Annual Meeting of the Japanese Society of Toxicology
Current issue
Displaying 1-50 of 555 articles from this issue
Invited Lecture
  • Nasir KHAN, Rohde CYNTHIA
    Session ID: IL
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Pharmaceutical R&D has played an extremely important role in improving human health, eradicating diseases, and in prolonging people’s lives. Discovering new medicines requires extensive collaborations among subject matter experts from various disciplines to advance molecules through the many stages of R&D assuring that the right targets and molecules are selected, and clinical development occurs in the appropriate patient population. Typically, it requires approximately 10 years to bring a molecule from benchtop to patients. However, special circumstances such as the emergence of SARS-CoV-2 at the end of 2019 (COVID-19) required the swift development of vaccines and drugs to address the pandemic in a timely manner. This success showcases the capability of modern molecular technologies and close collaborations among pharmaceutical industry, Regulators, Investigators, and patients to meet this challenge with the utmost speed and focus. The deep and diverse expertise from preclinical R&D disciplines were key in achieving these goals.

    Download PDF (3173K)
Special Lecture
  • Masashi YANAGISAWA
    Session ID: SL1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Sleep is a ubiquitous behavior in animals with a central nervous system. However, despite the fact that the executive neurocircuitry and neurochemistry for sleep/wake switching has been increasingly revealed in recent years, the fundamental mechanism for homeostatic regulation of sleep, as well as the neural substrate for "sleepiness" (sleep need), remains unknown. To crack open this black box, we have initiated a large-scale forward genetic screen of sleep/wake phenotype in mice based on true polysomnographic (EEG/EMG) measurements. We have so far screened >8,000 heterozygous ENU-mutagenized founders and established a number of pedigrees exhibiting heritable and specific sleep/wake abnormalities. By combining linkage analysis and the next-generation whole exome sequencing, we have molecularly identified and verified the causal mutation in several of these pedigrees. Biochemical and neurophysiological analyses of these mutations are underway. Since these dominant mutations cause strong phenotypic traits, we expect that the mutated genes will provide new insights into the elusive pathway regulating sleep/wakefulness. Indeed, through a systematic cross-comparison of the Sleepy mutants and sleep-deprived mice, we have recently found that the cumulative phosphorylation state of a specific set of mostly synaptic proteins may be the molecular substrate of sleep need.

    Download PDF (3014K)
  • Yuji IKEGAYA
    Session ID: SL2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Since 2018, I have been leading the ERATO Ikegaya Brain-AI Hybrid Project, which aims to combine machine learning with neuroscience research. Our project tackles complex questions, such as "What are the potential benefits of implanting an AI into the brain?", "How will the world change if our brains are connected to the internet?", and "What will be the impact on our minds if our brains are connected with each other?" We take these questions seriously and approach them with a sense of curiosity and wonder. For example, we have developed a chip with built-in information sensors that can be implanted into the brain. This chip can provide direct feedback to the brain about the environment and the body, such as changes in geomagnetism or blood pressure. By investigating how the brain responds to this new information, we hope to better understand how its abilities and behavior patterns can be enhanced.In addition, we are using AI to analyze information that the brain is sensing but not utilizing behaviorally. This research aims to extend the brain's function and determine if there is still room for optimization and evolution. These questions remain a mystery, but in my special lecture, I will discuss the state-of-the-art progress we have made in this field.

    Download PDF (3013K)
  • Tohru ISHITANI
    Session ID: SL3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    For preventing age-related decline and extending healthy life expectancy, preemptive medicine interfering the initial processes of aging and age-related diseases would be effective. To develop such medical approach, the processes priming aging and age-related diseases should be clarified. Our laboratory is tackling to unravel previously unidentified mechanisms of aging and cancer using small fish models and state-of-the-art technologies. Recently, by zebrafish imaging analyses, we discovered that a newly emerged precancerous cell with the RasG12V mutation in healthy epithelia are sensed and then eliminated by neighboring normal cells in an immune cell-independent manner. However, additional mutation of tumor suppressor gene or inflammation prevents the elimination of precancerous cells and then survived precancerous cells secrete inflammatory molecules that convert neighboring normal cells into either senescent or proliferative cells, generating a heterogeneous primary tumor. Thus, we revealed the novel mechanisms controlling the initial step of tumorigenesis (Nature Commun 2019; 2022). On the other hand, we are also tackling to clarify the fundamental mechanisms of systemic aging using an ultra-short-lived killifish Nothobranchius furzeri (N. furzeri) as a model. We successfully set up a large-scale breeding facility for N. furzeri and establish rapid reverse genetics methods for creating knockout and knock-in reporter N. furzeri (Sci Rep 2022). Utilizing these methods, we have identified several new factors that control systemic aging (in preparation). In this retreat, I would like to introduce these challenges and recent results.

