[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

Oxidative stress derived from various etiologies including environmental exposure, life-style changes, and systemic inflammation, can damage pancreatic β-cells, leading to the deficiency of insulin as type 1 diabetes, and also induce peripheral tissues as insulin resistance, leading to type 2 diabetes. Metabolic abnormalities in the body of individuals with diabetes cause mitochondrial superoxide overproduction that in turn activates multiple pathways: polyol pathway flux, increased formation of AGEs (advanced glycation end products), increased expression of the receptor for AGEs and its activating ligands, activation of protein kinase C isoforms, and overactivity of the hexosamine pathway, to generate excessive reactive oxygen or nitrogen species (ROSs or RNSs), which damages multiple organs, resulting in diabetic complications. Diabetic cardiomyopathy can occur independent of vascular disease, although the mechanisms are largely unknown. Current consensus is that the oxidative stress derived from metabolic syndrome causes cardiomyocyte abnormal gene expression, altered signal transduction, and the activation of pathways leading to programmed myocardial cell deaths. The resulting myocardial cell loss thus plays a critical role in the development of cardiac structural remodeling and dysfunction, “cardiomyopathy”. To support the above notion, our studies with in vitro and in vivo animal modes showed the prevention of diabetes and diabetic complications in the cardiomyocytes or transgenic mice with overexpression of antioxidant genes or supplementation of exogenous antioxidants. Among these clinical-translational antioxidants, metallothionein and its upstream nuclear transcriptional factor Nrf2 as well as their potent inducers have been received attention with greatly potential to be applied in clinics for patients with diabetes.

View full abstract

Methylmercury (MeHg) is a global pollutant that easily bioaccumulates in the marine food chain. This can be a detriment for human populations that rely on seafood as a principle source of nutrition since MeHg is a neurotoxin that disrupts brain development and function. The neurotoxicity of MeHg has been linked to its hydrophobicity and soft acid properties which instil it with the capacity to diffuse freely across the blood brain barrier and bind strongly to soft bases like sulfur on cysteine residues in proteins. Despite this, there seems to be a latency period between MeHg exposure and adverse neurological outcomes. This may be attributed to the gradual bioconversion of MeHg to inorganic mercury (IHg). Empirical data show that the reactive oxygen species (ROS) drive the bioconversion of MeHg to iHg. In particular, superoxide anion is the principle ROS responsible for catalyzing the demethylation of MeHg. This bioconversion mostly takes place inside the matrix of mitochondria which preferentially accumulates MeHg due to its high cysteine content. Studies using “omic” approaches have shown that MeHg disrupts mitochondrial function, in particular energy metabolism and antioxidant defense. With increasing awareness of environmental stewardship, incidence of acute MeHg poisoning from industrial pollution has become rare. However, the scale of chronic exposure to lower dose of MeHg as a result of global pollution or occupational hazard has grown. The talk will discuss the roles of toxicologists in the development of environmental management plan for MeHg in the local and global context.

View full abstract

Tanezumab is a monoclonal antibody (mAb) that binds to and inhibits the actions of nerve growth factor (NGF). Tanezumab is under clinical investigation for the treatment of pain associated with osteoarthritis, chronic low back pain, and cancer pain. On 14 December 2012, the U.S. Food and Drug (FDA) placed all anti-NGF mAb development programs on partial clinical hold due to adverse changes in the sympathetic nervous system of mature animals. The FDA indicated that to resume clinical studies, Pfizer must submit rigorous scientific data which characterize the sympathetic nervous system response to tanezumab and provide evidence that tanezumab may be used safely for the duration of the proposed clinical studies. Since the issuance of the partial clinical hold, Pfizer completed three studies in nonhuman primates to examine further the effects of tanezumab on the sympathetic nervous system. The results from these studies characterized the sympathetic nervous system response to tanezumab and clearly established that tanezumab does not cause neuronal cell death. In addition, these results show that exposure to tanezumab in non-human primates is associated with stereological changes in sympathetic ganglia, including smaller ganglion volume, smaller average neuron size/area, and lower estimated total neuron counts. These effects do not progress over time, are fully reversible, and are not associated with any adverse functional consequences. These data were submitted to the FDA in February 2015. In March 2015, FDA lifted the partial clinical hold on the tanezumab development program, allowing osteoarthritis and chronic low back pain studies with tanezumab to proceed.

