Prenatal exposure to mixture N1, a chemical mixture consisting of four phthalate diesters,three pesticides and Bisphenol A, has been associated with behavioural changes as well as changes in gene expression in mice. In this study it was investigated whether the changes in gene expression could be explained by changes in DNA methylation. Mixture N1 was found to significantly change DNA methylation in three different genes (Nr3c1, Nr3c2 and Crhr1) on totally eight different Cytosine Guanine dinucleotides (CpG) positions. To further investigate whether these changes could be induced already during differentiation of cells, a hippocampal cell model HT22 was exposed to mixture N1 during differentiation. In this mode, mixture N1 induced a statistically significant change in the promoter region of Nr3c1. Unfortunately, this change could however not be validated, and the experiments would need to be repeated. In conclusion, this study showed that exposure to mixture N1 can result in changes in DNA methylation.

京都大学 / 新制・課程博士 / 博士(工学) / 甲第24222号 / 工博第5050号 / 新制||工||1788(附属図書館) / 京都大学大学院工学研究科都市環境工学専攻 / (主査)教授 米田 稔, 教授 松井 康人, 准教授 松田 知成 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

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Hormones are endocrine disrupting compounds, which have been shown to alter the sexual development of aquatic organisms. Animal manure applications to agricultural fields for nutrient management can be a source of environmental hormones. This dissertation investigates the occurrence of hormones in fields applied with various manures and their adjacent streams, as well as the effect of manure application technologies on the fate of hormones in soils, sediments, and runoff. A total of 11 hormone compounds were studied. All studied analytes were quantified using liquid chromatography and triple-quadrupole mass spectrometry following various sample extraction and clean-up strategies. The spatial and temporal distribution of manure-associated hormones in a manure surface applied agricultural field and adjacent stream was studied at time points up to 7.5 months after a routine manure application. Hormones were detected mainly in the top 0-5cm soils. Significantly higher levels of hormones were found in the drystack applied area of the field when compared to dairy manure slurry applied portion. New technologies for the subsurface application of poultry litter show promise as a tool to reduce the transportation of environmental hormones in surface runoff. Once adequate sampling protocols were established; it was determined that subsurface injection of both dairy manure and poultry litter reduced the impact of manure surface runoff. Hormones also showed little vertical and lateral movement in the soil. The transformation rates of 1,4-androstadiene-3,17-dione, 4-androstene-3,17-dione and estrone were studied comparing the effects of temperature, soil type, and application type. The calculated half-life of 1,4-androstadiene-3,17-dione in poultry litter surface-applied soils was 1.9 times higher than that in the poultry litter subsurface-injected soils, indicating a faster dissipation rate in the injection slits. Estrone persisted at detectable levels for the duration of the study in all treatments. The continued use of best management practices and innovative manure management techniques for the reduction of nutrients, sediment and other contaminants has the potential to also reduce hormone transport to the natural environment. Monitoring many different types of hormones in all areas of an environmental system will continue to provide better information on the occurrence and fate of hormones sourced from manure amended soils. / Ph. D.