    Download PDF (3025K)
  • Yasuhiro KAZUKI
    Session ID: SL4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    The development of vectors for expressing genes in mammalian cells and animals has not only been a tool for analyzing gene functions, but also has played an important role in industrial and medical applications. In the conventional transgenic technology, the DNA that can be introduced is usually limited to several hundred kb, and it has been impossible to introduce genes or gene clusters having a size exceeding 1 Mb. To solve these problems, we used chromosome engineering technology to develop human artificial chromosomes (HAC) and mouse artificial chromosomes (MAC) that can introduce large human genes, multiple human genes in a stable manner. We have succeeded in loading the human dystrophin gene (2.5Mb), the human drug-metabolizing enzyme gene cluster (1.5Mb), the human antibody genes (3.5Mb), etc. into the HAC/MAC vector. Furthermore, by introducing a "Designed Chromosome" into mice and rats, in which a large gene is loaded into HAC/MAC, "Designed Animals" were created. Using these animals, we have worked on drug metabolism and pharmacokinetic research, toxicity research, production of antibody drug candidates, etc. Furthermore, we are working on basic research and applied research by "Designed Cell" using "Designed Chromosome". In this presentation, I will introduce new drug discovery tools (fully human antibody-producing animal containing human Ig locus, mouse/rat (or human cell lines) expressing human drug metabolism, etc.) developed by HAC/MAC technology, and further introduce new combined technologies of DNA synthesis and HAC/MAC for biomedical research.

    Download PDF (3014K)
  • Masatoshi HAGIWARA
    Session ID: SL5
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    To realize the new therapeutics and rescue patients who suffer incurable diseases, we set up comprehensive drug development system in our Kyoto University, under the strategy to maximize the serendipity for the academic drug discovery with open innovation. In our medical school, everybody can access to our original chemical libraries to screen compounds with own ideas using powerful cell-based assay machines in our Drug Discovery Center. If somebody succeeds to find out a hopeful small molecule, he/she can analyze the mode of action in Sections of Genetic Information Analysis, Mass Spectrometry, and Bio-imaging, synthesize structurally-developed compounds in MedChem Support Section, and further test the compounds in animal facilities. Then he/she can consult stuffs of Institute for Advancement of Clinical and Translational Science (iACT) to accomplish the non-clinical animal tests and prepare protocols of the clinical trial in Kyoto University Hospital. Using this system, we succeeded to develop several“Academia Drugs” in the fields of viral infections, cancers, and chronic pain. Even for genetic diseases such as Familial dysautonomia, EDA-ID (anhidrotic ectodermal dysplasia with immunodeficiency), cardiac Fabry disease, and type V cystic fibrosis, we found therapeutic small molecules.

    Download PDF (3026K)
  • Toshihiko SHIROISHI
    Session ID: SL6
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    The laboratory mouse strains, such as C57BL/6, have mosaic genomes with different subspecific origins. Their genomes are derived predominantly from Western European subspecies Mus musculus domesticus, with the remaining sequence (6-7% of the whole genome) derived mostly from Japanese subspecies M. m. molossinus. We reported that genome introgression fromJapanese fancy mice, ancestor of extant JF1/Ms strain, provided the origin of the M. m. molossinus genome in the laboratory strains, and largely contributed to their genome diversity. According to old literatures, we infer that the JF1/Ms ancestry, raised at the end of the Edo period, was transferred to Europe, where they were bred with mice from Western European subspecies. From the breeding colonie the laboratory mice were established. The admixture of different subspecies has created genomic and phenotypic diversity among the laboratory strains used in biomedical science, and this should be kept in mind when designing experiments with the laboratory mouse strains.

    A similar mosaic structure is also found in genome of modern humans, in which a small portion of Neanderthal or Denisovan genome is interspersed in the genome of Homo sapiens. Interestingly, in both human and mouse, it is likely that the mosaic genome structure was not created at random, but formed under genetic constraints. The laboratory mice and JF1/Ms strain might provide unique research materials to clarify the genetic constrains, which had contributed to configuration of the mammalian mosaic genomes.

    Download PDF (3022K)
  • Akio KOIZUMI
    Session ID: SL7
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Pefluoroalkyl substances (PFASs) are synthetic chemical compounds with at least a perfluorinated methyl group (–CF3) or a perfluorinated methylene group (–CF2–) in the molecule. Approximately 5000 distinct PFASs are known. Those compounds are chemically stable and have been used in variety applications; i.e., Teflon emulsion vehicle, fire-fighting foams, surfactants and cosmetic foundations. Because they are stable in the environment, legacy PFAS, PFOS, PFOA and PFHxS have been registered as persistent organic pollutants (POPs) in the Stockholm Convention. Those three PFASs have very long biological half-lives in human ranging from 2y to 6y years and are not metabolized in the body. There have been several lines of epidemiological evidence that these PFASs impair lipid metabolisms, have immunotoxicity, retard fetal and neonatal growth and increase the risk of renal cell cancer. In my presentation, I would like to discuss the environmental problems in Japan and basic toxicology of legacy PFASs, PFOS and PFOA mainly based on our data. As for the basic toxicology information, I would like to show 1) the enterohepatic circulation of PFOS and PFOA, by which their biological half-lives are extended, 2) modification of hepatic toxicity by PPARα (peroxisome proliferator activated receptor alpha), 3) a candidate transporter and 4) the effects on Na and Ca channels. In closing, I would like to address importance of providing mechanistic insights in transporters, immunotoxicology and carcinogenesis to support epidemiological studies.

