synthase kinase-3 (GSK-3) inhibitor. This screen illustrates a practical approach to identify small-molecule inhibitors of Wnt signaling that can seed the development of agents suitable to treat patients with Wnt-dependent tumors. Cancer Res; 70(14); 5963-73. (C) 2010 AACR..
Bavetsias, V., Large, J.M., Sun, C., Bouloc, N., Kosmopoulou, M., Matteucci, M., Wilsher, N.E., Martins, V., Reynisson, J., Atrash, B., et al.
(2010). Imidazo[4,5-b]pyridine derivatives as inhibitors of Aurora kinases: lead optimization studies toward the identification of an orally bioavailable preclinical development candidate. J med chem,
Lead optimization studies using 7 as the starting point led to a new class of imidazo[4,5-b]pyridine-based inhibitors of Aurora kinases that possessed the 1-benzylpiperazinyl motif at the 7-position, and displayed favorable in vitro properties. Cocrystallization of Aurora-A with 40c (CCT137444) provided a clear understanding into the interactions of this novel class of inhibitors with the Aurora kinases. Subsequent physicochemical property refinement by the incorporation of solubilizing groups led to the identification of 3-((4-(6-bromo-2-(4-(4-methylpiperazin-1-yl)phenyl)-3H-imidazo[4,5-b]pyridin-7-yl)piperazin-1-yl)methyl)-5-methylisoxazole (51, CCT137690) which is a potent inhibitor of Aurora kinases (Aurora-A IC(50) = 0.015 +/- 0.003 muM, Aurora-B IC(50) = 0.025 muM, Aurora-C IC(50) = 0.019 muM). Compound 51 is highly orally bioavailable, and in in vivo efficacy studies it inhibited the growth of SW620 colon carcinoma xenografts following oral administration with no observed toxicities as defined by body weight loss..
Workman, P. & Collins, I.
(2010). Probing the probes: fitness factors for small molecule tools. Chem biol,
Chemical probes for interrogating biological processes are of considerable current interest. Cell permeable small molecule tools have a major role in facilitating the functional annotation of the human genome, understanding both physiological and pathological processes, and validating new molecular targets. To be valuable, chemical tools must satisfy necessary criteria and recent publications have suggested objective guidelines for what makes a useful chemical probe. Although recognizing that such guidelines may be valuable, we caution against overly restrictive rules that may stifle innovation in favor of a "fit-for-purpose" approach. Reviewing the literature and providing examples from the cancer field, we recommend a series of "fitness factors" to be considered when assessing chemical probes. We hope this will encourage innovative chemical biology research while minimizing the generation of poor quality and misleading biological data, thus increasing understanding of the particular biological area, to the benefit of basic research and drug discovery..
Gaspar, N., Sharp, S.Y., Pacey, S., Jones, C., Walton, M., Vassal, G., Eccles, S., Pearson, A. & Workman, P.
(2009). Acquired resistance to 17-allylamino-17-demethoxygeldanamycin (17-AAG, tanespimycin) in glioblastoma cells. Cancer res,
Heat shock protein 90 (HSP90) inhibitors, such as 17-allylamino-17-demethoxygeldanamycin (17-AAG, tanespimycin), which is currently in phase II/phase III clinical trials, are promising new anticancer agents. Here, we explored acquired resistance to HSP90 inhibitors in glioblastoma (GB), a primary brain tumor with poor prognosis. GB cells were exposed continuously to increased 17-AAG concentrations. Four 17-AAG-resistant GB cell lines were generated. High-resistance levels with resistance indices (RI = resistant line IC(50)/parental line IC(50)) of 20 to 137 were obtained rapidly (2-8 weeks). After cessation of 17-AAG exposure, RI decreased and then stabilized. Cross-resistance was found with other ansamycin benzoquinones but not with the structurally unrelated HSP90 inhibitors, radicicol, the purine BIIB021, and the resorcinylic pyrazole/isoxazole amide compounds VER-49009, VER-50589, and NVP-AUY922. An inverse correlation between NAD(P)H/quinone oxidoreductase 1 (NQO1) expression/activity and 17-AAG IC(50) was observed in the resistant lines. The NQO1 inhibitor ES936 abrogated the differential effects of 17-AAG sensitivity between the parental and resistant lines. NQO1 mRNA levels and NQO1 DNA polymorphism analysis indicated different underlying mechanisms: reduced expression and selection of the inactive NQO1*2 polymorphism. Decreased NQO1 expression was also observed in a melanoma line with acquired resistance to 17-AAG. No resistance was generated with VER-50589 and NVP-AUY922. In conclusion, low NQO1 activity is a likely mechanism of acquired resistance to 17-AAG in GB, melanoma, and, possibly, other tumor types. Such resistance can be overcome with novel HSP90 inhibitors..
van Montfort, R.L. & Workman, P.
(2009). Structure-based design of molecular cancer therapeutics. Trends biotechnol,
Structure-based approaches now impact across the whole continuum of drug discovery, from new target selection through the identification of hits to the optimization of lead compounds. Optimal application of structure-based design involves close integration with other discovery technologies, including fragment-based and virtual screening. Here, we illustrate the use of structural information and of structure-based drug design approaches in the discovery of small-molecule inhibitors for cancer drug targets and provide an outlook on the exploitation of structural information in future cancer drug discovery. Examples include high profile protein kinase targets and structurally related PI3 kinases, histone deacetylases, poly(ADP-ribose)polymerase and the molecular chaperone HSP90. Structure-based design approaches have also been successfully applied to the protein-protein interaction targets p53-MDM2 and the Bcl-2 family..
Raynaud, F.I., Eccles, S.A., Patel, S., Alix, S., Box, G., Chuckowree, I., Folkes, A., Gowan, S., De Haven Brandon, A., Di Stefano, F., et al.
