DNA adduction by the potent carcinogen aflatoxin B1 : mechanistic studies

Aflatoxin B1, a potently carcinogenic fungal metabolite, is converted to the biologically active form by chemical oxidation using dimethyldioxirane and enzymatically by cytochrome P450 mixed-function oxidases. Both processes give rise to mixtures of the exo- and endo-8,9-epoxides. Methanolysis studies reveal exclusive trans opening of both epoxides under neutral conditions in CH3OH and CH3OH/H2O mixtures; an SN2 mechanism is postulated. Under acidic conditions, the exo isomer gives mixtures of trans and cis solvolysis products, suggesting that the reaction is, at least in part, SN1; the endo isomer gives only the trans product. The exo isomer reacts with DNA by attack of the nitrogen atom at the 7 position of guanine on C8 of the epoxide to give the trans adduct; the endo epoxide fails to form an adduct at this or any other site in DNA. The exo isomer is strongly mutagenic in a base-pair reversion assay employing Salmonella typhimurium; the endo isomer is essentially nonmutagenic. Aflatoxin B1 and its derivatives intercalate in DNA. These results are consistent with a mechanism in which intercalation of the exo epoxide optimally orients the epoxide for an SN2 reaction with guanine but intercalation of the endo isomer places the epoxide in an orientation which precludes reaction. Thus, while the exo epoxide is a potent mutagen, the endo epoxide fails to react with DNA.

A new Salmonella typhimurium NM5004 strain expressing rat glutathione S-transferase 5-5 : use in detection of genotoxicity of dihaloalkanes using an SOS/umu test system

The Escherichia coli mu operon was subcloned into a pKK233-2 vector containing rat glutathione S-transferase (GST) 5-5 cDNA and the plasmid thus obtained was introduced into Salmonella typhimurium TA1535. The newly developed strain S.typhimurium NM5004, was found to have 52-fold greater GST activity than the original umu strain S.typhimurium TA1535/pSK1002. We compared sensitivities of these two tester strains, NM5004 and TA1535/ pSK1002, for induction of umuC gene expression with several dihaloalkanes which are activated or inactivated by GST 5-5 activity. The induction of umuC gene expression by these chemicals was monitored by measuring the cellular P-galactosidase activity produced by umuC'lacZ fusion gene in these two tester strains. Ethylene dibromide, 1-bromo-2-chloroethane, 1,2-dichloroethane, and methylene dichloride induced umuC gene expression more strongly in the NM5004 strain than the original strain, 4-Nitroquinoline 1-oxide and N-methyl-N'-nitro-N-nitrosoguanidine were found to induce umuC gene expression to similar extents in both strains. In the case of 1-nitropyrene and 2-nitrofluorene, however, NM5004 strain showed weaker umuC gene expression responses than the original TA1535/ pSK1002 strain, 1,2-Epoxy-3-(4'-nitrophenoxy)propane, a known substrate for GST 5-5, was found to inhibit umuC induction caused by 1-bromo-2-chloroethane. These results indicate that this new tester NM5004 strain expressing a mammalian GST theta class enzyme may be useful for studies of environmental chemicals proposed to be activated or inactivated by GST activity.

Enhancement of bacterial mutagenicity of bifunctional alkylating agents by expression of mammalian glutathione s-transferase