    Download PDF (3018K)
  • Takahiro OCHIYA
    Session ID: SL8
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Extracellular vesicles (EVs) are being developed as therapeutics including regenerative and immune-modulatory therapies, and drug delivery system (DDS). Especially, among several stem cells, mesenchymal stem cells-derived EVs (MSC-EVs) have been focused as therapeutics for variety of human diseases including liver fibrosis, lung infection, kidney disorders. For the development of therapeutics of such stem cell-derived EVs, toxicity evaluation is the important first step in the nonclinical phase to ensure the safety of EV therapeutics. The values of MSC-EVs have been verified in a variety of experimental animal models, but the toxicity evaluation is not adequately performed. The aim of this talk is to evaluate toxicological profiles with MSC-EVs and other stem cell-derived EVs in according to the regulatory science.

    Download PDF (3175K)
Educational Lecture
  • Maki FUKAMI
    Session ID: EL1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Recent advances in cytogenetic technologies have enabled to detection of various chromosomal alterations. In particular, sex chromosomes often develop numerical or structural changes. For example, the Y chromosome contains a region enriched by repetitive sequences (“the AZF region”). This region is a hotspot for microdeletions and duplications, and more than 30% of Japanese men have microdeletions in this region. Copy-number variants in this region are known as risk factors for azoospermia, although the Japanese-specific microdeletion does not affect spermatogenesis.

    Of note, recent studies have shown that Y chromosomes are often lost in the somatic cells of elderly people. In these men, 45,X cells gradually accumulate in the body. Reportedly, 45,X cells are present in about 40% of men aged 70 years. Age-related Y chromosome loss is known as a risk factor for early death, cancer, and Alzheimer's disease. We found that somatic Y-chromosomal loss is not limited to elderly men, but also occurs at any time of life. Mosaic Y chromosomal loss in children and young men is a risk for disorders of sex development, growth retardation, and infertility. The frequency of Y chromosomal loss is significantly higher in smokers than in non-smokers, indicating that environmental factors play an important role in this phenomenon. On the other hand, we have found that the Japanese-specific AZF deletion does not increase the risk of Y chromosomal loss.

    In my talk, I would like to introduce new findings on sex chromosomal variations and discuss possible roles of environmental factors in the development of such chromosomal changes.

    Download PDF (3019K)
  • Keiko TAGUCHI
    Session ID: EL2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Cancer formation needs “optimum soil” in cellular microenvironment. ”Cell Competition” is an interesting mechanism in which mutated cells are eliminated by the surrounding normal cells. Professor Emanuel Farber and his colleagues at University of Toronto, Canada, demonstrated that carcinogens bind to genomic DNA, which then create specific DNA adducts to form tumors. This is the Resistant Hepatocyte rat model (also called as Solt-Farber model) established in 1970’s. This “Chemical Carcinogenesis” model is a combination of diethylnitrosamine (DEN), a tumor initiator, a proliferation inhibitor AAF and 70% partial hepatectomy that enhances proliferation, resulting in liver tumors. Preneoplastic lesions emerged in such a severe environment are consisted of hepatocytes positive to glutathione S-transferase P (GSTP). Professor Amedeo Columbano at University of Cagliari, Italy, who was a postdoctoral researcher in Farber’s laboratory reported in 2010’s that somatic mutations in NRF2 accumulated in the GSTP-positive lesions. The gain-of function mutations in NRF2 confer cells an ability to activate NRF2 constitutively. GSTP is a target gene of a transcription factor NRF2. On the other hand, somatic mutations in NRF2 and KEAP1 that regulates NRF2 activation have been reported in human cancer cells since 2006. As NRF2-addicted cancers occupy approximately 30% in certain types of cancers, such as in lung and esophagus, NRF2-targeting drugs would be critically important for the future therapies specific to the patients with NRF2-addicted cancers.

    Download PDF (3020K)
  • Kazutaka IKEDA, Soichiro IDE
    Session ID: EL3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Addictive substances are diverse and have varying degrees of neurotoxicity. In the world, opioid abuse is a serious problem. For example, approximately 70,000 people per year die due to opioid overdoses in the United States. In Japan, abuse of methamphetamine, which is classified as a stimulant, is a major problem. Recently, abuse of marijuana and prescription drugs has expanded, especially among young people, including junior high school and high school students. In addition, the problem of excessive caffeine intake from energy drinks and other sources is becoming more serious, especially among young people. On the other hand, many addictive substances have useful effects such as stress relief, pain relief, sleep onset, and antidepressant effects. Therefore, there is a need to clarify the mechanisms of toxicity and usefulness, and to properly use and regulate these substances. In this educational lecture, first, the various addictive substances will be classified according to their target molecules, neuropsychiatric effects, and legal regulations, and the overall picture will be introduced. Second, the toxicity and usefulness of individual addictive substances, such as alcohol and methamphetamine, and their mechanisms will be explained in detail. Finally, the latest research trends on addiction, including not only substance dependence but also behavioral addiction, will be introduced.

