148 results for Denny, William, ResearchSpace@Auckland

  • 7-substituted 2-nitro-5,6-dihydroimidazo[2,1-b][1,3]oxazines: Novel antitubercular agents lead to a new preclinical candidate for visceral leishmaniasis

    Thompson, Andrew; O'Connor, PD; Marshall, AJ; Yardley, V; Maes, L; Gupta, S; Launay, D; Braillard, S; Chatelain, E; Franzblau, SG; Wan, B; Wang, Y; Ma, Z; Cooper, CB; Denny, William (2017-05)

    Journal article
    The University of Auckland Library

    Within a backup program for the clinical investigational agent pretomanid (PA-824), scaffold hopping from delamanid inspired the discovery of a novel class of potent antitubercular agents that unexpectedly possessed notable utility against the kinetoplastid disease visceral leishmaniasis (VL). Following the identification of delamanid analogue DNDI-VL-2098 as a VL preclinical candidate, this structurally related 7-substituted 2-nitro-5,6-dihydroimidazo[2,1-b][1,3]oxazine class was further explored, seeking efficacious backup compounds with improved solubility and safety. Commencing with a biphenyl lead, bioisosteres formed by replacing one phenyl by pyridine or pyrimidine showed improved solubility and potency, whereas more hydrophilic side chains reduced VL activity. In a Leishmania donovani mouse model, two racemic phenylpyridines (71 and 93) were superior, with the former providing >99% inhibition at 12.5 mg/kg (b.i.d., orally) in the Leishmania infantum hamster model. Overall, the 7R enantiomer of 71 (79) displayed more optimal efficacy, pharmacokinetics, and safety, leading to its selection as the preferred development candidate.

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  • Biological characterization of SN32976, a selective inhibitor of PI3K and mTOR with preferential activity to PI3K??, in comparison to established pan PI3K inhibitors

    Rewcastle, Gordon; Kolekar, S; Buchanan, Christina; Gamage, Swarnalatha; Giddens, Anna; Tsang, KY; Kendall, Jackie; Singh, R; Lee, W-J; Smith, GC; Han, W; Matthews, DJ; Denny, William; Shepherd, PR; Jamieson, Stephen (2017-07-18)

    Journal article
    The University of Auckland Library

    Multiple therapeutic agents have been developed to target the phosphatidylinositol 3-kinase (PI3K) signaling pathway, which is frequently dysregulated in cancer promoting tumor growth and survival. These include pan PI3K inhibitors, which target class Ia PI3K isoforms and have largely shown limited single agent activity with narrow therapeutic windows in clinical trials. Here, we characterize SN32976, a novel pan PI3K inhibitor, for its biochemical potency against PI3K isoforms and mTOR, kinase selectivity, cellular activity, pharmacokinetics, pharmacodynamics and antitumor efficacy relative to five clinically-evaluated pan PI3K inhibitors: buparlisib, dactolisib, pictilisib, omipalisib and ZSTK474. SN32976 potently inhibited PI3K isoforms and mTOR, displaying preferential activity for PI3K?? and sparing of PI3K?? relative to the other inhibitors, while showing less off-target activity than the clinical inhibitors in a panel of 442 kinases. The major metabolites of SN32976 were also potent PI3K inhibitors with similar selectivity for PI3K?? as the parent compound. SN32976 compared favorably with the clinically-evaluated PI3K inhibitors in cellular assays, inhibiting pAKT expression and cell proliferation at nM concentrations, and in animal models, inducing a greater extent and duration of pAKT inhibition in tumors than pictilisib, dactolisib and omipalisib at similarly tolerated dose levels and inhibiting tumor growth to a greater extent than dactolisib and ZSTK474 and with similar efficacy to pictilisib and omipalisib. These results suggest that SN32976 is a promising clinical candidate for cancer therapy with enhanced kinase selectivity and preferential inhibition of PI3K?? compared to first generation pan PI3K inhibitors, while retaining comparable anticancer activity.