(2009). Biological properties of potent inhibitors of class I phosphatidylinositide 3-kinases: from PI-103 through PI-540, PI-620 to the oral agent GDC-0941. Mol cancer ther,
The phosphatidylinositide 3-kinase pathway is frequently deregulated in human cancers and inhibitors offer considerable therapeutic potential. We previously described the promising tricyclic pyridofuropyrimidine lead and chemical tool compound PI-103. We now report the properties of the pharmaceutically optimized bicyclic thienopyrimidine derivatives PI-540 and PI-620 and the resulting clinical development candidate GDC-0941. All four compounds inhibited phosphatidylinositide 3-kinase p110alpha with IC(50) < or = 10 nmol/L. Despite some differences in isoform selectivity, these agents exhibited similar in vitro antiproliferative properties to PI-103 in a panel of human cancer cell lines, with submicromolar potency in PTEN-negative U87MG human glioblastoma cells and comparable phosphatidylinositide 3-kinase pathway modulation. PI-540 and PI-620 exhibited improvements in solubility and metabolism with high tissue distribution in mice. Both compounds gave improved antitumor efficacy over PI-103, following i.p. dosing in U87MG glioblastoma tumor xenografts in athymic mice, with treated/control values of 34% (66% inhibition) and 27% (73% inhibition) for PI-540 (50 mg/kg b.i.d.) and PI-620 (25 mg/kg b.i.d.), respectively. GDC-0941 showed comparable in vitro antitumor activity to PI-103, PI-540, and PI-620 and exhibited 78% oral bioavailability in mice, with tumor exposure above 50% antiproliferative concentrations for >8 hours following 150 mg/kg p.o. and sustained phosphatidylinositide 3-kinase pathway inhibition. These properties led to excellent dose-dependent oral antitumor activity, with daily p.o. dosing at 150 mg/kg achieving 98% and 80% growth inhibition of U87MG glioblastoma and IGROV-1 ovarian cancer xenografts, respectively. Together, these data support the development of GDC-0941 as a potent, orally bioavailable inhibitor of phosphatidylinositide 3-kinase. GDC-0941 has recently entered phase I clinical trials..
Brough, P.A., Barril, X., Borgognoni, J., Chene, P., Davies, N.G., Davis, B., Drysdale, M.J., Dymock, B., Eccles, S.A., Garcia-Echeverria, C., et al.
(2009). Combining hit identification strategies: fragment-based and in silico approaches to orally active 2-aminothieno[2,3-d]pyrimidine inhibitors of the Hsp90 molecular chaperone. J med chem,
Inhibitors of the Hsp90 molecular chaperone are showing considerable promise as potential molecular therapeutic agents for the treatment of cancer. Here we describe novel 2-aminothieno[2,3-d]pyrimidine ATP competitive Hsp90 inhibitors, which were designed by combining structural elements of distinct low affinity hits generated from fragment-based and in silico screening exercises in concert with structural information from X-ray protein crystallography. Examples from this series have high affinity (IC50 = 50-100 nM) for Hsp90 as measured in a fluorescence polarization (FP) competitive binding assay and are active in human cancer cell lines where they inhibit cell proliferation and exhibit a characteristic profile of depletion of oncogenic proteins and concomitant elevation of Hsp72. Several examples (34a, 34d and 34i) caused tumor growth regression at well tolerated doses when administered orally in a human BT474 human breast cancer xenograft model..
Smith, J.R., Clarke, P.A., de Billy, E. & Workman, P.
(2009). Silencing the cochaperone CDC37 destabilizes kinase clients and sensitizes cancer cells to HSP90 inhibitors. Oncogene,
The cochaperone CDC37 promotes the association of HSP90 with the protein kinase subset of client proteins to maintain their stability and signalling functions. HSP90 inhibitors induce depletion of clients, which include several oncogenic kinases. We hypothesized that the targeting of CDC37 using siRNAs would compromise the maturation of these clients and increase the sensitivity of cancer cells to HSP90 inhibitors. Here, we show that silencing of CDC37 in human colon cancer cells diminished the association of kinase clients with HSP90 and reduced levels of the clients ERBB2, CRAF, CDK4 and CDK6, as well as phosphorylated AKT. CDC37 silencing promoted the proteasome-mediated degradation of kinase clients, suggesting a degradation pathway independent from HSP90 binding. Decreased cell signalling through kinase clients was also demonstrated by reduced phosphorylation of downstream substrates and colon cancer cell proliferation was subsequently reduced by the inhibition of the G1/S-phase transition. Furthermore, combining CDC37 silencing with the HSP90 inhibitor 17-AAG induced more extensive and sustained depletion of kinase clients and potentiated cell cycle arrest and apoptosis. These results support an essential role for CDC37 in concert with HSP90 in maintaining oncogenic protein kinase clients and endorse the therapeutic potential of targeting CDC37 in cancer..
Gorsuch, S., Bavetsias, V., Rowlands, M.G., Aherne, G.W., Workman, P., Jarman, M. & McDonald, E.
(2009). Synthesis of isothiazol-3-one derivatives as inhibitors of histone acetyltransferases (HATs). Bioorg med chem,
High-throughput screening led to the identification of isothiazolones 1 and 2 as inhibitors of histone acetyltransferase (HAT) with IC50s of 3 microM and 5 microM, respectively. Analogues of these hit compounds with variations of the N-phenyl group, and with variety of substituents at C-4, C-5 of the thiazolone ring, were prepared and assayed for inhibition of the HAT enzyme PCAF. Potency is modestly favoured when the N-aryl group is electron deficient (4-pyridyl derivative 10 has IC(50)=1.5 microM); alkyl substitution at C-4 has little effect, whilst similar substitution at C-5 causes a significant drop in potency. The ring-fused compound 38 has activity (IC(50)=6.1 microM) to encourage further exploration of this bicyclic structure. The foregoing SAR is consistent with an inhibitory mechanism involving cleavage of the S-N bond of the isothiazolone ring by a catalytically important thiol residue..
(2009). Drugging the cancer genome. Drug discovery today,
Banerji, U., Affolter, A., Judson, I., Marais, R. & Workman, P.
(2008). BRAF and NRAS mutations in melanoma: potential relationships to clinical response to HSP90 inhibitors. Mol cancer ther,
Oncogenic BRAF and NRAS mutations are frequent in malignant melanoma. BRAF that is activated by the common V600E and other mutations, as well as by upstream NRAS mutations, has been shown to require the molecular chaperone heat shock protein 90 (HSP90) for stabilization and is depleted by the HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG)]. Here, we explore the possible relationship between tumor BRAF and NRAS mutations and clinical response to 17-AAG in six patients with metastatic malignant melanoma who received pharmacologically active doses of 17-AAG as part of a phase I clinical trial. One patient with disease stabilization for 49 months had a (G13D)NRAS mutation and (WT)BRAF. A second patient who had stable disease for 15 months had a (V600E)BRAF mutation and (WT)NRAS. These preliminary results suggest that BRAF and NRAS mutation status should be determined in prospective phase II studies of HSP90 inhibitors in melanoma..
Chung, Y.L., Troy, H., Kristeleit, R., Aherne, W., Jackson, E., Atadja, P., Griffiths, J.R., Judson, I.R., Workman, P., Leach, M.O., et al.