    Download PDF (3177K)
Presidential Lecture
  • Satoshi KITAJIMA
    Session ID: PL
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    A paradigm shift is a revolutionary or dramatic change in a paradigm, i.e., a perception, idea, or social value system that were taken for granted in each given era or field. Paradigm is a concept originally proposed by Thomas Samuel Kuhn, a philosopher of science, in his main work "The Structure of Scientific Revolutions" (1962), but it has been expanded from its original meaning and generalized for use. Exceptional problems that cannot be explained by the old paradigm gradually accumulate, and as a result, the paradigm is plunged into crisis. Eventually, from among the ideas considered heretical, some emerge that are effective in solving problems, and then a new paradigm appears. This is how science has progressed.

    Noteworthy advances in toxicology are, in my opinion, voraciously absorbing the revolutionary progress that can be described as a paradigm shift in life science. To be precise, there shifts are from “Qualitative” to “Quantitative” study, from “Correlation” to “Causation” study, as well as from “Inductive inference of general rules” to “Deductive inference of the entire system based on genomes”. Another example is “Regulatory Science,” which contributes to adjusting to a desirable form for human and societal benefit. Toxicology, even more, also proactively contributes to the introduction and utilization of artificial intelligence (AI) technology by using deep learning. Furthermore, it seems that the realization of "crossing the barrier between functional science and morphology" by introducing molecular biology has been reached relatively early. How advanced and intriguing! —I know of no other discipline with such impressive features, other than toxicology.

    I sincerely hope that toxicology will continue to be an academic field that welcomes, rather than fears, the emergence of new paradigms.

    Download PDF (3179K)
Symposium 1: Current Situation and Prospects of Biomarker-based Strategies in Drug Development
  • Takayo UENO
    Session ID: S1-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    In the course of drug development, it’s important to find an effective dose in a defined group of patients with the fewest side effects. In recent years, it has become difficult to proceed with drug development without exploring the relationship between biomarkers representing genetic characteristics, environmental factors and detailed characteristics of diseases, and efficacy and safety. Utilization of biomarkers lead the shift change from a “one-drug-fits-all” to a “personalized approach”, placing the drug development industry in a highly dynamic landscape, having to navigate such disruptive trends. In response to this, innovative clinical trial designs have been key in realizing biomarker-driven drug development. Regulatory approvals of cancer genome sequencing panels and associated targeted therapies have brought personalized medicines to the clinic. Large-scale genome analysis technology and information processing technology are rapidly progressing in the medical field, which leads a global-wide transformation of the entire medical care. The advancement of technologies also promotes utilization of clinical biomarkers, and clinical biomarker-based medical innovation is currently taking place in various fields in drug development. The presentation will include an overview of biomarker-driven drug development and summarizes the current state of biomarker utilization in drug development. I will also touch on the development phases of clinical biomarkers divided into Discovery, Translational and Qualification.

    Download PDF (3020K)
  • Takuya FUJITA
    Session ID: S1-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Safety biomarkers are important drug development tool for monitoring and predicting the safety of new drug candidate compounds in nonclinical and clinical study. Although classical kidney injury biomarkers, such as serum creatinine and BUN continue to be utilized as primary renal markers in nonclinical study, they are considered poor indicators of early renal dysfunction due to limited sensitivity. Recently, FDA Biomarker Qualification Program qualified six urine biomarkers (KIM-1, clusterin, cystatin-C, N-acetyl-beta-D-glucosaminidase (NAG), neutrophil gelatinase-associagted lipocalin (NGAL), and osteopontin) to be used in standard measures of kidney function to detect early drug-induced renal tubular toxicity in phase I clinical trials. Thus, the safety biomarkers have the promise to accelerate and improve the drug development process. The urine biomarkers can be used for safety monitoring in clinical trials when nonclinical toxicology studies with a study drug demonstrate evidence of reversible histologic renal tubule damage that is associated with an elevation in any of the six urine biomarkers. However, urinary biomarkers sometimes show unexpected variability, which can complicate our understood. In this session, we will present the case of biomarker responses of drug-induced kidney toxicity in nonclinical study.

    Download PDF (3018K)
  • Yoshiro SAITO, Kosuke SAITO, Noriaki ARAKAWA
    Session ID: S1-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Ministry of Health, Labour and Welfare has been promoting predictive and preventive drug safety measures. Drug-induced interstitial pneumonia (DILD) is a severe adverse drug reaction that is frequently reported in Japanese. Among various types of DILD, the diffuse alveolar damage (DAD) type is particularly important to identify in early stage and initiate appropriate treatment, because it may cause death and sequelae. We collaborated with four university hospitals in Japan to search biomarkers of DILD.

    As a result, we found stratifin as a serum DAD marker that is elevated in the acute phase and decreased in the recovery phase, and was useful for discriminating DAD from other DILD types (such as organizing pneumonia and non-specific interstitial pneumonia) (Arakawa N. et al., Nat Commun. 2022;13:5854). These findings were successfully replicated in another cohort and was also useful for discriminating the DAD type of DILD from other related diseases (bacterial pneumonia, lung cancer, COPD, etc.). Note that employed analytical method was constructed and validated for the necessary parameters. Furthermore, the immunohistochemical analysis of autopsy samples revealed stratifin expression in the lung epithelium of DAD patients, and the results using cultured cells suggested p53- and apoptosis-dependent release of stratifin to the extracellular space. These results strongly suggested that stratifin is a useful DILD marker for DAD type. Separately, lysophosphatidylcholine was also found as a biomarker that could discriminate between DILD and idiopathic ILD (Saito K. et al., Sci Rep. 2022 ;12:19819). Based on these results, we received the pharmacogenomics and biomarker consultation of PMDA.

