9^th Euro-Global Summit on Toxicology and Applied Pharmacology June 22-24, 2017 Paris, France

Biography:

Abstract:

Endocrine-disrupting chemicals (EDCs) interfere with normal function of the endocrine system and have been associated with metabolic disorders, immune dysfunction, developmental defects, and cancer. Thus, presence of EDCs in the environment and water sources in particular, is a major health concern. Standard detection of EDCs in the environment relies on a laborious analysis of chemical structures using HPLC, MS/GS and related technologies. These methods are costly, time consuming and frequently fail to identify a specific chemical structure as many chemicals are subjected to bio-modifications in the environment. These derivatives cannot be easily identified and are not present in the currently existing libraries. Consequently, their levels are not efficiently monitored or regulated. In addition, it is unclear whether the EDCs detected by chemical methods elicit biological responses in mammalian systems. To overcome these obstacles, we developed a high-throughput assay for biological testing of EDCs using mammalian cell lines that express GFP-tagged nuclear steroid receptor constructs. This assay is based on translocation of fluorescently labeled nuclear receptor construct from the cytoplasm to the nucleus in the presence of EDCs acting on this particular receptor. Using this assay, we detected androgen activity in 35% of the tested water samples, and a previously unrecognized glucocorticoid (GC) activity in 27% of the samples. We also developed a novel call line expressing a GFP-tagged glucocorticoid receptor-thyroid receptor (GFP-GR-TRβ) chimeric construct which could detect known EDCs such as BPA and TBBPA as well as TRβ-interacting contaminants in water samples. We conclude that the wide-spread contamination of water sources with different classes of EDCs is a possible health hazard not only for the aquatic ecosystems, but may also negatively impact the human population. Largely unrestricted human activity with respect to pharmaceuticals and other potential endocrine disruptors is of concern and represents one of the main reasons for these wide-spread contaminations

Biography:

Branko Petrinec obtained his PhD degree from the Physics department of the University of Zagreb. He is a Research Associate at the Institute for Medical Research and Occupational Health in Zagreb, and an Assistant Professor in the Department of Physics, Josip Juraj Strossmayer University in Osijek. He has published more than 20 papers in reputed journals. He was the President for the Scientific Committee of the 10^th Symposium of the Croatian Radiation Protection Association.

Abstract:

We collected samples in accordance with the standard procedures recommended by the International Atomic Energy Association, within an internal project of the Institute for Medical Research and Occupational Health and in collaboration with the Physics department of the University of Osijek. The activity concentration of ¹³⁷Cs was determined using a high-purity germanium gamma-ray spectrometry system (FWHM of 2.24 keV and relative efficiency of 74.2%, all at 1.33 MeV ⁶⁰Co). The measured activity concentrations in soil from 11 locations range from 1.81±0.16 Bq/kg (Kozjak) to 10.65±0.26 Bq/kg (Hulovski channel), averaging to of 7.20±5.57 Bq/kg which is comparable to other parts of Croatia. Data sets consisting of measured activity concentrations of ¹³⁷Cs in soil and water samples within the boundaries of the Nature Park Kopacki Rit were used for the risk assessment for the wildlife and dose rates for freshwater and terrestrial organisms were estimated. For the freshwater ecosystem in Kopački Rit, the highest risk quotient (RQ) values were obtained for “pelagic fish” 0.038 and “amphibian” 0.037, respectively. Appropriate dose rates were below the screening level of 10 μGy h^-1: amphibian 0.37 μGy h^-1, bird 0.27 μGy h^-1, insect larvae 0.12 μGy h^-1, mammal 0.29 μGy h^-1, pelagic fish 0.38 μGy h^-1 and vascular plant 0.022 μGy h^-1. Also, for terrestrial ecosystems, all RQs are far below 1 (the highest values are 0.0013 for “lichen”, 0.0019 for “small mammal” and 0.0027 for “large mammal”) and the dose rates to species studied were clearly below the screening level of 10 μGy h^-1 (one-thousandth of this value: 0.0128 μGy h^-1 for the lichen, 0.0196 μGy h^-1 and 0.0277 μGy h^-1 for small and large mammals, respectively) indicating no significant impact of the ¹³⁷Cs fallout on all of the investigated species.

Biography:

Gyuseong Cho is a Professor of the Department of Nuclear and Quantum Engineering at KAIST (Korea Advanced Institute of Science and Technology) since 1994. He has expertise in design and evaluation of radiation detectors and radiation imaging systems for medical and industrial applications. Particularly he contributed to the development of digital radiography detectors based on the amorphous silicon as well as the brain PET systems based on the silicon photomultiplier. Currently he is the President of the Korean Society of Radiation Industry.