(2008). Noninvasive magnetic resonance spectroscopic pharmacodynamic markers of a novel histone deacetylase inhibitor, LAQ824, in human colon carcinoma cells and xenografts. Neoplasia,
The aim of this work was to use phosphorus magnetic resonance spectroscopy (P-31 MRS) to investigate the pharmacodynamic effects of LAQ824, a histone deacetylase (HDAC) inhibitor. Human HT29 colon carcinoma cells were examined by 31P MRS after treatment with LAQ824 and another HDAC inhibitor, suberoylanilide hydroxamic acid. HT29 xenografts and tumor extracts were also examined using P-31 MRS, pre- and post-LAQ824 treatment. Histone H3 acetylation was determined using Western blot analysis, and tumor microvessel density by immunohistochemical staining of CD31. Phosphocholine showed a significant increase in HT29 cells after treatment with LAQ824 and suberoylanilide hydroxamic acid. In vivo, the ratio of phosphomonoester/total phosphorus (TotP) signal was significantly increased in LAQ824-treated HT29 xenografts, and this ratio was inversely correlated with changes in tumor volume. Statistically significant decreases in intracellular pH, beta-nucleoside triphosphate (beta-NTP)/TotP, and beta-NTP/inorganic phosphate (Pi) and an increase in Pi/TotP were also seen in LAQ824-treated tumors. Tumor extracts showed many significant metabolic changes after LAQ824 treatment, in parallel with increased histone acetylation and decreased microvessel density. Treatment with LAQ824 resulted in altered phospholipid metabolism and compromised tumor bioenergetics. The phosphocholine and phosphomonoester increases may have the potential to act as pharmacodynamic markers for noninvasively monitoring tumor response after treatment with LAQ824 or other HDAC inhibitors..
Powers, M.V., Clarke, P.A. & Workman, P.
(2008). Dual targeting of HSC70 and HSP72 inhibits HSP90 function and induces tumor-specific apoptosis. Cancer cell,
Heat-shock protein 70 (HSP70) isoforms contribute to tumorigenesis through their well-documented antiapoptotic activity and via their role as cochaperones for the HSP90 molecular chaperone. HSP70 expression is induced following treatment with HSP90 inhibitors, which may attenuate the cell death effects of this class of inhibitor. Here we show that silencing either heat-shock cognate 70 (HSC70) or HSP72 expression in human cancer cell lines has no effect on HSP90 activity or cell proliferation. However, simultaneously reducing the expression of both of these isoforms induces proteasome-dependent degradation of HSP90 client proteins, G1 cell-cycle arrest, and extensive tumor-specific apoptosis. Importantly, simultaneous silencing of HSP70 isoforms in nontumorigenic cell lines does not result in comparable growth arrest or induction of apoptosis, indicating a potential therapeutic window..
Caldwell, J.J., Davies, T.G., Donald, A., McHardy, T., Rowlands, M.G., Aherne, G.W., Hunter, L.K., Taylor, K., Ruddle, R., Raynaud, F.I., et al.
(2008). Identification of 4-(4-aminopiperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidines as selective inhibitors of protein kinase B through fragment elaboration. J med chem,
Fragment-based screening identified 7-azaindole as a protein kinase B inhibitor scaffold. Fragment elaboration using iterative crystallography of inhibitor-PKA-PKB chimera complexes efficiently guided improvements in the potency and selectivity of the compounds, resulting in the identification of nanomolar 6-(piperidin-1-yl)purine, 4-(piperidin-1-yl)-7-azaindole, and 4-(piperidin-1-yl)pyrrolo[2,3- d]pyrimidine inhibitors of PKBbeta with antiproliferative activity and showing pathway inhibition in cells. A divergence in the binding mode was seen between 4-aminomethylpiperidine and 4-aminopiperidine containing molecules. Selectivity for PKB vs PKA was observed with 4-aminopiperidine derivatives, and the most PKB-selective inhibitor (30-fold) showed significantly different bound conformations between PKA and PKA-PKB chimera..
Rayter, S., Elliott, R., Travers, J., Rowlands, M.G., Richardson, T.B., Boxall, K., Jones, K., Linardopoulos, S., Workman, P., Aherne, W., et al.
(2008). A chemical inhibitor of PPM1D that selectively kills cells overexpressing PPM1D. Oncogene,
The PPM1D gene is aberrantly amplified in a range of common cancers and encodes a protein phosphatase that is a potential therapeutic target. However, the issue of whether inhibition of PPM1D in human tumour cells that overexpress this protein compromises their viability has not yet been fully addressed. We show here, using an RNA interference (RNAi) approach, that inhibition of PPM1D can indeed reduce the viability of human tumour cells and that this effect is selective; tumour cell lines that overexpress PPM1D are sensitive to PPM1D inhibition whereas cell lines with normal levels are not. Loss of viability associated with PPM1D RNAi in human tumour cells occurs via the activation of the kinase P38. To identify chemical inhibitors of PPM1D, a high-throughput screening of a library of small molecules was performed. This strategy successfully identified a compound that selectively reduces viability of human tumour cell lines that overexpress PPM1D. As expected of a specific inhibitor, the toxicity to PPM1D overexpressing cell lines after inhibitor treatment is P38 dependent. These results further validate PPM1D as a therapeutic target and identify a proof-of-principle small molecule inhibitor..
Holmes, J.L., Sharp, S.Y., Hobbs, S. & Workman, P.
(2008). Silencing of HSP90 cochaperone AHA1 expression decreases client protein activation and increases cellular sensitivity to the HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin. Cancer res,
AHA1 (activator of HSP90 ATPase) is a cochaperone of the ATP-dependent molecular chaperone, HSP90, which is involved in the maturation, stabilization/degradation, and function of oncogenic proteins. HSP90 operates in a multimeric complex driven by the binding and hydrolysis of ATP. Treatment of cells with the HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) results in the degradation of client proteins via the ubiquitin-proteasome pathway. As AHA1 increases the ATPase activity of HSP90, we hypothesized that modulation of AHA1 expression could influence the activity of client proteins and/or the cellular response to 17-AAG. We show that the basal expression of AHA1 is different across a panel of human cancer cell lines, and that treatment with 17-AAG resulted in sustained AHA1 up-regulation. Increasing the expression of AHA1 did not affect the sensitivity to 17-AAG, but did increase C-RAF activity and the levels of phosphorylated MEK1/2 and ERK1/2 without affecting total levels of these proteins or of client proteins C-RAF, ERBB2, or CDK4. Conversely, small interfering RNA-selective knockdown of >80% of AHA1 expression decreased C-RAF activity and reduced the levels of MEK1/2 and ERK1/2 phosphorylation. Moreover, the AHA1 knockdown resulted in a significant (P < 0.05) increase in sensitivity to 17-AAG, due in part to a 2- to 3-fold increase in apoptosis. These results show that the reduction of AHA1 levels could decrease the phosphorylation of key signal transduction proteins, and for the first time, separate the activation and stabilization functions of HSP90. Furthermore, AHA1 knockdown could sensitize cancer cells to 17-AAG. We conclude that modulation of AHA1 might be a potential therapeutic strategy to increase sensitivity to HSP90 inhibitors..