    In our presentation, after introducing the above examples, we would like to propose an appropriate strategy for establishing safety biomarkers.

    Download PDF (3030K)
  • Takasumi SHIMOMOTO
    Session ID: S1-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    While the use of biomarkers in drug development will lead to the realization of drugs with high therapeutic efficacy and few side effects, their use without the sufficient consideration may bring about wrong decisions. Therefore, it is essential to verify the qualification before the official use in drug development. Qualification is a conclusion that, within the stated context of use, the results of assessment with a biomarker can be relied upon to adequately reflect a biological process, response or event, and support use of the biomarker during drug development, ranging from discovery through post-approval (ICH E16).

    In 2009, Pharmaceuticals and Medical Devices Agency (PMDA) launched Omics Project Team to internally unify the way of thinking about biomarkers. The team has dealt with pharmacogenomics / biomarker consultation to evaluate and interpret data unrelated to particular pharmaceutical candidates, playing a role in the preparation of administrative notifications related to pharmacogenomics in cooperation with the Ministry of Health, Labour and Welfare (MHLW). Appropriately setting and revising the context of use will lead to the appropriate use and expansion of qualification, and the continuous and stepwise efforts will result in the realization of drugs with high therapeutic efficacy and few side effects guided by truly clinically meaningful biomarkers.

    In this presentation, I would like to introduce points to consider when considering drug development using safety biomarker candidates found in non-clinical studies.

    Download PDF (3014K)
Symposium 2: ToxicoEpigenetics as a leading edge of Toxicomics and the role of AI to its deployment
  • Masatoshi HAGIWARA
    Session ID: S2-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    In life-threatening scenarios, enhanced noradrenaline release induces endogenous analgesia through α2 adrenoceptors of the descending pain inhibitory system. Although α2-adrenergic agonists such as dexmedetomidine have analgesic and opioid-sparing effects, their use is restricted to hospital settings due to potential risks of acute hypertension/hypotension and bradycardia.Recently we found that a newly identified α2B subtype-specific antagonist, promotes noradrenaline release in murine spinal fluid and produces analgesic effects by stimulating the α2A-dependent pain inhibitory pathway. Orally administered ADRIANA has potent analgesic effects in several nociceptive pain models of mice and non-human primates. Genetic loss of the α2B adrenoceptor neutralizes the analgesic effect of ADRIANA in mice. As ADRIANA showed no significant adverse effects in non-clinical tests, we started a phase I/II clinical trial (jRCT2051220144) of the oral tablet of ADRIANA to examine the safety profile and the suppressive effect on postoperative pain in human.

    Download PDF (3013K)
  • Takuro NAKAMURA, Miwa TANAKA
    Session ID: S2-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Sarcomas are rare malignancies that exhibit diverse biological, genetic, morphological and clinical characteristics. Genetic alterations, such as gene fusions, mutations in transcriptional machinery components, histones, and DNA methylation regulatory molecules, play an essential role in sarcomagenesis. These mutations induce and/or cooperate with specific epigenetic aberrations required for the growth and maintenance of sarcomas. Studies using the mouse models for human sarcomas have demonstrated major advances in our understanding the developmental processes as well as tumor microenvironment of sarcomas. We will present our mouse models for fusion gene-associated sarcoma and its advantages. Recent technological progresses in epigenome editing will not only improve the studies using animal models but also provide a direct clue for epigenetic therapies. Potential applications of CRISPR/dCas9-based epigenetic editing in sarcoma studies and therapeutics will be introduced.

    Download PDF (3013K)
  • Jun KANNO, Ken-ichi AISAKI, Ryuichi ONO, Satoshi KITAJIMA
    Session ID: S2-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    We developed a new repeated dosing protocol; 14-day repeated dosing to all mice followed by a Percellome Project protocol study that monitors gene expression at 2, 4, 8, and 24 hours. The effect of repeated dosing can be decomposed into two elements, Transient Response (TR) seen within 24 hours after the last exposure, and the Basal Response (BR) as a drift in the basal expression due to repeated dosing. To decipher the BR, DNA methylation and histone modification were comprehensively monitored by WGBS and ChIP-seq against H3K4me3, H3K27me3, H3K27Ac, and H3K9me3. In case of clofibrate (CFB), a known PPARa activator, the repeated dosing has affected the TR and BR of many genes. For example, the TR of pdk4 and cyp4a14 had decreased and analysis on their enhancer-promoter region suggested that it can be explained by a decrease in PPARa signaling.