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  • 6-Nitro-2,3-dihydroimidazo[2,1-b][1,3]thiazoles: Facile synthesis and comparative appraisal against tuberculosis and neglected tropical diseases

    Thompson, Andrew; Blaser, Adrian; Palmer, Brian; Anderson, Robert; Shinde, SS; Launay, D; Chatelain, E; Maes, L; Franzblau, SG; Wan, B; Wang, Y; Ma, Z; Denny, William (2017-06)

    Journal article
    The University of Auckland Library

    As part of a quest for backups to the antitubercular drug pretomanid (PA-824), we investigated the unexplored 6-nitro-2,3-dihydroimidazo[2,1-b][1,3]-thiazoles and related -oxazoles. The nitroimidazothiazoles were prepared in high yield from 2-bromo-4-nitroimidazole via heating with substituted thiiranes and diisopropylethylamine. Equivalent examples of these two structural classes provided broadly comparable MICs, with 2-methyl substitution and extended aryloxymethyl side chains preferred; albeit, S-oxidised thiazoles were ineffective for tuberculosis. Favourable microsomal stability data for a biaryl thiazole (45) led to its assessment in an acute Mycobacterium tuberculosis mouse model, alongside the corresponding oxazole (48), but the latter proved to be more efficacious. In vitro screening against kinetoplastid diseases revealed that nitroimidazothiazoles were inactive versus leishmaniasis but showed interesting activity, superior to that of the nitroimidazooxazoles, against Chagas disease. Overall, "thio-delamanid" (49) is regarded as the best lead.

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  • Exploration of a series of 5-arylidene-2-thioxoimidazolidin-4-ones as inhibitors of the cytolytic protein perforin

    Spicer, Julie; Lena, G; Lyons, DM; Huttunen, KM; Miller, Christian; O'Connor, Patrick; Bull, Matthew; Helsby, Nuala; Jamieson, Stephen; Denny, William; Ciccone, A; Browne, KA; Lopez, JA; Rudd-Schmidt, J; Voskoboinik, I; Trapani, JA (2013)

    Journal article
    The University of Auckland Library

    A series of novel 5-arylidene-2-thioxoimidazolidin-4-ones were investigated as inhibitors of the lymphocyte-expressed pore-forming protein perforin. Structure-activity relationships were explored through variation of an isoindolinone or 3,4-dihydroisoquinolinone subunit on a fixed 2-thioxoimidazolidin-4-one/thiophene core. The ability of the resulting compounds to inhibit the lytic activity of both isolated perforin protein and perforin delivered in situ by natural killer cells was determined. A number of compounds showed excellent activity at concentrations that were nontoxic to the killer cells, and several were a significant improvement on previous classes of inhibitors, being substantially more potent and soluble. Representative examples showed rapid and reversible binding to immobilized mouse perforin at low concentrations (???2.5 ??M) by surface plasmon resonance and prevented formation of perforin pores in target cells despite effective target cell engagement, as determined by calcium influx studies. Mouse PK studies of two analogues showed T1/2 values of 1.1-1.2 h (dose of 5 mg/kg i.v.) and MTDs of 60-80 mg/kg (i.p.).

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  • Development of PIK-75 nanosuspension formulation with enhanced delivery efficiency and cytotoxicity for targeted anti-cancer therapy

    Talekar, M; Ganta, S; Amiji, M; Jamieson, Stephen; Kendall, Jackie; Denny, William; Garg, Sanjay (2013-06)