Eccles, S.A., Massey, A., Raynaud, F.I., Sharp, S.Y., Box, G., Valenti, M., Patterson, L., de Haven Brandon, A., Gowan, S., Boxall, F., et al.
(2008). NVP-AUY922: a novel heat shock protein 90 inhibitor active against xenograft tumor growth, angiogenesis, and metastasis. Cancer res,
We describe the biological properties of NVP-AUY922, a novel resorcinylic isoxazole amide heat shock protein 90 (HSP90) inhibitor. NVP-AUY922 potently inhibits HSP90 (K(d) = 1.7 nmol/L) and proliferation of human tumor cells with GI(50) values of approximately 2 to 40 nmol/L, inducing G(1)-G(2) arrest and apoptosis. Activity is independent of NQO1/DT-diaphorase, maintained in drug-resistant cells and under hypoxic conditions. The molecular signature of HSP90 inhibition, comprising induced HSP72 and depleted client proteins, was readily demonstrable. NVP-AUY922 was glucuronidated less than previously described isoxazoles, yielding higher drug levels in human cancer cells and xenografts. Daily dosing of NVP-AUY922 (50 mg/kg i.p. or i.v.) to athymic mice generated peak tumor levels at least 100-fold above cellular GI(50). This produced statistically significant growth inhibition and/or regressions in human tumor xenografts with diverse oncogenic profiles: BT474 breast tumor treated/control, 21%; A2780 ovarian, 11%; U87MG glioblastoma, 7%; PC3 prostate, 37%; and WM266.4 melanoma, 31%. Therapeutic effects were concordant with changes in pharmacodynamic markers, including induction of HSP72 and depletion of ERBB2, CRAF, cyclin-dependent kinase 4, phospho-AKT/total AKT, and hypoxia-inducible factor-1alpha, determined by Western blot, electrochemiluminescent immunoassay, or immunohistochemistry. NVP-AUY922 also significantly inhibited tumor cell chemotaxis/invasion in vitro, WM266.4 melanoma lung metastases, and lymphatic metastases from orthotopically implanted PC3LN3 prostate carcinoma. NVP-AUY922 inhibited proliferation, chemomigration, and tubular differentiation of human endothelial cells and antiangiogenic activity was reflected in reduced microvessel density in tumor xenografts. Collectively, the data show that NVP-AUY922 is a potent, novel inhibitor of HSP90, acting via several processes (cytostasis, apoptosis, invasion, and angiogenesis) to inhibit tumor growth and metastasis. NVP-AUY922 has entered phase I clinical trials..
de Bono, J.S., Kristeleit, R., Tolcher, A., Fong, P., Pacey, S., Karavasilis, V., Mita, M., Shaw, H., Workman, P., Kaye, S., et al.
(2008). Phase I pharmacokinetic and pharmacodynamic study of LAQ824, a hydroxamate histone deacetylase inhibitor with a heat shock protein-90 inhibitory profile, in patients with advanced solid tumors. Clin cancer res,
To determine the safety, maximum tolerated dose, and pharmacokinetic-pharmacodynamic profile of a histone deacetylase inhibitor, LAQ824, in patients with advanced malignancy. Patients and Methods: LAQ824 was administered i.v. as a 3-h infusion on days 1, 2, and 3 every 21 days. Western blot assays of peripheral blood mononuclear cell lysates and tumor biopsies pretherapy and posttherapy evaluated target inhibition and effects on heat shock protein-90 (HSP90) client proteins and HSP72..
Brough, P.A., Aherne, W., Barril, X., Borgognoni, J., Boxall, K., Cansfield, J.E., Cheung, K.-., Collins, I., Davies, N.G., Drysdale, M.J., et al.
(2008). 4,5-diarylisoxazole Hsp90 chaperone inhibitors: potential therapeutic agents for the treatment of cancer. J med chem,
Inhibitors of the Hsp90 molecular chaperone are showing considerable promise as potential chemotherapeutic agents for cancer. Here, we describe the structure-based design, synthesis, structure-activity relationships and pharmacokinetics of potent small-molecule inhibitors of Hsp90 based on the 4,5-diarylisoxazole scaffold. Analogues from this series have high affinity for Hsp90, as measured in a fluorescence polarization (FP) competitive binding assay, and are active in cancer cell lines where they inhibit proliferation and exhibit a characteristic profile of depletion of oncogenic proteins and concomitant elevation of Hsp72. Compound 40f (VER-52296/NVP-AUY922) is potent in the Hsp90 FP binding assay (IC50 = 21 nM) and inhibits proliferation of various human cancer cell lines in vitro, with GI50 averaging 9 nM. Compound 40f is retained in tumors in vivo when administered i.p., as evaluated by cassette dosing in tumor-bearing mice. In a human colon cancer xenograft model, 40f inhibits tumor growth by approximately 50%..
Vaughan, C.K., Mollapour, M., Smith, J.R., Truman, A., Hu, B., Good, V.M., Panaretou, B., Neckers, L., Clarke, P.A., Workman, P., et al.
(2008). Hsp90-dependent activation of protein kinases is regulated by chaperone-targeted dephosphorylation of Cdc37. Mol cell,
Activation of protein kinase clients by the Hsp90 system is mediated by the cochaperone protein Cdc37. Cdc37 requires phosphorylation at Ser13, but little is known about the regulation of this essential posttranslational modification. We show that Ser13 of uncomplexed Cdc37 is phosphorylated in vivo, as well as in binary complex with a kinase (C-K), or in ternary complex with Hsp90 and kinase (H-C-K). Whereas pSer13-Cdc37 in the H-C-K complex is resistant to nonspecific phosphatases, it is efficiently dephosphorylated by the chaperone-targeted protein phosphatase 5 (PP5/Ppt1), which does not affect isolated Cdc37. We show that Cdc37 and PP5/Ppt1 associate in Hsp90 complexes in yeast and in human tumor cells, and that PP5/Ppt1 regulates phosphorylation of Ser13-Cdc37 in vivo, directly affecting activation of protein kinase clients by Hsp90-Cdc37. These data reveal a cyclic regulatory mechanism for Cdc37, in which its constitutive phosphorylation is reversed by targeted dephosphorylation in Hsp90 complexes..