    Here, we performed a 4-day new repeated dosing study on perfluorooctanic acid (PFOA). Firstly, the 24-hour no-effect dose (used for dose setting of Percellome Project) of single dose of PFOA was 10mg/kg whereas that of 4-day dosing was 0.1mg/kg (CFB was 100mg/kg and 70mg/kg respectively). Pdk4 gene, known to be located downstream of PPARa, was gradually induced towards 24 hours after the last dosing, and cyp4a14 showed increase in both TR and BR, suggesting difference from CFB. Further analysis on the difference based on the network description will be presented with an attempt to look in to regulatory mechanisms by referring to the epigenetic information of CFB and other examples in our database. (Research Grant of MHLW)

    Download PDF (3017K)
  • Yayoi NATSUME-KITATANI, Ken-ichi AISAKI, Satoshi KITAJIMA, Jun KANNO
    Session ID: S2-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    The Percellome database [1], which enables quantitative comparisons of gene expression profiles (microarray data) induced by toxic chemicals by the process of estimating the number of mRNA copies per cell, is a useful resource for inferring the molecular mechanisms of toxicity caused by chemical exposure. The database contains gene expression levels (the number of mRNA copies per cell estimated by the above process) for each exposure dose and time in mice. By quantitatively capturing the dynamic changes in gene expression, it is possible to extract "what kind of molecular network changes can be linked to toxicity expression due to chemical exposure. We selected known PPAR alpha (Peroxisome Proliferator Activated Receptor Alpha) ligands and chemicals that have been implied to be PPAR alpha ligands based on our previous research results (clofibrate, valproic acid, estragole, DEHP (di(2-ethylhexyl)phthalate) and PB (phenobarbital)) for our analysis. We compared the patterns of dynamic changes in gene expression levels of these five chemicals, and detected common and unique patterns among them. In this presentation, a wide variety of biological responses induced by thesechemical exposures will be reported and useful analytical tools will be introduced.

    [1] Kanno J. et al., J. Toxicol. Sci. 2013;38(4): 643-654

    Download PDF (3016K)
  • Samik GHOSH
    Session ID: S2-5
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    The science of toxicology forms the cornerstone of modern medicine and healthcare. In toxicology, computational methods from omics-level data analysis to multi-level (cell, tissue, organ, and whole body) modeling and simulation techniques have been increasingly applied. New approaches in Analytical AI based on deep neural networks and representation learning hold further promise to enhance the field, providing the ability to query multiple modalities of data (quantitative, imaging, text processing) and higher predictive quality of results.

    However, gaps exist in the adoption of such cutting-edge methods in modern machine learning by practicing toxicologists, where descriptor-based quantitative structure–activity relationship (QSAR) methods are still widely used in this field. With the exponential progress witnessed in the field of AI, and more recently the rise of Generative AI and large language models (LLMs), it is critical to Review, Re-assess, and Re-imagine the role of AI in toxicology.

    In this talk, we highlight, through different case studies, the various applications of traditional and modern machine learning techniques in toxicology - ranging from large-scale risk assessment of specific compounds to the prediction of the impact of combination trial ingredients in cosmetology. Modern methods are typically data-hungry and require training on many examples. We also explore how emerging methods in generative AI can be developed to fine-tune existing large, pre-trained models on a large corpus of multi-modal data and applied to focused problems in toxicology.

    Novel modeling architectures which leverage existing computational methods, together with modern neural networks, representational learning methods, and large language models hold significant promise to re-imagine the science of toxicology - from specific drug ADMEtox properties to whole body risk assessment for human health and ecotoxicity.

    Download PDF (3027K)
Symposium 3: Biometals Specialty Section Symposium - 50 years of metal toxicology and expectations for the next 50 years
  • Toshiyuki KAJI
    Session ID: S3-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    “Deductive toxicology" is a deductive approach to explain the toxicity of chemical substances to humans by collecting reproducible scientific findings on the toxicity. In accidental or incidental poisoning cases, it is impossible or extremely difficult to reproduce the exposure conditions, and "deductive toxicology" is effective in such cases. Based on this thinking method, we worked on the environmental toxicology of cadmium, lead, and methylmercury. Additionally, we pioneered "vascular toxicology" and obtained new findings useful for understanding not only vascular lesions but also organ toxicity of cadmium, lead, and methylmercury. On the other hand, we have conceived of a new molecular probe that takes advantage of the strong bioactivity of metals to analyze their biological functions. This is a new research strategy, bioorganometallics, utilizing organic-inorganic hybrid molecules. We started by investigating the cytotoxicity of organic-inorganic hybrid molecules to clarify their properties and mechanisms. Then, using the organic-inorganic hybrid molecules as molecular probes, we analyzed the mechanism underlying metallothionein induction in vascular endothelial cells, the mechanism of regulation of proteoglycan synthesis in endothelial cells, and the mechanism of synthesis of supersulfides in endothelial cells. I would like to confirm the development of the above studies, the new findings obtained, and the importance of free development of toxicological research, thereby contributing to the future of toxicology in some small way.

    Download PDF (3014K)
  • Akira NAGANUMA
    Session ID: S3-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Minamata disease was first discovered in Minamata, Japan in 1956. It was caused by methylmercury discharged from a factory to Minamata Bay. The history of Minamata disease will be introduced in this presentation.