    Journal article
    The University of Auckland Library

    PIK-75 is a phosphatidylinositol 3-kinase (PI3K) inhibitor that shows selectivity toward p110-?? over the other PI3K class Ia isoforms p110-?? and p110-??, but it lacks solubility, stability and other kinase selectivity. The purpose of this study was to develop folate-targeted PIK-75 nanosuspension for tumor targeted delivery and to improve therapeutic efficacy in human ovarian cancer model. High pressure homogenization was used to prepare the non-targeted and targeted PIK-75 nanosuspensions which were characterized for size, zeta potential, entrapment efficiency, morphology, saturation solubility and dissolution velocity. In vitro analysis of drug uptake, cell viability and cell survival was conducted in SKOV-3 cells. Drug pharmacokinetics and pAkt expression were determined in SKOV-3 tumor bearing mice. PIK-75 nanosuspensions showed an improvement in dissolution velocity and an 11-fold increase in saturation solubility over pre-milled PIK-75. In vitro studies in SKOV-3 cells indicated a 2-fold improvement in drug uptake and 0.4-fold decrease in IC50 value of PIK-75 following treatment with targeted nanosuspension compared to non-targeted nanosuspension. The improvement in cytotoxicity was attributed to an increase in caspase 3/7 and hROS activity. In vivo studies indicated a 5-10-fold increased PIK-75 accumulation in the tumor with both the nanosuspension formulations compared to PIK-75 suspension. The targeted nanosuspension showed an enhanced downregulation of pAkt compared to non-targeted formulation system. These results illustrate the opportunity to formulate PIK-75 as a targeted nanosuspension to enhance uptake and cytotoxicity of the drug in tumor.

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  • Synthesis and biological evaluation of novel phosphatidylinositol 3-kinase inhibitors: Solubilized 4-substituted benzimidazole analogs of 2-(difluoromethyl)-1-[4,6-di(4-morpholinyl)-1,3,5-triazin-2-yl]-1H-benzimidazole (ZSTK474)

    Rewcastle, Gordon; Gamage, Swarnalatha; Flanagan, Jack; Kendall, Jackie; Denny, William; Baguley, Bruce; Buchanan, Christina; Chao, M; Kestell, P; Kolekar, Sharada; Lee, W-J; Lill, CL; Malik, A; Singh, R; Jamieson, Stephen; Shepherd, Peter (2013-06)

    Journal article
    The University of Auckland Library

    A range of 4-substituted derivatives of the pan class I PI 3-kinase inhibitor 2-(difluoromethyl)-1-[4,6-di-(4-morpholinyl)-1,3,5-triazin-2-yl]-1H-benzimidazole (ZSTK474) were prepared in a search for more soluble analogs. 4-Aminoalkoxy substituents provided the most potent derivatives, with the 4-O(CH2)3NMe2 analog (compound 14) being identified as displaying the best overall activity in combination with good aqueous solubility (25 mg/mL for the hydrochloride salt). This compound was tested in a U87MG xenograft model, but displayed less potency than ZSTK474 as a result of an unfavorable pharmacokinetic profile.

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  • Novel pyrazolo[1,5-a]pyridines with improved aqueous solubility as p110??-selective PI3 kinase inhibitors

    Kendall, Jackie; Giddens, Anna; Tsang, KY; Marshall, ES; Lill, CL; Lee, W-J; Kolekar, S; Chao, M; Malik, A; Yu, S; Chaussade, C; Buchanan, Christina; Jamieson, Stephen; Rewcastle, Gordon; Baguley, Bruce; Denny, William; Shepherd, PR (2017-01)

    Journal article
    The University of Auckland Library

    As part of our investigation into pyrazolo[1,5-a]pyridines as novel p110?? selective PI3 kinase inhibitors, we report a range of analogues with improved aqueous solubility by the addition of a basic amine. The compounds demonstrated comparable p110?? potency and selectivity to earlier compounds but with up to 1000?? greater aqueous solubility, as the hydrochloride salts. The compounds also displayed good activity in a cellular assay of PI3 kinase activity.

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  • Benzenesulphonamide inhibitors of the cytolytic protein perforin

    Spicer, Julie; Miller, Christian; O'Connor, PD; Jose, Jiney; Huttunen, KM; Jaiswal, Jagdish; Denny, William; Akhlaghi, H; Browne, KA; Trapani, JA (2017-02)

    Journal article
    The University of Auckland Library

    The pore-forming protein perforin is a key component of mammalian cell-mediated immunity and essential to the pathway that allows elimination of virus-infected and transformed cells. Perforin activity has also been implicated in certain auto-immune conditions and therapy-induced conditions such as allograft rejection and graft versus host disease. An inhibitor of perforin activity could be used as a highly specific immunosuppressive treatment for these conditions, with reduced side-effects compared to currently accepted therapies. Previously identified first-in-class inhibitors based on a 2-thioxoimidazolidin-4-one core show suboptimal physicochemical properties and toxicity toward the natural killer (NK) cells that secrete perforin in vivo. The current benzenesulphonamide-based series delivers a non-toxic bioisosteric replacement possessing improved solubility.