View full abstract

Modern bio-imaging technologies like magnetic resonance imaging and spectroscopy (MRI/MRS) as well as some others have attained an important role in medical research due to low invasiveness and ability to provide functional information about biological systems in vivo. Such information could be obtained from the same subject repeatedly and with the least possible interference, which makes bio-imaging a unique and indispensable tool in toxicological and drug safety research. Most of bio-imaging modalities and techniques are intrinsically translatable – the same methodology can be applied pre-clinically in animals and in clinical settings. A great opportunity exists to leverage the advances in clinical imaging to improve the translatability of nonclinical safety assessments. Development of non-invasive imaging biomarkers of toxicity holds the promise to drastically improve contemporary safety assessment paradigms. This talk will provide the technical description of bio-imaging technologies and challenges of their use in the development of imaging biomarkers to support drug safety and other toxicological research settings.

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

Metallothioneins (MT) are ubiquitous low molecular weight cysteine-rich (30%) proteins with different biological roles in nutrition and toxicology. MT binds with both essential metals (Zn & Cu) and toxic metals (Cd & Hg). MT levels are high in organs of young mammals and proliferating epithelial cells, including certain cancer cells with nuclear localization. It can store Zn & Cu, and protect against toxicity of Cd & Hg. The expression of MT can be induced by certain metals and other factors. The high MT levels in metastatic cancer cells may be linked to the increased demand for metal ions in these cells. In addition, MT in cancer cells can protect against persistent oxidative stress. The high expression of MT has been associated with protection against DNA damage, oxidative stress and apoptosis. In cancer cells, the tumor suppressor gene p53 status influences the intracellular redox conditions and also affect the express of MT. The various protective mechanisms of MT will be discussed. (Rad. Res 148:235-39, 1977. J. Tr. Ele. Med. Biol 35:18-29, 2016).

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

That respiratory defense mechanisms are stimulated by oxidants/electrophiles has long been appreciated. Our work has focused on characterizing two such defense pathways from an inhalation toxicological perspective: 1) neuronal reflex responses, initiated by the transient receptor potential ankyrin 1 (TRPA1) receptor, and 2) cellular antioxidant defenses initiated via the nuclear factor erythroid 2-related factor (NRF2). TRPA1 and NRF2 are oxidant sensitive receptors expressed throughout the respiratory tract. They are activated by reactive inhaled agents (acrolein) or by local metabolic activation of inhaled or systemically delivered agents (naphthalene, acetaminophen). Exposure to inhaled and systemic oxidants in combination results in synergistic TRPA1 and NRF2 responses. Oxidant responses have also been characterized among respiratory tract regions and between electrophiles with differing reactivity. When normalized to delivered dose, inhaled oxidant/electrophile induction of pro-inflammatory genes is similar throughout the airways, however, differences exist in the NRF2-dependent antioxidant gene induction patterns between regions. Activation of both TRPA1 and NRF2 depends on toxicant interaction with sulfhydryl moieties. The oxidant exposure levels necessary for activation of TRPA1 and NRF2 responses are similar suggesting both oxidant sensors are of comparable sensitivity. While soft electrophiles, such as acrolein, react with sulfhydryls, hard electrophiles, such as diacetyl, do not. Unlike acrolein, inhaled diacetyl is only a weak activator of either neuronal reflexes or NRF2 pathways, suggesting the chemical electrophilic reactivity profiles of TRPA1 and NRF2 are comparable. Overall, the oxidant sensitive receptors TRPA1 and NRF2 demonstrate many toxicologically relevant similarities

View full abstract

Our body has ability to sense environmental insults and to activate cellular defense enzyme genes. Transcription factor Nrf2 is essential for the coordinated induction of cellular defense enzymes and protection of lung tissues through association with CNC-sMaf binding element (CsMBE or ARE/EpRE). This notion has been supported by experiments using animal models, showing that Nrf2-null mice are sensitive to a wide variety of toxic electrophiles and ROS. Keap1 acts as a subunit of ubiquitin-E3 ligase that degrades Nrf2 constitutively and as sensors for electrophilic and oxidative stresses, and covalent modifications of the cysteine residues abrogate the ubiquitin ligase activity. This system has been referred to as the Cysteine Code. The two-site recognition / hinge and latch model proposed for the Keap1-Nrf2 system describes the mechanism of nuclear accumulation of Nrf2 in a Cul3-Keap1 E3 ubiquitin ligase-dependent manner. We have verified this model through structure biology, mouse genetics, and human disease analyses. Many missense mutations have been identified in KEAP1 and NRF2 genes of human lung cancers. These mutations disrupt the KEAP1-NRF2 complex and result in constitutive activation of NRF2. Subsequently, elevated expression of NRF2 target genes confers advantages on the growth of cancer cells through the metabolic reprogramming. Thus, the Keap1-Nrf2 system opens a new avenue to the understanding of the signal transduction and regulatory processes underlying the stress response and cancer progression.