    Download PDF (3011K)
  • Seiichiro HIMENO
    Session ID: S3-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Selenium (Se) has triple faces as an essential, toxic, and pharmacological element. The dose-response relationships of these three characteristics fall in a narrow range. Dietary Se intake levels vary largely between nations and areas, depending on soil and crop Se concentrations. Humans and domestic animals have experienced Se deficiency and poisoning due to low and high Se dietary intake, respectively. In the U.S., Se poisoning among domestic animals occurred in South Dakota, but the north to the central area, including South Dakota, is the main production zone for floor and corn, resulting in higher Se intake among the U.S. population. A part of China and Nordic countries showed low Se intake levels. Most epidemiological studies showing the benefits of Se, including its anticancer effects, have been reported from these areas. The cancer chemoprevention trial by Se supplementation failed to prevent prostate cancer but increased the incidence of diabetes. The anticancer activity of Se could be attributed to its pharmacological activities, which occur at concentrations close to toxic doses, rather than to its antioxidant activities. Se supplementation to Se-deficient humans results in elevated antioxidant enzyme activities. However, after Se-dependent antioxidant activity reaches the plateau level, further Se supplementation would not increase antioxidant activity, and in some cases, it would cause adverse effects of Se, such as diabetes. Misunderstanding these points would cause unnecessary, sometimes harmful, Se supplementation.

    Download PDF (3015K)
  • Masahiko SATOH
    Session ID: S3-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Cadmium (Cd), a harmful metal, causes the damages in the kidney, liver, bone, respiratory and circulatory. In particular, chronic renal toxicity, whose main symptom is renal tubular dysfunction, is subject to the safety evaluation of Cd. The chronic renal toxicity occurred in many Cd-polluted areas of Japan. The itai-itai disease, whose main symptoms are renal toxicity and osteomalacia, has occurred in elderly multiparous women as a result of long-term oral intake of Cd due to environmental pollution of Cd in the Jinzu River area in Toyama. With the itai-itai disease as an opportunity, the evaluation of health risk of Cd has been studied, and further research has been carried out to elucidate the mechanism of Cd toxicity. Recently, Cd poisoning due to industrial workplaces and environmental pollution has hardly occurred in Japan. However, the health effects of long-term exposure to trace amounts of Cd have become a problem for the elderly, since Cd is taken into the body through foods such as rice over the course of a lifetime, and its persistence in the body is high. On the other hand, metallothionein (MT) was isolated as a Cd-binding protein from the renal cortex of horses in 1957. Since MT contains a large amount of cysteine, it has a high affinity for metals and has a free radical scavenging effect. Previous studies have clarified that MT is involved in the maintenance of homeostasis of essential metals such as copper and zinc, and plays a protective role against the toxicity of various harmful factors such as Cd and various diseases caused by oxidative stress. In this symposium, it will be introduced the findings on the toxicity of Cd and the biological defense of MT, and also referred to future issues.

    Download PDF (3015K)
  • Hiroyoshi FUJITA, Chie SUGIMOTO, Hiroshi WAKAO
    Session ID: S3-5
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    In 1970s, marked inhibition of 5-aminolevulinate dehydratase (ALAD) was well known as the specific and sensitive indicator of lead., resulting hypochromic anemia. Replacement of zinc by lead inhibited purified ALAD, which was reactivated by reconstruction of zinc binding to essential SH residues. It was showen that almost 90% of ALAD activity in vivo was determined by molar ratios among ALAD, and lead as well as zinc and GSH in all tissues examined. The result agreed with observations supporting absence of isozyme.Generating trichloroethylene-epoxide also efficiently inhibited ALAD, without any reactivation. Hepatic ALAD inhibition decreased either CYP and heme saturation of tryptophan pyrrolase, showing deficient free heme pool. Furthermore, hepatic 5-aminolevulinate synthase (ALAS), the rate-limiting step, was markedly induced, suggesting a negative feedback. Erythroid ALAS, immunochemically isozyme of hepatic ALAS, showed a positive regulation by heme, a part of which was mediated by Bach-1. These results were in good agreement with a difference between hepatic and erythropoietic porphyria, i.e. presence and absent of acute neuro-onset. Based on studies by others as well as ours, Japanese Government approved hemin as orphan drug for hepatic porphyria, in 2011.In 1990s, past exposure of asbestos and coming increase of mesothelioma became one of the most sever health problems in Japan. One of members in our laboratory (H. W.) succeeded to prevent carcinogenesis by NKT cells in mice, hence we started to develop similar method in human on 2006. Mucosal associated invariant T (MAIT) cells whose characteristics resembles NKT cells were abundant in human. By 2013, human MAIT cells derived from iPS cells (reMAIT) indicated ability to prevent infection of mycobacteria. Successive studies reveal preventive activity of reMAIT cells on cancer in 2022.