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  • Evidence that phospholipase C is involved in the antitumour action of NSC768313, a new thieno[2,3-b]pyridine derivative

    Reynisson, Johannes; Jaiswal, Jagdish; Barker, David; D'Mello, Stacey; Denny, William; Baguley, Bruce; Leung, Yee Fun (2016-03)

    Journal article
    The University of Auckland Library

    The thieno[2,3-b]pyridines were discovered by virtual high throughput screening as potential inhibitors of phospholipase C (PLC) isoforms and showed potent growth inhibitory effects in National Cancer Institute's human tumour cell line panel (NCI60). The mechanism of the anti-proliferative activity of thieno[2,3-b]pyridines is explored here.We aimed to investigate the basis for the anti-proliferative activity of these thieno[2,3-b]pyridines and to determine whether the cellular inhibition was related to their inhibition of PLC.Four breast cancer cell lines were used to assess the anti-proliferative effects (IC50 values) of six representative thieno[2,3-b]pyridines. The most potent compound (derivative 3; NSC768313), was further studied in MDA-MB-231 cells. DNA damage was examined by ??H2AX expression level, and cell cycle arrest by flow cytometry. Cell morphology was examined by tubulin antibody staining. The growth inhibitory effect of combination treatment with derivative 3 and paclitaxel (tubulin inhibitor), doxorubicin (topoisomerase II inhibitor) or camptothecin (topoisomerase I inhibitor) was evaluated. A preliminary mouse toxicity assay was used to evaluate the pharmacological properties.Addition of the thieno[2,3-b]pyridine derivative 3 to the MDA-MB-231 cells induced G2/M growth inhibition, cell cycle arrest in G2-phase, membrane blebbing and the formation of multinucleated cells. It did not induce DNA damage, mitotic arrest or changes in calcium ion flux. Combination of derivative 3 with paclitaxel showed a high degree of synergy, while combinations with doxorubicin and camptothecin showed only additive effects. A mouse pharmacokinetic study of derivative 3 showed that after intraperitoneal injection of a single does (10??mg/Kg), the Cmax was 0.087????mol/L and the half-life was 4.11??h.The results are consistent with a mechanism in which thieno[2,3-b]pyridine derivatives interact with PLC isoforms (possibly PLC-??), which in turn affect the cellular dynamics of tubulin-??, inducing cell cycle arrest in G2-phase. We conclude that these compounds have novelty because of their PLC target and may have utility in combination with mitotic poisons for cancer treatment.

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  • Synthesis and cytotoxicity of thieno[2,3-b]quinoline-2-carboxamide and cycloalkyl[b]thieno[3,2-e]pyridine-2-carboxamide derivatives

    Leung, Yee Fun; Pilkington, Lisa; van Rensburg, M; Jeon, CY; Song, M; Arabshahi, HJ; De Zoysa, Gayan; Sarojini Amma, Vijayalekshmi; Denny, William; Reynisson, Johannes; Barker, David (2016)

    Journal article
    The University of Auckland Library

    Seventy nine derivatives of thieno[2,3-b]quinolines, tetrahydrothieno[2,3-b]quinoline, dihydrocyclopenta[b]thieno[3,2-e]pyridine, cyclohepta[b]thieno[3,2-e]pyridine and hexahydrocycloocta[b]thieno[3,2-e]pyridine were either synthesized or obtained commercially and tested for their antiproliferative activity against HCT116, MDA-MB-468 and MDA-MB-231 human cancer cell lines. The most potent eight compounds were active against all cell lines with IC50 values in the 80-250nM range. In general hexahydrocycloocta[b]thieno[3,2-e]pyridines were most active with increasing activity observed as larger cycloalkyl rings were fused to the pyridine ring.