Abstract:

We are proposing a new radiation scale, RAIN (radiation index) for the general public who often suffers from misunderstanding and unnecessary fear of the radiation and radiation release accidents. RAIN is defined in dimensionless such as the seismic scale. RAIN is defined to scale the individual accumulated radiation dose originating from an arbitrarily-defined particular event such as a single medical CT examination, an intake of contaminated foods or an exposure due to a certain incident for a certain period of time. The practical range of RAIN value varies from zero to 10. To maximize the simplicity for the general public, the numerical value of RAIN shall retain no more than a single significant digit after decimal point. The reference dose is selected as 10 µSv in a year in this paper, based on the exemption and clearance levels which were suggested by IAEA. Some example values of RAIN are 0.8 for a typical chest X-ray examination (0.07 mSv), 2.9 for a CT examination (7.4 mSv), 3.7 for the maximum annual effective dose limit of a radiation worker (50 mSv), 5.6 for the lethal dose LD50/60 (4000 mSv) etc. As a quick aid for the general public, the last column of Table 1 provides the three zones of RAIN values (Green, Yellow, and Red zones) which are characterized by the degree of severity of radiation exposure. We anticipate that this new definition of RAIN may serve the public and the experts to converse more correctly, easily and fruitfully.

Biography:

Danju Zhang has her expertise in silviculture, forest ecology, chemical ecology. She along with her team has focused on the short rotation Eucalyptus plantations for over 25 years. She identified the volatile organic compounds and water-soluble phenol allelochemicals and evaluate their physio-ecological effects on plant and soil biota.

Abstract:

Allelopathy has been identified as an underlying mechanism of detrimental environmental impacts within commercial plantations. Eucalyptus spp. is known to generate huge amounts of volatile organic compounds (VOCs) that can function as phytotoxins and thus inhibit other plants. In the present study, biochemical markers, including activities of acetylcholinesterase (AChE) and oxidative stress enzymes, such as superoxide dismutase (SOD) and glutathione S-transferase (GST), were assayed to assess changes in Eisenia fetida at the physiological level induced by different doses of VOCs as part of an acute toxicity test over 7 and 14-day exposures. In addition, the toxicities of VOCs were investigated using a soil avoidance test and comet assay. The results revealed that E. fetida exhibited significant avoidance behavior towards the highest concentrations of undecane, decane, 2,4-dimethyl heptane, and 2,2,4,6,6-pentametyl heptane. The tail DNA percentages were significantly increased for all experimental treatments relative to control. However, under the treatments of VOCs, Olive tail moment content and comet tail length also display an obvious increase compared to control, except for that of octane, undecane and decane treatments. As VOC concentrations and durations increased in the soil, activities of AChE, SOD, and GST were either stimulated or inhibited. Among the VOCs, decane, 2,4-dimethyl heptane, 2,2,4,6,6-pentamethyl heptane, and 2,4-di tert buyl phenol exerted stronger effects on enzymatic activities. In summary, VOCs in rhizosphere soils of E. grandis might exert a toxic impact on E. fetida, among which 2,4-dimethyl heptane, 2,2,4,6,6-pentamethyl heptane, and 2,4-di tert buyl phenol have the strongest effects. It was found that 4 and 8 year might be the important turning points for the dynamics of the allelochemicals. The effects of water-soluble allelochemicals were stronger than that of VOCs. At present, selection of the main effective phenol compounds through UPLC-MS and their physioecological effects are in progress.

Biography:

Vijay Jagdale is Veterinarian by profession and has expertise in Preclinical Toxicology as a Toxicologist as well as Pathologist working with various CRO’s and discovery/generic pharmaceutical companies. He has contribution in building strategic discovery platform for preclinical assays to facilitate smooth pathway for the lead candidates. He has expertise in the area of preclinical research in drug discovery and development in CROs with adequate exposure to preclinical toxicology, toxico-pathology, DMPK and laboratory animal facility set up, management and operation. Complete understanding of preclinical GLP toxicology with multiple types of therapeutics areas adhering to the various national and international regulatory guidelines. He also has experience in establishment, operation and management of national and international (AAALAC/OLAW) accredited rodent facilities with preclinical toxicology laboratory.