View full abstract

Epidemiological studies indicate a strong link between inhaled particulate matter (PM) and cardiovascular and neurological disorders. Additionally, there is evidence that certain gases, such as ozone, and mixed gas-PM exposures may have similar effect beyond the lung. Therefore, we have explored the role of serum components in driving cerebrovascular endothelial inflammatory responses and dysfunction that result from exposure to a wide array of PM and gas mixtures. Using multiwalled carbon nanotubes (MWCNT) via pulmonary delivery in mice, we observed acute deficits in blood brain barrier, activation of astrocytes, and induction of cortical CCL5 and interleukin-6 (IL6) mRNA that were abrogated by coadministration of a rho kinase inhibitor that improves endothelial barrier integrity. This outcome was replicated in a rat model of inhaled ozone, suggesting pathways common to both gases and PM. We further explored the bioactivity of serum from exposed rodents, in terms of the ability to induce inflammatory responses in cerebrovascular endothelial cells in vitro, and found that, again, both MWCNT and ozone exposures led to compositional changes that elicited transcriptional changes and surface receptor expression of inflammatory adhesion molecules, and also diminished regrowth of cerebrovascular endothelial cells. Serum proteomic analysis reveals <1,000 high confidence peaks induced by ozone and peptide sequencing suggests that many of these peaks reflect fragmented or oxidatively modified endogenous peptides, rather than synthesis or secretion of new proteins. Similar serum alterations were noted following exposure to MWCNT. Using myographic approaches, we subsequently confirmed that the fraction of serum components <10 kDa could reduce ex vivo vasorelaxation responses and that this serum bioactivity was absent in matrix metalloproteinase-9-deficient mice, suggesting a role for protease activity in generating pathological circulating constituents. These studies provide mechanistic evidence for a role for circulating components to drive the systemic vascular effects of inhaled pollutants, but further characterization of the serum compositional changes is needed to fully understand this phenomenon.

View full abstract

Toxicity of volatile organic compounds (VOCs) in indoor air, such as formaldehyde (FA), Xylene (Xy) and Paradichlorobenzene (pDB), at the levels of Sick House/Building Syndrome (SHS) is difficult to assess by the ordinary inhalation animal studies; histopathological endpoints are negative for toxicity at such concentration level. Here we applied our Percellome Toxicogenomics Project that had been launched to develop a comprehensive gene network for the mechanism-based predictive toxicology using time- and dose-dependent transcriptomic responses induced by a chemical. This Project was initiated to reinforce and eventually replace the “safety factor (uncertainty factor)” widely used for the extrapolation of experimental animal data to humans. For this purpose, a normalization method designated as “Percellome” is developed (BMC Genomics 7:64, 2006) to generate mRNA expression values in “copy numbers per one cell” from microarrays and Q-PCR. Over 100 chemicals have already been tested in this Project. Here, we report that the Percellome analysis is capable of predicting functional insults that might lead to chronic toxicity.
FA, Xy and pDB at concentrations 0.08, 0.20 and 0.04 ppm, respectively, close to the “Indicative indoor exposure value of SHS (MHLW, Japan)”, were applied to the C57BL/6J mice 22hr/day x 7 days inhalation-exposure protocol (4 concentrations x 4 time points, triplicate). Lung, liver and hippocampus were analyzed. Strong suppression of gene expression related to neuronal activity in hippocampus, i.e. the immediate early genes (IEGs) including Arc, Dusp1 and Fos was shown commonly among the three chemicals. Lung and liver Percellome analysis pointed out a candidate cytokine upstream of IEGs. Our finding may be considered as a first substantial data that would explain the indefinite or unidentified complaint in SHS patients. In addition, Percellome analysis of the orally exposed lung and liver to FA, Xy and pDB will be presented for further clarification of the inter-organ relationship.

View full abstract

With the requirement for electronic data standards scheduled to begin December 18, 2016, the pharma community is on the edge of a major change in the way data need to be collected and reported, as well as the way regulatory authorities will be able to analyze data. To go along with these changes are benefits from data warehouses and sophisticated data analysis tools that can offer immediate access to data in ways not currently feasible. As we approach the start of FDA's mandate for SEND, the quality of SEND implementation for submissions becomes critical. In this presentation, best practices for SEND implementation based on extensive FDA SEND submission experience will be shared. SEND Readiness in terms of the ability to use electronic raw data to automatically create SEND datasets will be reviewed, along with suggestions for protocol designs that best conform to the SEND model. RecommendatioNoncompliance to the standard can result in Refusal to File by FDA ns will be presented for SEND dataset review, both for accuracy and fitness of data for analysis. Updates from CDISC and PhUSE, including SENDIG 3.1 timelines and challenges, Define standard, and Study Data Reviewer's Guide (SDRG) content and format will be highlighted.

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract

[in Japanese]

View full abstract