Recently, we inserted the plasmid vector pKK233-2 containing rat GSH S-transferase (GST) 5-5 cDNA into Salmonella typhimurium TA1535 and found that these bacteria [GST 5-5(+)] expressed the protein and produced mutations when ethylene or methylene dihalides were added [Thier, R., Taylor, J. B., Pemble, S. E., Ketterer, B., Persmark, M., Humphreys, W. G., and Guengerich, F. P. (1993) Proc. Natl. Acad. Sci. U.S.A. 90, 8576-8580]. After exposure to the known GST 5-5 substrate 1,2-epoxy-3-(4′-nitrophenoxy)propane, the GST 5-5(+) strain showed fewer mutants than the bacteria transfected with the cDNA clone in a reverse orientation [GST 5-5(-)], suggesting a protective role of GST 5-5. However, mutations were considerably enhanced in the GST 5-5(+) strain [as compared to GST 5-5(-)] when 1,2,3,4-diepoxybutane (butadiene diepoxide) or 1,2-epoxy-4-bromobutane was added. The GST 5-5(+) and GST 5-5(-) bacterial stains showed similar responses to 1,2-epoxypropane, 3,4-epoxy-1-butene, and 1,4-dibromobutane. The results suggest that some bifunctional activated butanes are transformed to mutagenic products through GSH conjugation. We also found that the GST 5-5(+) strain showed enhanced mutagenicity with 1,4-dibromo-2,3-epoxybutane, 1,2-epoxy-3-bromopropane (epibromohydrin), and (±)-1,4-dibromo-2,3-dihydroxybutane. The possibility was considered that a 5-membered thialonium ion may be involved in the mutagenicity. Model thialonium compounds were rather stable to hydrolysis in aqueous solution at pH 7.4 and slowly alkylated 4-(4-nitrobenzyl)pyridine. The presence of a hydroxyl group β to the sulfur did not enhance reactivity. Mechanisms involving episulfonium ions are considered more likely. Potential oxidation products of the toxic pesticide 1,2-dibromo-3-chloropropane (DBCP) were also considered in this system. DBCP itself gave rather similar results in the two strains. Others have reported that oxidation of DBCP is required for mutagenicity, along with GST-catalyzed GSH conjugation [Simula, T. P., Glancey, M. J., Söderlund, E. J., Dybing, E., and Wolf, C. R. (1993) Carcinogenesis 14, 2303-2307]. The putative oxidation product 1,2-dibromopropional did not show a difference between the two strains. However, 1,3-dichloroacetone, a model for the putative oxidation product 1-bromo-3-chloroacetone, was considerably more mutagenic in the GST 5-5(+) strain.

Human glutathione S-transferase T1-1 enhances mutagenicity of 1,2-dibromoethane, dibromomethane and 1,2,3,4-diepoxybutane in Salmonella typhimurium

The rat theta class glutathione S-transferase (GST) 5-5 has been shown to affect the mutagenicity of halogenated alkanes and epoxides. In Salmonella typhimurium TA1535 expressing the rat GST5-5 the number of revertants was increased compared to the control strain by CH2Br2, ethylene dibromide (EDB) and 1,2,3,4-diepoxybutane (BDE); in contrast, mutagenicity of 1,2-epoxy-3-(4'-nitrophenoxy)propane (EPNP) was reduced. S.typhimurium TA1535 cells were transformed with an expression plasmid carrying the cDNA of the human theta ortholog GST1-1 either in sense or antisense orientation, the latter being the control. These transformed bacteria were utilized for mutagenicity assays. Mutagenicity of EDB, BDE, CH2Br2, epibromohydrin and 1,3-dichloroacetone was higher in the S.typhimurium TA1535 expressing GSTT1-1 than in the control strain. The expression of active enzyme did not affect the mutagenicity of 1,2-epoxy-3-butene or propylene oxide, GSTT1-1 expression reduced the mutagenicity of EPNP. Glutathione S-transferase 5-5 and GSTT1-1 modulate genotoxicity of several industrially important chemicals in the same way. Polymorphism of the GSTT1 locus in humans may therefore cause differences in cancer susceptibility between the two phenotypes.

Technicolouring the fridge : reducing food waste through uses of colour-coding and cameras

Domestic food wastage is a growing problem for the environment and food security. Some causes of domestic food wastes are attributed to a consumer’s behaviours during food purchasing, storage and consumption, such as: excessive food purchases and stockpiling in storage. Recent efforts in human-computer interaction research have examined ways of influencing consumer behaviour. The outcomes have led to a number of interventions that assist users with performing everyday tasks. The Internet Fridge is an example of such an intervention. However, new pioneering technologies frequently confront barriers that restrict their future impact in the market place, which has prompted investigations into the effectiveness of behaviour changing interventions used to encourage more sustainable practices. In this paper, we investigate and compare the effectiveness of two interventions that encourage behaviour change: FridgeCam and the Colour Code Project. We use FridgeCam to examine how improving a consumer’s food supply knowledge can reduce food stockpiling. We use the Colour Code Project to examine how improving consumer awareness of food location can encourage consumption of forgotten foods. We explore opportunities to integrate these interventions into commercially available technologies, such as the Internet Fridge, to: (i) increase the technology’s benefit and value to users, and (ii) promote reduced domestic food wastage. We conclude that interventions improving consumer food supply and location knowledge can promote behaviours that reduce domestic food waste over a longer term. The implications of this research present new opportunities for existing and future technologies to play a key role in reducing domestic food waste.