Abstract:

Developing a new drug from original idea to the launch of a finished product is a complex process where strategic planning plays crucial role to reduce usual time (12–15 years) as well as cost ( 1 billion $) for all the processes. The pharmaceutical industry has been evolving worldwide in the recent years, which made numerous CROs and biotech companies conducting such programs through services in various countries. The proper selection and applications of correct models, as well as appropriate data interpretation, are critically important in decision making and successful advancement of drug candidates. The successful discovery program includes strategic execution and read outs from initial assays like physicochemical characterization (kinetic solubility, thermodynamic solubility, pH dependent solubility and lipophilicity), absorption (PAMPA, MDCK permeability, Caca-2 permeability, MDCK-MDR1 permeability), metabolism (liver microsomes, hepatocytes, S9 fraction, reaction phenotyping, CYP inhibition and time dependent inhibition) and other preliminary assays (PPB, brain/tissue binding, blood to plasma ration, plasma stability, SGF/SIF stability, buffer stability, etc.) to prioritize leads for the further in-vivo assays. Pharmacokinetics assays needs appropriate execution to rank order compounds based on promising clearance (CL), bioavailability (F%), exposure (AUC), half-life (t_1/2), and distribution volume (L). Selective in-vivo PK studies are valuable to confirm whether the applied in vitro assays (in vitro metabolism and absorption) can serve as good predictive models for in vivo PK in terms of plasma clearance and bioavailability. The early application of preclinical safety assessment-both new molecular technologies as well as more established approaches such as standard repeat-dose rodent toxicology studies can identify predictable safety issues earlier in the testing paradigm. These earlier identification of dose-limiting toxicities will provide chemists and toxicologists the opportunity to characterize the dose-limiting toxicities, determine structure–toxicity relationships and minimize or circumvent adverse safety liabilities. Early stages includes nonclinical safety studies on candidate drugs to assess general toxicology (through in vivo experiments), safety pharmacology (effects on major organ systems) and basic genetic toxicity tests during early discovery phase in non-GLP conditions. Because of this, there is a strong need for personnel involved with toxicology and pharmacology studies need to understand the varied tools and approaches to perform early drug discovery safety analysis.

Biography:

Lucija Peterlin Mašič is an Associate Professor for Medicinal Chemistry and an Assistant Professor for Toxicological Chemistry. She has expertise in Medicinal Chemistry and Toxicology. She has studied the influence of metabolism on endocrine activities of hormone disrupting chemicals such as analogs of bisphenol A and novel brominated flame retardants with different in vitro assays. Her research interests are also in vitro studies of drug metabolism, testing compounds acting on nuclear receptors (estrogen, thyroid, androgen, glucocorticoid, PXR, FXR and PPARs), metabolism of xenobiotics and activity studies of their metabolites, bimolecular mechanism studies of toxicity, structure and ligand based drug design, etc.

Abstract:

Statement of the Problem: Structural analogs of bisphenol A (BPF, BPAF, BPS) are commonly used as its alternatives in industrial and commercial applications. Nevertheless, the question arises whether the use of other bisphenols is justified as replacements for bisphenol A in mass production of plastic materials. Knowledge about the metabolic pathways and enzymes involved in metabolic bio-transformations is essential for understanding and predicting mechanisms of toxicity. The activities on different nuclear receptors of the new brominated flame retardants 2-ethylhexyl 2,3,4,5-tetrabromobenzoate (TBB) and bis(2-ethylhexyl) 2,3,4,5-tetrabromophthalate (TBPH), and their main carboxylic acid metabolites 2,3,4,5-tetrabromobenzoic acid (TBBA) and mono(2-ethylhexyl) tetrabromophthalate (TBMEPH) were also investigated.

Methodology & Theoretical Orientation: The bisphenols metabolism using pooled liver and intestine microsomes, as well as recombinant human cytochromes (CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2E1, and CYP3A4) was tested using LC-MS/MS. For nuclear receptor testing in vitro systems were used to evaluate the estrogenic, androgenic, glucocorticoid, thyroid, PXR and PPARs activities of parent chemicals and their metabolites.