Maloney, A., Clarke, P.A., Naaby-Hansen, S., Stein, R., Koopman, J.O., Akpan, A., Yang, A., Zvelebil, M., Cramer, R., Stimson, L., et al.
(2007). Gene and protein expression profiling of human ovarian cancer cells treated with the heat shock protein 90 inhibitor 17-allylamino-17-demethoxygeldanamycin. Cancer res,
The promising antitumor activity of 17-allylamino-17-demethoxygeldanamycin (17AAG) results from inhibition of the molecular chaperone heat shock protein 90 (HSP90) and subsequent degradation of multiple oncogenic client proteins. Gene expression microarray and proteomic analysis were used to profile molecular changes in the A2780 human ovarian cancer cell line treated with 17AAG. Comparison of results with an inactive analogue and an alternative HSP90 inhibitor radicicol indicated that increased expression of HSP72, HSC70, HSP27, HSP47, and HSP90beta at the mRNA level were on-target effects of 17AAG. HSP27 protein levels were increased in tumor biopsies following treatment of patients with 17AAG. A group of MYC-regulated mRNAs was decreased by 17AAG. Of particular interest and novelty were changes in expression of chromatin-associated proteins. Expression of the heterochromatin protein 1 was increased, and expression of the histone acetyltransferase 1 and the histone arginine methyltransferase PRMT5 was decreased by 17AAG. PRMT5 was shown to be a novel HSP90-binding partner and potential client protein. Cellular protein acetylation was reduced by 17AAG, which was shown to have an antagonistic interaction on cell proliferation with the histone deacetylase inhibitor trichostatin A. This mRNA and protein expression analysis has provided new insights into the complex molecular pharmacology of 17AAG and suggested new genes and proteins that may be involved in response to the drug or be potential biomarkers of drug action..
Smith, N.F., Raynaud, F.I. & Workman, P.
(2007). The application of cassette dosing for pharmacokinetic screening in small-molecule cancer drug discovery. Mol cancer ther,
Pharmacokinetic evaluation is an essential component of drug discovery and should be conducted early in the process so that those compounds with the best chance of success are prioritized and progressed. However, pharmacokinetic analysis has become a serious bottleneck during the 'hit-to-lead' and lead optimization phases due to the availability of new targets and the large numbers of compounds resulting from advances in synthesis and screening technologies. Cassette dosing, which involves the simultaneous administration of several compounds to a single animal followed by rapid sample analysis by liquid chromatography/tandem mass spectrometry, was developed to increase the throughput of in vivo pharmacokinetic screening. Although cassette dosing is advantageous in terms of resources and throughput, there are possible complications associated with this approach, such as the potential for compound interactions. Following an overview of the cassette dosing literature, this article focuses on the application of the technique in anticancer drug discovery. Specific examples are discussed, including the evaluation of cassette dosing to assess pharmacokinetic properties in the development of cyclin-dependent kinase and heat shock protein 90 inhibitors. Subject to critical analysis and validation in each case, the use of cassette dosing is recommended in appropriate chemical series to enhance the efficiency of drug discovery and reduce animal usage..
Hardcastle, A., Tomlin, P., Norris, C., Richards, J., Cordwell, M., Boxall, K., Rowlands, M., Jones, K., Collins, I., McDonald, E., et al.
(2007). A duplexed phenotypic screen for the simultaneous detection of inhibitors of the molecular chaperone heat shock protein 90 and modulators of cellular acetylation. Mol cancer ther,
Histone deacetylases (HDACs), histone acetyltransferases (HATs), and the molecular chaperone heat shock protein 90 (HSP90) are attractive anticancer drug targets. High-throughput screening plays a pivotal role in modern molecular mechanism-based drug discovery. Cell-based screens are particularly useful in that they identify compounds that are permeable and active against the selected target or pathway in a cellular context. We have previously developed time-resolved fluorescence cell immunosorbent assays (TRF-Cellisas) for compound screening and pharmacodynamic studies. These assays use a primary antibody to the single protein of interest and a matched secondary immunoglobulin labeled with an europium chelate (Eu). The availability of species-specific secondary antibodies labeled with different lanthanide chelates provides the potential for multiplexing this type of assay. The approach has been applied to the development of a 384-well duplexed cell-based screen to simultaneously detect compounds that induce the co-chaperone HSP70 as a molecular marker of potential inhibitors of HSP90 together with those that modulate cellular acetylation (i.e., potential inhibitors of histone deacetylase or histone acetyltransferase activity). The duplexed assay proved reliable in high-throughput format and approximately 64,000 compounds were screened. Following evaluation in secondary assays, 3 of 13 hits from the HSP70 arm were confirmed. Two of these directly inhibited the intrinsic ATPase activity of HSP90 whereas the third seems to have a different mechanism of action. In the acetylation arm, two compounds increased cellular acetylation, one of which inhibited histone deacetylase activity. A third compound decreased cellular histone acetylation, potentially through a novel mechanism of action..
Chan, F., Sun, C., Perumal, M., Nguyen, Q.D., Bavetsias, V., McDonald, E., Martins, V., Wilsher, N.E., Raynaud, F.I., Valenti, M., et al.
(2007). Mechanism of action of the Aurora kinase inhibitor CCT129202 and in vivo quantification of biological activity. Mol cancer ther,
(12 Pt 1),
The Aurora family of serine/threonine kinases is important for the regulation of centrosome maturation, chromosome segregation, and cytokinesis during mitosis. Overexpression of Aurora kinases in mammalian cells leads to genetic instability and transformation. Increased levels of Aurora kinases have also been linked to a broad range of human tumors. Here, we describe the properties of CCT129202, a representative of a structurally novel series of imidazopyridine small-molecule inhibitors of Aurora kinase activity. This compound showed high selectivity for the Aurora kinases over a panel of other kinases tested and inhibits proliferation in multiple cultured human tumor cell lines. CCT129202 causes the accumulation of human tumor cells with >or=4N DNA content, leading to apoptosis. CCT120202-treated human tumor cells showed a delay in mitosis, abrogation of nocodazole-induced mitotic arrest, and spindle defects. Growth of HCT116 xenografts in nude mice was inhibited after i.p. administration of CCT129202. We show that p21, the cyclin-dependent kinase inhibitor, is induced by CCT129202. Up-regulation of p21 by CCT129202 in HCT116 cells led to Rb hypophosphorylation and E2F inhibition, contributing to a decrease in thymidine kinase 1 transcription. This has facilitated the use of 3'-deoxy-3'[(18)F]fluorothymidine-positron emission tomography to measure noninvasively the biological activity of the Aurora kinase inhibitor CCT129202 in vivo..