Designing for grassroots food production : an event-based urban agriculture community

As urbanisation of the global population has increased above 50%, growing food in urban spaces increases in importance, as it can contribute to food security, reduce food miles, and improve people’s physical and mental health. Approaching the task of growing food in urban environments is a mixture of residential growers and groups. Permablitz Brisbane is an event-centric grassroots community that organises daylong ‘working bee’ events, drawing on permaculture design principles in the planning and design process. Permablitz Brisbane provides a useful contrast from other location-centric forms of urban agriculture communities (such as city farms or community gardens), as their aim is to help encourage urban residents to grow their own food. We present findings and design implications from a qualitative study with members of this group, using ethnographic methods to engage with and understand how this group operates. Our findings describe four themes that include opportunities, difficulties, and considerations for the creation of interventions by Human-Computer Interaction (HCI) designers.

From bench to bedside : immunotherapy for prostate cancer

The mainstay therapeutic strategy for metastatic castrate-resistant prostate cancer (CRPC) continues to be androgen deprivation therapy usually in combination with chemotherapy or androgen receptor targeting therapy in either sequence, or recently approved novel agents such as Radium 223. However, immunotherapy has also emerged as an option for the treatment of this disease following the approval of sipuleucel-T by the FDA in 2010. Immunotherapy is a rational approach for prostate cancer based on a body of evidence suggesting these cancers are inherently immunogenic and, most importantly, that immunological interventions can induce protective antitumour responses. Various forms of immunotherapy are currently being explored clinically, with the most common being cancer vaccines (dendritic-cell, viral, and whole tumour cell-based) and immune checkpoint inhibition. This review will discuss recent clinical developments of immune-based therapies for prostate cancer that have reached the phase III clinical trial stage. A perspective of how immunotherapy could be best employed within current treatment regimes to achieve most clinical benefits is also provided.

Meta-analysis of microarray data identifies GAS6 expression as an independent predictor of poor survival in ovarian cancer

Seeking new biomarkers for epithelial ovarian cancer, the fifth most common cause of death from all cancers in women and the leading cause of death from gynaecological malignancies, we performed a meta-analysis of three independent studies and compared the results in regard to clinicopathological parameters. This analysis revealed that GAS6 was highly expressed in ovarian cancer and therefore was selected as our candidate of choice. GAS6 encodes a secreted protein involved in physiological processes including cell proliferation, chemotaxis, and cell survival. We performed immunohistochemistry on various ovarian cancer tissues and found that GAS6 expression was elevated in tumour tissue samples compared to healthy control samples (P < 0.0001). In addition, GAS6 expression was also higher in tumours from patients with residual disease compared to those without. Our data propose GAS6 as an independent predictor of poor survival, suggesting GAS6, both on the mRNA and on the protein level, as a potential biomarker for ovarian cancer. In clinical practice, the staining of a tumour biopsy for GAS6 may be useful to assess cancer prognosis and/or to monitor disease progression.

Fine-mapping the HOXB region detects common variants tagging a rare coding allele : evidence for synthetic association in prostate cancer

The HOXB13 gene has been implicated in prostate cancer (PrCa) susceptibility. We performed a high resolution fine-mapping analysis to comprehensively evaluate the association between common genetic variation across the HOXB genetic locus at 17q21 and PrCa risk. This involved genotyping 700 SNPs using a custom Illumina iSelect array (iCOGS) followed by imputation of 3195 SNPs in 20,440 PrCa cases and 21,469 controls in The PRACTICAL consortium. We identified a cluster of highly correlated common variants situated within or closely upstream of HOXB13 that were significantly associated with PrCa risk, described by rs117576373 (OR 1.30, P = 2.62×10(-14)). Additional genotyping, conditional regression and haplotype analyses indicated that the newly identified common variants tag a rare, partially correlated coding variant in the HOXB13 gene (G84E, rs138213197), which has been identified recently as a moderate penetrance PrCa susceptibility allele. The potential for GWAS associations detected through common SNPs to be driven by rare causal variants with higher relative risks has long been proposed; however, to our knowledge this is the first experimental evidence for this phenomenon of synthetic association contributing to cancer susceptibility.