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  • Tyrosine Kinase Inhibitors. 20. Optimization of Substituted Quinazoline and Pyrido[3,4- d ]pyrimidine Derivatives as Orally Active, Irreversible Inhibitors of the Epidermal Growth Factor Receptor Family

    Smaill, Jeffrey; Gonzales, AJ; Spicer, Julie; Lee, H; Reed, JE; Sexton, K; Althaus, IW; Zhu, T; Black, Shannon; Blaser, Adrian; Denny, William; Ellis, PA; Fakhoury, S; Harvey, PJ; Hook, K; McCarthy, FOJ; Palmer, Brian; Rivault, F; Schlosser, K; Ellis, T; Thompson, Andrew; Trachet, E; Winters, RT; Tecle, H; Bridges, A (2016-09-08)

    Journal article
    The University of Auckland Library

    Structure???activity relationships for inhibition of erbB1, erbB2, and erbB4 were determined for a series of quinazoline- and pyrido[3,4-d]pyrimidine-based analogues of the irreversible pan-erbB inhibitor, canertinib. Cyclic amine bearing crotonamides were determined to provide rapid inhibition of cellular erbB1 autophosphorylation and good metabolic stability in liver microsome and hepatocyte assays. The influence of 4-anilino substitution on pan-erbB inhibitory potency was investigated. Several anilines were identified as providing potent, reversible pan-erbB inhibition. Optimum 4- and 6-substituents with known 7-substituents provided preferred irreversible inhibitors for pharmacodynamic testing in vivo. Quinazoline 54 and pyrido[3,4-d]pyrimidine 71 were identified as clearly superior to canertinib. Both compounds possess a piperidinyl crotonamide Michael acceptor and a 3-chloro-4-fluoroaniline, indicating these as optimized 6- and 4-substituents, respectively. Pharmacokinetic comparison of compounds 54 and 71 across three species selected compound 54 as the preferred candidate. Compound 54 (PF-00299804) has been assigned the nomenclature of dacomitinib and is currently under clinical evaluation.

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  • A Selective and Slowly Reversible Inhibitor of l-Type Amino Acid Transporter 1 (LAT1) Potentiates Antiproliferative Drug Efficacy in Cancer Cells

    Huttunen, KM; Gynther, M; Huttunen, J; Puris, E; Spicer, Julie; Denny, William (2016-06-23)

    Journal article
    The University of Auckland Library

    The l-type amino acid transporter 1 (LAT1) is a transmembrane protein carrying bulky and neutral amino acids into cells. LAT1 is overexpressed in several types of tumors, and its inhibition can result in reduced cancer cell growth. However, known LAT1 inhibitors lack selectivity over other transporters. In the present study, we designed and synthesized a novel selective LAT1 inhibitor (1), which inhibited the uptake of LAT1 substrate, l-leucin as well as cell growth. It also significantly potentiated the efficacy of bestatin and cisplatin even at low concentrations (25 ??M). Inhibition was slowly reversible, as the inhibitor was able to be detached from the cell surface and blood-brain barrier. Moreover, the inhibitor was metabolically stable and selective toward LAT1. Since the inhibitor was readily accumulated into the prostate after intraperitoneal injection to the healthy mice, this compound may be a promising agent or adjuvant especially for the treatment of prostate cancer.