Raynaud, F.I., Eccles, S., Clarke, P.A., Hayes, A., Nutley, B., Alix, S., Henley, A., Di-Stefano, F., Ahmad, Z., Guillard, S., et al.
(2007). Pharmacologic characterization of a potent inhibitor of class I phosphatidylinositide 3-kinases. Cancer res,
Extensive evidence implicates activation of the lipid phosphatidylinositide 3-kinase (PI3K) pathway in the genesis and progression of various human cancers. PI3K inhibitors thus have considerable potential as molecular cancer therapeutics. Here, we detail the pharmacologic properties of a prototype of a new series of inhibitors of class I PI3K. PI103 is a potent inhibitor with low IC50 values against recombinant PI3K isoforms p110alpha (2 nmol/L), p110beta (3 nmol/L), p110delta (3 nmol/L), and p110gamma (15 nmol/L). PI103 also inhibited TORC1 by 83.9% at 0.5 micromol/L and exhibited an IC50 of 14 nmol/L against DNA-PK. A high degree of selectivity for the PI3K family was shown by the lack of activity of PI103 in a panel of 70 protein kinases. PI103 potently inhibited proliferation and invasion of a wide variety of human cancer cells in vitro and showed biomarker modulation consistent with inhibition of PI3K signaling. PI103 was extensively metabolized, but distributed rapidly to tissues and tumors. This resulted in tumor growth delay in eight different human cancer xenograft models with various PI3K pathway abnormalities. Decreased phosphorylation of AKT was observed in U87MG gliomas, consistent with drug levels achieved. We also showed inhibition of invasion in orthotopic breast and ovarian cancer xenograft models and obtained evidence that PI103 has antiangiogenic potential. Despite its rapid in vivo metabolism, PI103 is a valuable tool compound for exploring the biological function of class I PI3K and importantly represents a lead for further optimization of this novel class of targeted molecular cancer therapeutic..
Whittaker, S.R., Te Poele, R.H., Chan, F., Linardopoulos, S., Walton, M.I., Garrett, M.D. & Workman, P.
(2007). The cyclin-dependent kinase inhibitor seliciclib (R-roscovitine; CYC202) decreases the expression of mitotic control genes and prevents entry into mitosis. Cell cycle,
The cyclin-dependent kinase (CDK) inhibitor seliciclib (R-roscovitine, CYC202) shows promising antitumor activity in preclinical models and is currently undergoing phase II clinical trials. Inhibition of the CDKs by seliciclib could contribute to cell cycle arrest and apoptosis seen with the drug. However, it is common for drugs to exert multiple effects on gene expression and biochemical pathways. To further our understanding of the molecular pharmacology of seliciclib, we employed cDNA microarrays to determine changes in gene expression profiles induced by the drug in HT29 human colon cancer cells. Concentrations of seliciclib were used that inhibited RB phosphorylation and cell proliferation. An increase in the mRNA expression for CJUN and EGR1 was confirmed by Western blotting, consistent with activation of the ERK1/2 MAPK pathway by seliciclib. Transcripts of key genes required for the progression through mitosis showed markedly reduced expression, including Aurora-A/B (AURK-A/B), Polo-like kinase (PLK), cyclin B2 (CCNB2), WEE1 and CDC25C. Reduced expression of these mitotic genes was also seen at the protein level. siRNA-mediated depletion of Aurora-A protein led to an arrest of cells in the G(2)/M phase, consistent with the effects of seliciclib treatment. Inhibition of mitotic entry following seliciclib treatment was indicated by a reduction of histone H3 phosphorylation, which is catalyzed by Aurora-B, and by decreased expression of mitotic markers, including phospho-protein phosphatase 1 alpha. The results indicate a potential mechanism through which seliciclib prevents entry into mitosis. Gene expression profiling has generated hypotheses that led to an increase in our knowledge of the cellular effects of seliciclib and could provide potential pharmacodynamic or response biomarkers for use in animal models and clinical trials..
Sanderson, S., Valenti, M., Gowan, S., Patterson, L., Ahmad, Z., Workman, P. & Eccles, S.A.
(2006). Benzoquinone ansamycin heat shock protein 90 inhibitors modulate multiple functions required for tumor angiogenesis. Mol cancer ther,
Heat shock protein 90 (Hsp90) is a molecular chaperone involved in maintaining the correct conformation and stability of its client proteins. This study investigated the effects of Hsp90 inhibitors on client protein expression and key cellular functions required for tumor angiogenesis. The benzoquinone ansamycin Hsp90 inhibitors geldanamycin and/or its derivatives 17-allylamino-17-demethoxygeldanamycin (17-AAG) and 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin inhibited production of vascular endothelial growth factor (VEGF)-A by tumor cells and blocked proliferative responses of human endothelial cells at nanomolar concentrations. 17-AAG also significantly reduced endothelial cell migration, tubular differentiation, invasion through Matrigel, and secretion of urokinase-type plasminogen activator at concentrations at or below those that inhibited proliferation. 17-AAG significantly reduced expression of VEGF receptor (VEGFR)-2 and established Hsp90 client proteins in human endothelial cells in vitro as well as in mouse vena cava, mesenteric vessels, and blood vessels within human tumor xenografts in vivo; this was associated with decreased tumor microvessel density. Finally, we showed for the first time that Hsp90 inhibitors also reduce expression of VEGFR-1 on human vascular endothelial cells, VEGFR-3 on lymphatic endothelial cells in vitro, and all three VEGFRs on mouse vasculature in vivo. Thus, we identify Hsp90 inhibitors as important regulators of many aspects of tumor angiogenesis (and potentially lymphangiogenesis) and suggest that they may provide therapeutic benefit not only via direct effects on tumor cells but also indirectly by inhibiting the production of angiogenic cytokines and responses of activated endothelial cells that contribute to tumor progression and metastasis..
Newbatt, Y., Burns, S., Hayward, R., Whittaker, S., Kirk, R., Marshall, C., Springer, C., McDonald, E., Marais, R., Workman, P., et al.