Development of a polymer theranostic for prostate cancer

Theranostics offers an improved treatment strategy for prostate cancer by facilitating simultaneous targeting of tumour cells with subsequent drug delivery and imaging. In this report we describe the synthesis of hyperbranched polymers that are biocompatible, can specifically target and be internalised by prostate cancer cells (through targeting of prostate-specific membrane antigen – PSMA) and ultimately facilitate controlled delivery of a model drug. The theranostic also incorporates a far-red fluorescent dye that allows tracking of the polymer via optical imaging. Controlled synthesis of the polymer is achieved via reversible addition fragmentation chain transfer polymerisation of polyethylene glycol monomethyl methacrylate, with ethylene glycol dimethacrylate as the branching agent. Incorporation of 20 mol% of an hydrazide-methacrylate monomer allows post-ligation of a model drug, fluorene-2-carboxaldehyde, through a hydrolytically-degradable hydrazone linkage. The rate of degradation of this particular linker was enhanced at endosomal pH (pH = 5.5) where [similar]95% of the model drug was released in 4 hours compared to less than 5% released over the same period at physiological pH. The theranostic showed high uptake into prostate cancer cells expressing prostate-specific membrane antigen, while minimal uptake was observed in PC3 cells negative for PSMA, highlighting the enhanced efficacy of the targeting ligand.

Pharmacokinetics and tumor disposition of PEGylated, methotrexate conjugated poly-l-lysine dendrimers

Dendrimers have potential for delivering chemotherapeutic drugs to solid tumours via the enhanced permeation and retention (EPR) effect. The impact of conjugation of hydrophobic anticancer drugs to hydrophilic PEGylated dendrimer surfaces, however, has not been fully investigated. The current study has therefore characterised the effect on dendrimer disposition of conjugating α-carboxyl protected methotrexate (MTX) to a series of PEGylated 3H-labelled poly-L-lysine dendrimers ranging in size from generation 3 (G3) to 5 (G5) in rats. Dendrimers contained 50% surface PEG and 50% surface MTX. Conjugation of MTX generally increased plasma clearance when compared to conjugation with PEG alone. Conversely, increasing generation reduced clearance, increased metabolic stability and reduced renal elimination of the administered radiolabel. For constructs with molecular weights >20 kDa increasing the molecular weight of conjugated PEG also reduced clearance and enhanced metabolic stability but had only a minimal effect on renal elimination. Tissue distribution studies revealed retention of MTX conjugated smaller (G3-G4) PEG570 dendrimers (or their metabolic products) in the kidneys. In contrast, the larger G5 dendrimer was concentrated more in the liver and spleen. The G5 PEG1100 dendrimer was also shown to accumulate in solid Walker 256 and HT1080 tumours and comparative disposition data in both rats (1 to 2% dose/g in tumour) and mice (11% dose/g in tumour) are presented. The results of this study further illustrate the potential utility of biodegradable PEGylated poly-L-lysine dendrimers as long circulating vectors for the delivery and tumour-targeting of hydrophobic drugs.

PSMA-targeting iron oxide magnetic nanoparticles enhance MRI of preclinical prostate cancer

Aim Evaluate potential of newly-developed, biocompatible iron oxide magnetic nanoparticles (MNPs) conjugated with J591, an antibody to an extracellular epitope of prostate specific membrane antigen (PSMA), to enhance MRI of prostate cancer (PCa). Materials & Methods Specific binding to PSMA by J591-MNP was investigated in vitro. MRI studies were performed on orthotopic tumor-bearing NOD.SCID mice 2h and 24hr after intravenous injection of J591-MNPs, or non-targeting MNPs. Results and Conclusions In vitro, MNPs did not affect PCa cell viability, and conjugation to J591 did not compromise antibody specificity and enhanced cellular iron uptake. In vivo, PSMA-targeting MNPs increased MR contrast of tumors, but not by non-targeting MNPs. This provides proof-of-concept that PSMA-targeting MNPs have potential to enhance MR detection/localization of PCa.,

DIY media architecture: Open and participatory approaches to community engagement

Media architecture’s combination of the digital and the physical can trigger, enhance, and amplify urban experiences. In this paper, we examine how to bring about and foster more open and participatory approaches to engage communities through media architecture by identifying novel ways to put some of the creative process into the hands of laypeople. We review technical, spatial, and social aspects of DIY phenomena with a view to better understand maker cultures, communities, and practices. We synthesise our findings and ask if and how media architects as a community of practice can encourage the ‘open-sourcing’ of information and tools allowing laypeople to not only participate but become active instigators of change in their own right. We argue that enabling true DIY practices in media architecture may increase citizen control. Seeking design strategies that foster DIY approaches, we propose five areas for further work and investigation. The paper begs many questions indicating ample room for further research into DIY Media Architecture.