The Potomac River Basin is significant for both public and ecological health as it flows directly into the ecologically-sensitive Chesapeake Bay. It is a drinking water source for about 5 million people living in Maryland, Virginia, and Washington D.C. The discovery of intersex fish, an indicator of poor ecological health, in the Chesapeake Bay occurred in the 2000s, and has led to a series of studies in the watershed to determine the sources and magnitude of endocrine disruption. Endocrine disrupting compounds (EDCs) are exogenous chemicals that interfere with the endocrine system and can cause detrimental health effects at low concentrations. This study aims to understand a best management practice referred to as planned indirect potable reuse (IPR) and its impacts on EDCs. The Occoquan Watershed is a planned IPR subwatershed of the Potomac River Basin. Water samples were collected at the water reclamation plant discharge (Upper Occoquan Service Authority), up- and downstream of that location along Bull Run, and at the water treatment plant intake (Frederick P. Griffith WTP) in the Occoquan Watershed to assess planned IPR. Samples were also collected at a water treatment plant (James J. Corbalis WTP) along the Potomac River for comparison as an unplanned IPR location. These two groups of samples were analyzed for EDCs (categorized into two groups: estrogen hormones and other synthetic organic compounds (SOCs)), nutrients, and other water quality parameters. The infrequency of estrogen hormones and SOC patterns indicate planned and unplanned IPR are both viable approaches to provide safe drinking water / Master of Science / Our river systems are important to maintain both for human and environmental health. The Potomac River Basin is the area of land drained by the Potomac River and its tributaries. The Potomac River Basin is significant for both public and ecological health as it flows directly into the ecologically-sensitive Chesapeake Bay. It is a drinking water source for about 5 million people living in Maryland, Virginia, and Washington D.C. The discovery of intersex fish, or when a single fish has both male and female characteristics, occurred in the Chesapeake Bay in the 2000s. Fish health is often an indicator of poor environmental health, and in this case endocrine disruption. This discovery led to a series of studies in the watershed to determine the sources and magnitude of endocrine disruption. Endocrine disrupting compounds (EDCs) are external chemicals that interfere with the endocrine system once they enter the body of a human or another organism, and can cause detrimental health effects even at low concentrations. This study aims to understand a best management practice, or a type of water pollution control, referred to as planned indirect potable reuse (IPR) and its impacts on EDCs. IPR occurs when wastewater from a community is discharged into to a river or a reservoir, and then downstream it is withdrawn from that same source for drinking water purposes. This can be either planned or unplanned. Planned IPR is becoming more common as population, especially in urban areas, increases. The Occoquan Watershed is a planned IPR subwatershed of the Potomac River Basin. Water samples were collected at the water reclamation plant discharge (Upper Occoquan Service Authority), up- and downstream of that location along Bull Run, and at the water treatment plant intake (Frederick P. Griffith WTP) in the Occoquan Watershed to assess planned IPR. Samples were also collected at a water treatment plant (James J. Corbalis WTP) along the Potomac River for comparison as an unplanned IPR location. These two groups of samples were analyzed for EDCs (categorized into two groups: estrogen hormones and other synthetic organic compounds (SOCs)), nutrients, and other water quality parameters. The infrequency of estrogen hormones and SOC patterns indicate planned and unplanned IPR are both viable approaches to provide safe drinking water.

No / Vascular disrupting agents (VDA) offer a strategy to starve solid tumors of nutrients and oxygen concomitant with tumor shrinkage. Several VDAs have progressed into early clinical trials, but their therapeutic value seems to be compromised by systemic toxicity. In this report, we describe the design and characterization of a novel VDA, ICT2588, that is nontoxic until activated specifically in the tumor by membrane-type 1 matrix metalloproteinase (MT1-MMP). HT1080 cancer cells expressing MT1-MMP were selectively chemosensitive to ICT2588, whereas MCF7 cells that did not express MT1-MMP were nonresponsive. Preferential hydrolysis of ICT2588 to its active metabolite (ICT2552) was observed in tumor homogenates of HT1080 relative to MCF7 homogenates, mouse plasma, and liver homogenate. ICT2588 activation was inhibited by the MMP inhibitor ilomastat. In HT1080 tumor-bearing mice, ICT2588 administration resulted in the formation of the active metabolite, diminution of tumor vasculature, and hemorrhagic necrosis of the tumor. The antitumor activity of ICT2588 was superior to its active metabolite, exhibiting reduced toxicity, improved therapeutic index, enhanced pharmacodynamic effect, and greater efficacy. Coadministration of ICT2588 with doxorubicin resulted in a significant antitumor response (22.6 d growth delay), which was superior to the administration of ICT2588 or doxorubicin as a single agent, including complete tumor regressions. Our findings support the clinical development of ICT2588, which achieves selective VDA targeting based on MT-MMP activation in the tumor microenvironment.