(2006). Identification of inhibitors of the kinase activity of oncogenic (V600E)BRAF in an enzyme cascade high-throughput screen. J biomol screen,
The Cancer Genome Project has identified several oncogenic mutations in BRAT that represent important opportunities for cancer drug discovery. The (V600E)BRAF mutation accounts for approximately 90% of the mutations identified. A strong case has emerged from molecular, cellular, and structural studies for the identification and development of inhibitors of this mutated BRAF protein. The authors have developed and run a high-throughput screen to find inhibitors of (V600E)BRAF using an enzyme cascade assay in which oncogenic BRAF activates MEK1, which in turn activates ERK2, which then phosphorylates the transcription factor ELK1. A phosphospecific antibody, Europium-labeled secondary antibody, and a time-resolved fluorescent readout were used to measure phosphorylation of ELK1. Overall assay variation was 12.4%. The assay was used to screen 64,000 compounds with an overall Z' factor of 0.58 +/- 0.12. A series of 3,5,di-substituted pyridines were identified as inhibitors of the cascade assay. These compounds did not inhibit a shortened activated MEK1 to ELK1 cascade but were active (0.5-27.9 mu M) in a (V600E)BRAF assay and represent a potential starting point for future drug discovery and development..
Smith, N.F., Hayes, A., James, K., Nutley, B.P., McDonald, E., Henley, A., Dymock, B., Drysdale, M.J., Raynaud, F.I. & Workman, P., et al.
(2006). Preclinical pharmacokinetics and metabolism of a novel diaryl pyrazole resorcinol series of heat shock protein 90 inhibitors. Mol cancer ther,
CCT018159 was recently identified as a novel inhibitor of heat shock protein (Hsp) 90, a promising target for cancer therapy. Pharmacokinetic and metabolic properties are likely to be important for efficacy and need to be optimized during drug development. Here, we define the preclinical metabolism and pharmacokinetics of CCT018159 and some early derivatives. In addition, we assess in vitro metabolic stability screening and in vivo cassette dosing (simultaneous administration of several compounds to a single animal) as approaches to investigate these compounds. The plasma clearance following individual i.v. administration to mice was rapid (0.128-0.816 L/h), exceeding hepatic blood flow. For CCT066950 and CCT066952, this could be attributed in part to extensive (>80%) blood cell binding. Oral bioavailability ranged from 1.8% to 29.6%. Tissue distribution of CCT066952 was rapid and moderate, and renal excretion of the compounds was minimal (<1% of dose excreted). Compounds underwent rapid glucuronidation both in vivo and following incubation with mouse liver microsomes. However, whereas CCT066965 was metabolized to the greatest extent in vitro, this compound displayed the slowest plasma clearance. The rank order of the compounds from the highest to lowest area under the curve was the same following discrete and cassette dosing. Furthermore, pharmacokinetic variables were similar whether the compounds were dosed alone or in combination. We conclude that the pharmacokinetics of CCT018159 are complex. Cassette dosing is currently the best option available to assess the pharmacokinetics of this promising series of compounds in relatively high throughput and is now being applied to identify compounds with optimal pharmacokinetic properties during structural analogue synthesis..
Burns, S., Travers, J., Collins, I., Rowlands, M.G., Newbatt, Y., Thompson, N., Garrett, M.D., Workman, P. & Aherne, W.
(2006). Identification of small-molecule inhibitors of protein kinase B (PKB/AKT) in an AlphaScreen (TM) high-throughput screen. J biomol screen,
Protein kinase B (PKB/AKT) has been identified as a promising cancer drug target downstream of PI3 kinase. To find novel inhibitors of PKB/AKT kinase activity for progression as anticancer agents, the authors have used a high-throughput screen based on AlphaScreen (TM) technology. A known kinase inhibitor, the isoquinoline H8, was used as a positive control with mean inhibition in the screen of 43.4% +/- 13.1%. The performance of the screen was highly acceptable with Z' and Z factors of 0.83 +/- 0.07 and 0.75 +/- 0.04, respectively. A number of confirmed hits (similar to 0.1% hit rate) were identified from 63,500 compounds screened. Five compounds have previously been described as PKB inhibitors, demonstrating the ability of the assay to find authentic inhibitors of the enzyme. Five hits had the potential to interfere with the assay signal and were deemed to be false positives. Two compounds were nonspecific inhibitors of PKB as enzyme inhibition in a filter-based assay was markedly reduced in the presence of 0.01% Triton X100. The authors now include an interference assay during hit confirmation procedures and check compound activity in the presence of Triton X100 in an attempt to eliminate nonspecific aggregators at an early stage..
Honma, M., Stubbs, M., Collins, I., Workman, P., Aherne, W. & Watt, F.M.
(2006). Identification of novel keratinocyte differentiation modulating compounds by high-throughput screening. J biomol screen,
The authors have designed high-throughput screens to identify compounds that promote or inhibit terminal differentiation of primary human epidermal keratinocytes. Eleven known inhibitors of signaling pathways and approximately 4000 compounds of diverse structure were screened using an In-Cell Western system based on immunofluorescent staining of the terminal differentiation marker, involucrin. Staurosporine, a nonspecific protein kinase C inhibitor, and H89, a protein kinase A inhibitor, promoted expression of involucrin. Conversely, U0126, a MEK inhibitor, and SAHA or SBHA, 2 histone deacetylase inhibitors, reduced the expression of involucrin during calcium-induced stratification. In addition, the authors found 1 novel compound that induced keratinocyte differentiation and 2 novel compounds that were inhibitory to calcium-induced differentiation. The differentiation-inducing compound also inhibited growth of a human squamous cell carcinoma line by stimulating both differentiation and apoptosis. Because the compound affected the tumor cells at a lower concentration than primary keratinocytes, it may have potential as an antitumor therapy..
Chau, N.-., Rogers, P., Aherne, W., Carroll, V., Collins, I., McDonald, E., Workman, P. & Ashcroft, M.
(2005). Identification of novel small molecule inhibitors of hypoxia-inducible factor-1 that differentially block hypoxia-inducible factor-1 activity and hypoxia-inducible factor-1alpha induction in response to hypoxic stress and growth factors. Cancer res,
Hypoxia-inducible factor-1 (HIF-1) is a transcriptional complex that is activated in response to hypoxia and growth factors. HIF-1 plays a central role in tumor progression, invasion, and metastasis. Overexpression of the HIF-1alpha subunit has been observed in many human cancers and is associated with a poor prognostic outcome with conventional treatments. Targeting HIF-1 using novel small molecule inhibitors is, therefore, an attractive strategy for therapeutic development. We have generated U2OS human osteosarcoma cells stably expressing a luciferase reporter construct under the control of a hypoxia response element (U2OS-HRE-luc). The U2OS-HRE-luc cells were robustly and reproducibly sensitive to hypoxic stress in a HIF-1-dependent manner. We developed an automated U2OS-HRE-luc cell-based assay that was used in a high-throughput screen to identify compounds that inhibited HIF-1 activity induced by treatment with the hypoxia mimetic, deferoxamine mesylate. We performed a pilot screen of the National Cancer Institute Diversity Set of 2,000 compounds. We identified eight hit compounds, six of these were also identified by Rapisarda et al. in an independent hypoxia screen. However, there were two novel hit compounds, NSC-134754 and NSC-643735, that did not significantly inhibit constitutive luciferase activity in U2OS cells (U2OS-luc). We showed that both NSC-134754 and NSC-643735 significantly inhibited HIF-1 activity and HIF-1alpha protein induced by deferoxamine mesylate. Interestingly, NSC-134754 but not NCS-643735 inhibited HIF-1 activity and HIF-1alpha protein induced by hypoxia and significantly inhibited Glut-1 expression. Finally, we showed that both NCS-134754 and NCS-643735 inhibited HIF-1alpha protein induced by insulin-like growth factor-1. Our cell-based assay approach has successfully identified novel compounds that differentially target hypoxia and/or growth factor-mediated induction of HIF-1alpha..