Conclusion & Significance: Once enter the body; the bisphenols are subjected to both oxidative metabolism and mainly conjugation. However, conjugation, which is mainly with glucuronic acid, is the predominant metabolic pathway for bisphenols and this is therefore considered to be an important mechanism for bisphenols detoxification. However, comprehensive testing of BPAF-glucuronide on all nuclear receptors has been performed and will be presented. On the other hand, it has been detected that metabolites of bisphenols and flame retardants have enhanced endocrine activities. For example, the estrogenic activities of MBP and hydroxycumyl alcohol are significantly higher than that of BPA. Even the formation of small amounts of those metabolites can significantly affect estrogenic activity. This was observed also for the novel brominated flame retardants TBB and TBPH, which showed weak or no activity on estrogen and androgen receptors, while their metabolites showed significant anti-estrogenic and anti-androgenic effects. Although conjugation is an extremely important detoxification pathway for bisphenols, it cannot completely eliminate their toxic effects in the body. Environmental estrogens can elicit their responses not only through binding to ERs, but also through numerous alternative pathways, and very low concentrations are needed for measurable effects. Despite their rapid glucuronidation, small amounts of bisphenols remain in their free form. As these can be further metabolized to their biologically active metabolites, these represent the risk for human health.

Biography:

Yassine Ait Bali is a Research Neurocientist at the Pharmacology, Neurobiology and Behavior Laboratory. His research interests have focused on the Neurotoxicology field. He is concentrating mainly on the relationship between pesticides side effects and the onset of brain diseases especially following developmental exposures covering all the aspects of neurological disease at different levels: Behavioral, morphological, physiological and molecular.

Abstract:

Statement of the Problem: The herbicide paraquat is a wide used product. Its role in the pathogenesis of some brain disorders has become intensely debated and gained increasing interest in recent years. The aim of our study is to investigate the developmental and neurobehavioral effects of prenatal exposure to this product in mice.

Material & Methodology: In the present work, we investigated the acute and developmental toxicity of PQ, from the 1^st or 6^th day of mating and throughout the gestation period. We have examined several parameters, including toxicity indices, reproductive performance and sensorimotor development, as well as anxiety and cognitive performance of the offspring.

Findings: Our results showed that exposure to 20 mg/kg of Paraquat during the first days of pregnancy completely prevent pregnancy in treated mice. Ingestion of a tolerable dose from the 6^th day of pregnancy caused an alteration in fertility and reproductive parameters and a decrease in litter size. In offspring, paraquat is responsible for a variety of behavioral disorders, manifested as an overall delay of innate reflexes and a deficit in motor development. All exposed animals showed a decrease in the level of locomotor activity, increased levels of anxiety and pronounced cognitive impairment in adulthood. Immunohistochemical studies have shown a decrease in the number of TH-immunoreactive neurons in the substantia nigra and intense glial proliferation in the hippocampus (using GFAP) in treated animals.

Conclusion & Significance: These results demonstrated that paraquat led to the onset of many behavioral changes that stem from the impairment of neuronal developmental processes in prenatally exposed mice.

Biography:

Jacqueline Chuah received her B.Sc (Hons) in Biological Sciences from the Nanyang Technological University, Singapore. In 2013 she joined Dr. Daniele Zink’s team at the Institute of Bioengineering and Nanotechnology of the Agency for Science, Technology and Research, Singapore. Dr. Zink’s lab develops organ-specific predictive screening technologies for in vitro toxicology/ nanotoxicology and conducts stem cell research with an emphasis on applications in predictive toxicology. Jacqueline’s work is focused on developing stem cell-based assays to predict kidney toxicity.

Abstract:

There is an increasing demand for alternative methods for the evaluation of compound toxicity in humans. Accepted alternative methods for the prediction of toxicities for human internal organs, such as liver and kidney, are not available. We have developed the first pre-validated platforms for the prediction of compound-induced nephrotoxicity in humans (Chuah and Zink, 2017; Kandasamy et al., 2015; Li et al., 2014; Li et al., 2013; Su et al., 2014; Su et al., 2016). These methods include the first high-throughput platform for the accurate prediction of nephrotoxicity (Su et al., 2016), as well as the first and only predictive stem cell-based methods (Li et al., 2014; Kandasamy et al., 2015). Our stem cell-based methods employ a rapid and robust one-step protocol for the differentiation of human induced pluripotent stem cells (iPSC) into renal proximal tubular cell (PTC)-like cells. PTC are a major target for compound-induced toxicity in the kidney due to their roles in compound transport and metabolism. iPSC-derived PTC-like cells have a purity of >90% after only 8 days of differentiation (Kandasamy et al., 2015). They can be directly applied for compound screening, as no cell purification is required. A method for the prediction of nephrotoxicity has been developed by combining use of iPSC-derived PTC-like cells with machine learning methods. This method has been pre-validated with 30 compounds and has a test balanced accuracy of 87% (Kandasamy et al. 2015). Furthermore, the underlying mechanisms of drug-induced cellular injury could also be correctly identified (Kandasamy et al., 2015). PTC-like cells are also suitable for applications in our high-throughput platform, which combines high-content imaging of renal cells with automated phenotypic profiling and machine learning methods (Su et al., 2016). This automated platform has been pre-validated with 44 compounds and has a test balanced accuracy ranging between 82% - 89%, depending on the human renal cell type used (Su et al., 2016). Based on these technologies we are currently developing a portfolio of platforms for the prediction of various organ-specific toxicities. In addition, we have established an organ-on-chip platform for repeated dose testing. This robust platform is based on simple and disposable chips and may be suitable for predicting quantitative parameters of the human dose response, such as No Observed Adverse Effect Levels.