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  • L-Type amino acid transporter 1 (lat1)-mediated targeted delivery of perforin inhibitors

    Huttunen, KM; Huttunen, J; Aufderhaar, I; Gynther, M; Denny, William; Spicer, Julie (2016-02)

    Journal article
    The University of Auckland Library

    Perforin is a cytolytic pore-forming glycoprotein secreted by cytotoxic effector cells. It is a key component of the immune response against virus-infected and transformed cells and has been implicated in a number of human diseases. Perforin activity can be inhibited by small-molecular-weight compounds, although less is known about their delivery to the site of action. Therefore, in the present study, it was explored if perforin inhibitors could be efficiently and site-selectively delivered firstly into the cytotoxic effector cells and secondly into lytic granules, in which perforin is stored. This was accomplished by designing and synthesizing four prodrugs of perforin inhibitors that could utilize l-type amino acid transporter (LAT1), since activated immune cells are known to over-express LAT1. The results demonstrate that cellular uptake of perforin inhibitors can be increased by LAT1-utilizing prodrugs (into human breast adenocarcinoma cells (MCF-7)). Furthermore, these prodrugs were also able to deliver perforin inhibitors into the cell organelles having lower pH (rat liver lysosomes). Therefore, by using these prodrugs, intracellular mechanisms of perforin inhibitory activity can be studied more thoroughly in future. Moreover, this prodrug approach can be applied for other drugs that would benefit from targeted delivery into cells expressing LAT1, such as cancer.

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  • The preclinical pharmacokinetic disposition of a series of perforin-inhibitors as potential immunosuppressive agents

    Bull, Matthew; Spicer, Julie; Huttunen, KM; Denny, William; Ciccone, A; Browne, KA; Trapani, JA; Helsby, Nuala (2015-12)

    Journal article
    The University of Auckland Library

    The cytolytic protein perforin is a key component of the immune response and is implicated in a number of human pathologies and therapy-induced conditions. A novel series of small molecule inhibitors of perforin function have been developed as potential immunosuppressive agents. The pharmacokinetics and metabolic stability of a series of 16 inhibitors of perforin was evaluated in male CD1 mice following intravenous administration. The compounds were well tolerated 6??h after dosing. After intravenous administration at 5??mg/kg, maximum plasma concentrations ranged from 532??????200 to 10,061??????12??ng/mL across the series. Plasma concentrations were greater than the concentrations required for in vitro inhibitory activity for 11 of the compounds. Following an initial rapid distribution phase, the elimination half-life values for the series ranged from 0.82??????0.25 to 4.38??????4.48??h. All compounds in the series were susceptible to oxidative biotransformation. Following incubations with microsomal preparations, a tenfold range in in vitro half-life was observed across the series. The data suggests that oxidative biotransformation was not singularly responsible for clearance of the compounds and no direct relationship between microsomal clearance and plasma clearance was observed. Structural modifications however, do provide some information as to the relative microsomal stability of the compounds, which may be useful for further drug development.

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  • Systemic and Brain Pharmacokinetics of Perforin Inhibitor Prodrugs

    Gynther, M; Pickering, DS; Spicer, Julie; Denny, William; Huttunen, KM (2016-07-05)

    Journal article
    The University of Auckland Library

    We have recently reported that by converting a perforin inhibitor into an l-type amino acid transporter 1 (LAT1)-utilizing prodrug its cellular uptake can be greatly increased. The aim of the present study was to determine the in vivo and brain pharmacokinetics of two perforin inhibitors and their LAT1-utilizing prodrugs 1 and 2. In addition, the brain uptake mechanism and entry into primary mouse cortical neurons and astrocytes were evaluated. After 23 ??mol/kg i.p. bolus injection, the prodrugs' unbound area under the concentration curve in brain was 0.3 nmol/g ?? min, whereas the parent drugs could not reach the brain. The unbound brain concentrations of the prodrugs after 100 ??M in situ mouse brain perfusion were 521.4 ?? 46.9 and 126.9 ?? 19.9 pmol/g for prodrugs 1 and 2, respectively. The combination of competing transporter substrates for LAT1, l-tryptophan, and for organic anion transporting polypeptides, probenecid, decreased the brain concentrations to 352.4 ?? 44.5 and 70.9 ?? 7.0 pmol/g, respectively. In addition, in vitro uptake studies showed that at 100 ??M prodrug 1 had 3.4-fold and 4.5-fold higher uptake rate into neurons and astrocytes, respectively, compared to its parent drug. Thus, the prodrugs enhance significantly the therapeutic potential of the parent drugs for the treatment of disorders of central nervous system in which neuroinflammation is involved.