Yes / Non-invasive methods to monitor tumour growth are an important goal in cancer drug development. Thermographic imaging systems offer potential in this area, since a change in temperature is known to be induced due to changes within the tumour microenvironment. This study demonstrates that this imaging modality can be applied to a broad range of tumour xenografts and also, for the first time, the methodology’s suitability to assess anti-cancer agent efficacy. Mice bearing subcutaneously implanted H460 lung cancer xenografts were treated with a novel vascular disrupting agent, ICT-2552, and the cytotoxin doxorubicin. The effects on tumour temperature were assessed using thermographic imaging over the first 6 hours post-administration and subsequently a further 7 days. For ICT-2552 a significant initial temperature drop was observed, whilst for both agents a significant temperature drop was seen compared to controls over the longer time period. Thus thermographic imaging can detect functional differences (manifesting as temperature reductions) in the tumour response to these anti-cancer agents compared to controls. Importantly, these effects can be detected in the first few hours following treatment and therefore the tumour is observable non-invasively. As discussed, this technique will have considerable 3Rs benefits in terms of reduction and refinement of animal use. / University of Bradford

No / Development of therapeutic strategies for tumor-selective delivery of therapeutics through exploitation of the proteolytic tumor phenotype has significant scope for improvement of cancer treatment. ICT2588 is a peptide-conjugated prodrug of the vascular disrupting agent (VDA) azademethylcolchicine developed to be selectively hydrolyzed by matrix metalloproteinase-14 (MMP-14) within the tumor. In this report, we extend our previous proof-of-concept studies and demonstrate the therapeutic potential of this agent against models of human colorectal, lung, breast, and prostate cancer. In all tumor types, ICT2588 was superior to azademethylcolchicine and was greater or comparable to standard clinically used agents for the respective tumor type. Prodrug activation in clinical human lung tumor homogenates relative to stability in human plasma and liver was observed, supporting clinical translation potential. A major limiting factor to the clinical value of VDAs is their inherent cardiovascular toxicity. No increase in plasma von Willebrand factor (vWF) levels, an indicator of systemic vascular dysfunction and acute cardiovascular toxicity, was detected with ICT2588, thereby supporting the tumor-selective activation and reduced potential of ICT2588 to cause cardiovascular toxicity. Our findings reinforce the improved therapeutic index and tumorselective approach offered by ICT2588 and this nanotherapeutic approach.

Steroid hormones belong to a group of compounds known as endocrine disruptors. They are hydrophobic compounds and are categorized as natural and synthetic estrogens. Some common household products have been implicated as estrogen mimics. Exposure effects of these compounds are felt by human and wildlife, such reproductive alterations in fish and frogs. They mainly introduced into the environment through veterinary medicines administration to animals and the discharges from wastewater treatment plants (WWTPs). In this study, a new alternative analytical procedure that is simple, rapid and fast for the determination and quantification of five steroidal hormones: estriol (E3), beta estradiol (β-E2), alpha estradiol (α-E2), testosterone (T), progesterone (P) and bisphenol A (BPA) using the High pressure liquid chromatography coupled to a charged aerosol detector (HPLC-CAD). These compounds were studied because of their strong endocrine-disrupting effects in the environment. Under optimum conditions, a linear graph was obtained with correlation coefficient (R2) ranging from 0.9952 - 0.9996. The proposed method was applied to the analysis of water samples from a wastewater plant and the results obtained were satisfactory. The limits of detection (LOD) for the target analytes in wastewater influent was between 0.0002 – 0.0004 μg/L and the limit of quantification (LOQ) was 0.001 μg/L respectively for each of the analytes. Enrichment factors of 148- 258, and extraction efficiency 84- 102% were obtained for the target analytes; relative standard deviations (% RSD) for m = 6 were between 2.8 and 7.6%. The concentration of the EDCs in environment sample was between 0.2 - 2.3 μg/L. / Chemistry / M. Sc. (Chemistry)