Recent Publications

1. Chuah J K C, Zink D (2017) Stem cell-derived kidney cells and organoids: Recent breakthroughs and emerging applications. Biotechnology Advances 35: 150-167.

2. Kandasamy K, Chuah J K C, Su R, Huang P, Eng K G, Xiong S, Li Y, Chia C S, Loo L H, Zink D (2015) Prediction of drug-induced nephrotoxicity and injury mechanisms with human induced pluripotent stem cell-derived cells and machine learning methods. Scientific Reports 5: 12337.

3. Li Y, Kandasamy K, Chuah J K C, Lam Y N, Toh W S, Oo Z Y, Zink D (2014). Identification of nephrotoxic compounds with embryonic stem cell-derived human renal proximal tubular-like cells. Molecular Pharmaceutics 11: 1982-1990.

4. Su, R., Li, Y., Zink, D., and Loo, L.-H. (2014). Supervised prediction of drug-induced nephrotoxicity based on interleukin-6 and -8 expression levels. BMC Bioinformatics 15, S16.

5. Su R, Xiong S, Zink D, Loo L H (2016) High-throughput imaging-based nephrotoxicity prediction for xenobiotics with diverse chemical structures. Archives of Toxicology 90: 2793-2808.

Biography:

Vijay Jagdale is Veterinarian by profession and has expertise in Preclinical Toxicology as a Toxicologist as well as Pathologist working with various CRO’s and discovery/generic pharmaceutical companies. He has contribution in building strategic discovery platform for preclinical assays to facilitate smooth pathway for the lead candidates. He has expertise in the area of preclinical research in drug discovery and development in CROs with adequate exposure to preclinical toxicology, toxico-pathology, DMPK and laboratory animal facility set up, management and operation. Complete understanding of preclinical GLP toxicology with multiple types of therapeutics areas adhering to the various national and international regulatory guidelines. He also has experience in establishment, operation and management of national and international (AAALAC/OLAW) accredited rodent facilities with preclinical toxicology laboratory.

Abstract:

Historically, animals have been used in a wide range of scientific research activities that have provided many benefits to society, particularly in relation to the advancement of scientific knowledge, human and veterinary medicine, and the safety of chemical products. These principles seem to unify concerns for better science with causing less pain and distress to the experimental animals. All research with the Animal has the potential to cause pain, suffering, distress or lasting harm to the animals used. Most animals are killed at the end of experiments. Practical advances in the scientific methods can reduce areas of conflict. For this reason, the importance of the three R’s (refinement, reduction and replacement), and especially of the need to find replacements, cannot be overstated. The CCAC's ethic of animal experimentation is based on Smyth's definition of alternatives, i.e. replacement, reduction and refinement; replacement alternative methods which avoid or replace the use of animals. This includes both absolute replacements and relative replacements. Reduction alternatives refers to any strategy that results in fewer animals being used to obtain sufficient and precise data to answer the research question, without compromising animal welfare. Refinement alternatives refer to the modification of experimental procedures to minimize pain and distress. The 3Rs are rich in ambiguities and any implementation requires resolving the dilemma that promoting one R will sometimes directly or indirectly conflict with promoting another. The role of national and international accreditations have considerable impact on maintaining standard animal care and use program while designing animal experiments during discovery phase. IACUC/IAEC authority has responsibility to monitor the animal program as well as approvals of experimentation with ethical considerations. The ideas behind the 3Rs are so intuitively compelling that it is tempting to believe that full implementation is merely a matter of time, and once the 3Rs are widely implemented, the public will fully support any continued laboratory animal use that is deemed necessary in the biomedical research.