Writing embodied practice from the inside : outside the exegesis

The textual turn is a good friend of expert spectating, where it assumes the role of writing-productive apparatus, but no friend at all of expert practices or practitioners (Melrose, 2003). Introduction The challenge of time-based embodied performance when the artefact is unstable As a former full-time professional practitioner with an embodied dance practice as performer, choreographer and artistic director for three decades, I somewhat unexpectedly entered the world of academia in 2000 after completing a practice-based PhD, which was described by its examiners as ‘pioneering’. Like many artists my intention was to deepen and extend my practice through formal research into my work and its context (which was intercultural) and to privilege the artist’s voice in a research world where it was too often silent. Practice as research, practice-based research, and practice-led research were not yet fully named. It was in its infancy and my biggest challenge was to find a serviceable methodology which did not betray my intentions to keep practice at the centre of the research. Over the last 15 years, practice led doctoral research, where examinable creative work is placed alongside an accompanying (exegetical) written component, has come a long way. It has been extensively debated with a range of theories and models proposed (Barrett & Bolt, 2007, Pakes, 2003 & 2004, Piccini, 2005, Philips, Stock & Vincs 2009, Stock, 2009 & 2010, Riley & Hunter 2009, Haseman, 2006, Hecq, 2012). Much of this writing is based around epistemological concerns where the research methodologies proposed normally incorporate a contextualisation of the creative work in its field of practice, and more importantly validation and interrogation of the processes of the practice as the central ‘data gathering’ method. It is now widely accepted, at least in the Australian creative arts context, that knowledge claims in creative practice research arise from the material activities of the practice itself (Carter, 2004). The creative work explicated as the tangible outcome of that practice is sometimes referred to as the ‘artefact’. Although the making of the artefact, according to Colbert (2009, p. 7) is influenced by “personal, experiential and iterative processes”, mapping them through a research pathway is “difficult to predict [for] “the adjustments made to the artefact in the light of emerging knowledge and insights cannot be foreshadowed”. Linking the process and the practice outcome most often occurs through the textual intervention of an exegesis which builds, and/or builds on, theoretical concerns arising in and from the work. This linking produces what Barrett (2007) refers to as “situated knowledge… that operates in relation to established knowledge” (p. 145). But what if those material forms or ‘artefacts’ are not objects or code or digitised forms, but live within the bodies of artist/researchers where the nature of the practice itself is live, ephemeral and constantly transforming, as in dance and physical performance? Even more unsettling is when the ‘artefact’ is literally embedded and embodied in the work and in the maker/researcher; when subject and object are merged. To complicate matters, the performing arts are necessarily collaborative, relying not only on technical mastery and creative/interpretive processes, but on social and artistic relationships which collectively make up the ‘artefact’. This chapter explores issues surrounding live dance and physical performance when placed in a research setting, specifically the complexities of being required to translate embodied dance findings into textual form. Exploring how embodied knowledge can be shared in a research context for those with no experiential knowledge of communicating through and in dance, I draw on theories of “dance enaction” (Warburton, 2011) together with notions of “affective intensities” and “performance mastery” (Melrose, 2003), “intentional activity” (Pakes, 2004) and the place of memory. In seeking ways to capture in another form the knowledge residing in live dance practice, thus making implicit knowledge explicit, I further propose there is a process of triple translation as the performance (the living ‘artefact’) is documented in multi-facetted ways to produce something durable which can be re-visited. This translation becomes more complex if the embodied knowledge resides in culturally specific practices, formed by world views and processes quite different from accepted norms and conventions (even radical ones) of international doctoral research inquiry. But whatever the combination of cultural, virtual and genre-related dance practices being researched, embodiment is central to the process, outcome and findings, and the question remains of how we will use text and what forms that text might take.

Citizen’s right to the digital city: Urban interfaces, activism, and placemaking

Edited by thought leaders of the fields of urban informatics and urban interaction design, this book brings together case studies and examples from around the world to discuss the role that urban Interfaces, citizen action, and city making play in the quest to create and maintain not only secure and resilient, but productive, sustainable, and liveable urban environments. The book debates the impact of these trends on theory, policy, and practice. The chapters in this book are sourced from blind peer reviewed contributions by leading researchers working at the intersection of the social / cultural, technical / digital, and physical / spatial domains of urbanism scholarship. The book appeals not only to research colleagues and students, but also to a vast number of practitioners in the private and public sector interested in accessible accounts that clearly and rigorously analyse the affordances and possibilities of urban interfaces, mobile technology, and location-based services to engage people towards open, smart and participatory urban environments.