Banerji, U., Walton, M., Raynaud, F., Grimshaw, R., Kelland, L., Valenti, M., Judson, I. & Workman, P.
(2005). Pharmacokinetic-pharmacodynamic relationships for the heat shock protein 90 molecular chaperone inhibitor 17-allylamino, 17-demethoxygeldanamycin in human ovarian cancer xenograft models. Clin cancer res,
(19 Pt 1),
To establish the pharmacokinetic and pharmacodynamic profile of the heat shock protein 90 (HSP90) inhibitor 17-allylamino, 17-demethoxygeldanamycin (17-AAG) in ovarian cancer xenograft models..
Stimson, L., Rowlands, M.G., Newbatt, Y.M., Smith, N.F., Raynaud, F.I., Rogers, P., Bavetsias, V., Gorsuch, S., Jarman, M., Bannister, A., et al.
(2005). Isothiazolones as inhibitors of PCAF and p300 histone acetyltransferase activity. Mol cancer ther,
Histone acetylation plays an important role in regulating the chromatin structure and is tightly regulated by two classes of enzyme, histone acetyltransferases (HAT) and histone deacetylases (HDAC). Deregulated HAT and HDAC activity plays a role in the development of a range of cancers. Consequently, inhibitors of these enzymes have potential as anticancer agents. Several HDAC inhibitors have been described; however, few inhibitors of HATs have been disclosed. Following a FlashPlate high-throughput screen, we identified a series of isothiazolone-based HAT inhibitors. Thirty-five N-substituted analogues inhibited both p300/cyclic AMP-responsive element binding protein-binding protein-associated factor (PCAF) and p300 (1 to >50 micromol/L, respectively) and the growth of a panel of human tumor cell lines (50% growth inhibition, 0.8 to >50 micromol/L). CCT077791 and CCT077792 decreased cellular acetylation in a time-dependent manner (2-48 hours of exposure) and a concentration-dependent manner (one to five times, 72 hours, 50% growth inhibition) in HCT116 and HT29 human colon tumor cell lines. CCT077791 reduced total acetylation of histones H3 and H4, levels of specific acetylated lysine marks, and acetylation of alpha-tubulin. Four and 24 hours of exposure to the compounds produced the same extent of growth inhibition as 72 hours of continuous exposure, suggesting that growth arrest was an early event. Chemical reactivity of these compounds, as measured by covalent protein binding and loss of HAT inhibition in the presence of DTT, indicated that reaction with thiol groups might be important in their mechanism of action. As one of the first series of small-molecule inhibitors of HAT activity, further analogue synthesis is being pursued to examine the potential scope for reducing chemical reactivity while maintaining HAT inhibition..
Banerji, U., O'Donnell, A., Scurr, M., Pacey, S., Stapleton, S., Asad, Y., Simmons, L., Maloney, A., Raynaud, F., Campbell, M., et al.
(2005). Phase I pharmacokinetic and pharmacodynamic study of 17-allylamino, 17-demethoxygeldanamycin in patients with advanced malignancies. J clin oncol,
To study the toxicity and pharmacokinetic-pharmacodynamic profile of 17-allylamino, 17- demethoxygeldanamycin (17-AAG) and to recommend a dose for phase II trials..
Folkes, A.J., Ahmadi, K., Alderton, W.K., Alix, S., Baker, S.J., Box, G., Chuckowree, I.S., Clarke, P.A., Depledge, P., Eccles, S.A., et al.
The identification of 2-(1H-indazol-4-yl)-6-(4-methanesulfonyl-piperazin-1-ylmethyl)-4-morpholin-4-yl-thieno[3,2-d]pyrimidine (GDC-0941) as a potent, selective, orally bioavailable inhibitor of class I PI3 kinase for the treatment of cancer. J med chem,
Phosphatidylinositol-3-kinase (PI3K) is an important target in cancer due to the deregulation of the PI3K/ Akt signaling pathway in a wide variety of tumors. A series of thieno[3,2-d]pyrimidine derivatives were prepared and evaluated as inhibitors of PI3 kinase p110alpha. The synthesis, biological activity, and further profiling of these compounds are described. This work resulted in the discovery of 17, GDC-0941, which is a potent, selective, orally bioavailable inhibitor of PI3K and is currently being evaluated in human clinical trials for the treatment of cancer..
Yap, T.A., Smith, A.D., Ferraldeschi, R., Al-Lazikani, B., Workman, P. & de Bono, J.S.
Drug discovery in advanced prostate cancer: translating biology into therapy. Nat rev drug discov,
Castration-resistant prostate cancer (CRPC) is associated with a poor prognosis and poses considerable therapeutic challenges. Recent genetic and technological advances have provided insights into prostate cancer biology and have enabled the identification of novel drug targets and potent molecularly targeted therapeutics for this disease. In this article, we review recent advances in prostate cancer target identification for drug discovery and discuss their promise and associated challenges. We review the evolving therapeutic landscape of CRPC and discuss issues associated with precision medicine as well as challenges encountered with immunotherapy for this disease. Finally, we envision the future management of CRPC, highlighting the use of circulating biomarkers and modern clinical trial designs..
Rye, C.S., Chessum, N.E., Lamont, S., Pike, K.G., Faulder, P., Demeritt, J., Kemmitt, P., Tucker, J., Zani, L., Cheeseman, M.D., et al.
Discovery of 4,6-disubstituted pyrimidines as potent inhibitors of the heat shock factor 1 (HSF1) stress pathway and CDK9. Medchemcomm,
A series of 4,6-disubstituted pyrimidines from a phenotypic screen targeting the HSF1 pathway are also potent inhibitors of CDK9.