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  • Effect of common and experimental anti-tuberculosis treatments on Mycobacterium tuberculosis growing as biofilms

    Dalton, JP; Uy, Benedict; Phummarin, N; Copp, Brent; Denny, William; Swift, Simon; Wiles, Siouxsie (2016)

    Journal article
    The University of Auckland Library

    Much is known regarding the antibiotic susceptibility of planktonic cultures of Mycobacterium tuberculosis, the bacterium responsible for the lung disease tuberculosis (TB). As planktonically-grown M. tuberculosis are unlikely to be entirely representative of the bacterium during infection, we set out to determine how effective a range of anti-mycobacterial treatments were against M. tuberculosis growing as a biofilm, a bacterial phenotype known to be more resistant to antibiotic treatment. Light levels from bioluminescently-labelled M. tuberculosis H37Rv (strain BSG001) were used as a surrogate for bacterial viability, and were monitored before and after one week of treatment. After treatment, biofilms were disrupted, washed and inoculated into fresh broth and plated onto solid media to rescue any surviving bacteria. We found that in this phenotypic state M. tuberculosis was resistant to the majority of the compounds tested. Minimum inhibitory concentrations (MICs) increased by 20-fold to greater than 1,000-fold, underlying the potential of this phenotype to cause significant problems during treatment.

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  • Screening of anti-mycobacterial compounds in a naturally infected zebrafish larvae model

    Dalton, JP; Uy, Benedict; Okuda, KS; Hall, Christopher; Denny, William; Crosier, Philip; Swift, Simon; Wiles, Siouxsie (2017-02)

    Journal article
    The University of Auckland Library

    Mycobacterium tuberculosis is a deadly human pathogen that causes the lung disease TB. M. tuberculosis latently infects a third of the world's population, resulting in ???1.5 million deaths per year. Due to the difficulties and expense of carrying out animal drug trials using M. tuberculosis and rodents, infections of the zebrafish Danio rerio with Mycobacterium marinum have become a useful surrogate. However, the infection methods described to date require specialized equipment and a high level of operator expertise.We investigated whether zebrafish larvae could be naturally infected with bioluminescently labelled M. marinum by immersion, and whether infected larvae could be used for rapid screening of anti-mycobacterial compounds using bioluminescence. We used rifampicin and a variety of nitroimidazole-based next-generation and experimental anti-mycobacterial drugs, selected for their wide range of potencies against M. tuberculosis, to validate this model for anti-mycobacterial drug discovery.We observed that five of the six treatments (rifampicin, pretomanid, delamanid, SN30488 and SN30527) significantly reduced the bioluminescent signal from M. marinum within naturally infected zebrafish larvae. Importantly, these same five treatments also retarded the growth of M. tuberculosis in vitro. In contrast, only three of the six treatments tested (rifampicin, delamanid and SN30527) retarded the growth of M. marinum in vitro.We have demonstrated that zebrafish larvae naturally infected with bioluminescent M. marinum M can be used for the rapid screening of anti-mycobacterial compounds with readily available equipment and limited expertise. The result is an assay that can be carried out by a wide variety of laboratories for minimal cost and without high levels of zebrafish expertise.

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  • 3-(3,4-Dihydroisoquinolin-2(1H)-ylsulfonyl)benzoic Acids: Highly Potent and Selective Inhibitors of the Type 5 17-??-Hydroxysteroid Dehydrogenase AKR1C3

    Jamieson, Stephen; Brooke, Darby; Heinrich, D; Atwell, GJ; Silva, S; Hamilton, EJ; Turnbull, AP; Rigoreau, LJ; Trivier, E; Soudy, C; Samlal, SS; Owen, PJ; Schroeder, E; Raynham, T; Flanagan, Jack; Denny, William (2012)

    Journal article
    The University of Auckland Library

    A high-throughput screen identified 3-(3,4-dihydroisoquinolin-2(1H)-ylsulfonyl)benzoic acid as a novel, highly potent (low nM), and isoform-selective (1500-fold) inhibitor of aldo-keto reductase AKR1C3: a target of interest in both breast and prostate cancer. Crystal structure studies showed that the carboxylate group occupies the oxyanion hole in the enzyme, while the sulfonamide provides the correct twist to allow the dihydroisoquinoline to bind in an adjacent hydrophobic pocket. SAR studies around this lead showed that the positioning of the carboxylate was critical, although it could be substituted by acid isosteres and amides. Small substituents on the dihydroisoquinoline gave improvements in potency. A set of "reverse sulfonamides" showed a 12-fold preference for the R stereoisomer. The compounds showed good cellular potency, as measured by inhibition of AKR1C3 metabolism of a known dinitrobenzamide substrate, with a broad rank order between enzymic and cellular activity, but amide analogues were more effective than predicted by the cellular assay.

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  • Morpholylureas are a new class of potent and selective inhibitors of the type 5 17-??-hydroxysteroid dehydrogenase (AKR1C3)

    Flanagan, Jack; Atwell, GJ; Heinrich, DM; Brooke, DG; Silva, S; Rigoreau, LJM; Trivier, E; Turnbull, AP; Raynham, T; Jamieson, Stephen; Denny, William (2014-02-01)

    Journal article
    The University of Auckland Library

    Inhibitors of the aldo-keto reductase enzyme AKR1C3 are of interest as potential drugs for leukemia and hormone-related cancers. A series of non-carboxylate morpholino(phenylpiperazin-1-yl)methanones were prepared by palladium-catalysed coupling of substituted phenyl or pyridyl bromides with the known morpholino(piperazin-1-yl)methanone, and shown to be potent (IC50???100nM) and very isoform-selective inhibitors of AKR1C3. Lipophilic electron-withdrawing substituents on the phenyl ring were positive for activity, as was an H-bond acceptor on the other terminal ring, and the ketone moiety (as a urea) was essential. These structure-activity relationships are consistent with an X-ray structure of a representative compound bound in the AKR1C3 active site, which showed H-bonding between the carbonyl oxygen of the drug and Tyr55 and His117 in the 'oxyanion hole' of the enzyme, with the piperazine bridging unit providing the correct twist to allow the terminal benzene ring to occupy the lipophilic pocket and align with Phe311.

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  • Arming antibodies with DNA cross-linking agents derived from the duocarmycins

    Lu, Guo-Liang; Tercel, Moana; Giddens, Anna; Lee, H; Bonnet, Muriel; Denny, William; Flygare, JA; Pillow, TA; Safina, BS; Stabenb, LR; Verma, VA; Wei, B (2016-08-28)

    Conference item
    The University of Auckland Library

    Antibody drug conjugates (ADCs) are composed of an antibody covalently linked to a therapeutic agent in such a way that the agent (or ???payload???) is released selectively by antigen-presenting cells. Two ADCs armed with cytotoxic microtubule-binding agents have recently been approved for the treatment of HER2-positive breast cancer and CD30-positive lymphoma, prompting much work on the development of alternative ADCs as anticancer agents. An active area of exploration concerns the search for suitable payloads outside the class of microtubule-binding agents. The duocarmycins are a small group of natural products that alkylate adenine in the minor groove of DNA. They possess several properties that make them attractive as ADC payloads, including high cytotoxic potency, activity against many multidrug-resistant cell lines, and activity against both cycling and non-cycling cells. Simplified and more synthetically accessible variants of the alkylating subunit have been reported which retain the cytotoxic potency of the natural products. This potency can be further enhanced by the preparation of dimeric analogues which cross-link DNA, providing, in some examples, remarkably toxic compounds with IC50s in the fM range. This presentation will report on the synthesis and properties of homo- and heterodimers incorporating duocarmycin analogues, the preparation and antibody-conjugation of drug-linker constructs of the same, and the properties of these new ADC candidates.

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