    Download PDF (3020K)
Symposium 4: Microglial Toxicology
  • Gi-Wook HWANG
    Session ID: S4-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Methylmercury is one of the most common environmental contaminants causing neurological disorders in the brain. In recent years, fetal health hazards due to exposure of pregnant women to methylmercury via fish diets are a worldwide concern. However, the mechanisms involved in methylmercury-induced neurological damage remain largely unknown. We have previously shown that methylmercury induces cell death via induction of expression of inflammatory cytokines such as oncostatin M (OSM), TNF-α, IL-1β, and CCL2, using mouse neural stem cell lines. We also found that the above proinflammatory cytokines are induced in microglia in the brains of mice treated with methylmercury. Moreover, OSM released from microglia induces cell death by binding to the extracellular domain of TNF receptor 3 (TNFR3) on neuronal cell membranes, and that this process is promoted by the direct binding of methylmercury to the 105th cysteine residue in OSM. Recently, we have shown that treatment of mouse brain slices with methylmercury induces neuronal cell death, which is inhibited by treatment with liposome encapsulated clodronate that selectively kill microglia. Mice exposed to methylmercury also showed impaired memory function, but this impairment was also suppressed by simultaneous administration of PLX3397, a microglia inhibitor. As described above, it has been suggested that methylmercury may damage surrounding neurons through induction of expression and production of inflammatory cytokines by activating microglia. This talk will introduce the role of microglia in methylmercury-induced neurological damage in the brain.

    Download PDF (3020K)
  • Ikuko MIYAZAKI, Masato ASANUMA
    Session ID: S4-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Parkinson’s disease (PD) is a progressive neurodegenerative disease with motor symptoms, such as akinesia/bradykinesia, tremor, rigidity, and postural instability, due to a loss of nigrostriatal dopaminergic neurons. Although various pathogenic factors, such as oxidative stress, neuroinflammation, mitochondrial impairment and α-synuclein toxicity, are indicated, the mechanism of neurodegeneration in PD remains unknown. Glial cells play pivotal role in maintenance of neuronal environment in the central nervous system by energy supply and immune defense; however, glial cells promote neuroinflammation followed by neurotoxicity. Recently, it is reported that activated microglia convert astrocytes to neurotoxic A1phenotype. Thus, contribution of astrocyte-microglia crosstalk has been received attention. Furthermore, glial cells exhibit not only morphological but also functional diversity depending on brain region. We have demonstrated that exposure to the pesticide rotenone, an environmental risk factor in PD, induced brain region-specific glial dysfunction mediated by astrocyte-microglia interactions and caused non-cell autonomous dopaminergic neurodegeneration. Furthermore, we found secreted molecules from microglia, which were treated with conditioned media from rotenone-treated astrocytes, induced dopaminergic neuronal death. In this symposium, we will outline recent findings on astrocyte-microglia crosstalk in PD pathology and would like to discuss the investigation on the mechanism of PD pathogenesis focusing on the regional specificity of glial cells.

    Download PDF (3015K)
  • Yasuhiro ISHIHARA
    Session ID: S4-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Recently, over 100,000 of chemicals are industrially produced and widely used in medical and industrial fields, and even our daily life. However, several types of chemicals can cause behavioral abnormalities after growth due to exposure during the developmental period from fetal to infancy. Taking an antiepileptic drug, valproic acid during pregnancy reportedly reduces intelligence quotient and increases autism risk in children after growth. Prenatal exposure to polychlorinated biphenyls causes postnatal memory and learning deficits and abnormal social behavior. Some heavy metals and pesticides are also recognized to have similar effects. Microglia, immune cells in the brain, selectively engulf unnecessary synapses during development to enhance the maturation of neural networks. On the other hand, excessive activation of microglia is considered to induce neuroinflammation and act destructively on the central nervous system. Recently, the relationship between abnormal microglial activity and neurological and psychiatric disorders has been actively discussed, and various chemicals such as heavy metals and polycyclic aromatic hydrocarbons were reported to activate microglia. In this presentation, I will show the results focusing on the interaction of activated microglia and neural network defects caused by prenatal valproic acid exposure, and also introduce the neurological effects of a pesticide neonicotinoid, and an air pollutant. PM2.5 when espousing to them during development.

    Download PDF (3020K)
  • Ryuta KOYAMA
    Session ID: S4-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Microglia are brain-resident macrophages that prune synapses through "find me" and "eat me" signals, which are fundamental to neural circuit reorganization. While Complement C1q has been identified as a "find me" and "eat me" signal for microglia, it can be randomly distributed in the brain parenchyma under certain circumstances, such as neonatal seizures. To investigate the mechanisms by which microglia determine which synapses to phagocytose after neonatal seizures, a live imaging system for microglia-synapse interactions was established in vitro. We found that microglia contacted and engulfed more active synapses when the neurons were activated. In a mouse model of neonatal febrile seizures, inhibitory, but not excitatory, synapses were pruned by microglia, resulting in hyperactivity of dentate neuronal circuits. We also discovered that the increased activity of dentate inhibitory neurons during hyperthermia-induced seizures resulted in preferential interactions between microglia and inhibitory synapses. These findings suggest that C1q may serve as an "eat-me" and increased neuronal activity as a "find-me" signal in the process of synaptic pruning by microglia, which may underlie the development of epilepsy following neonatal seizures.

    Download PDF (3020K)
Symposium 5: Current status and future perspectives of New Approach Methodologies (NAMs) in Next Generation Risk Assessment (NGRA)
  • Takashi YAMADA
    Session ID: S5-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Next Generation Risk Assessment (NGRA) is supported by New Approach Methodology (NAM), which consists of various in vitro and in silico approaches, and their integrated use is expected to improve the confidence of human safety assessment of chemical substances. A varieties of case studies have been recently developed for proof-of-concept of NGRA. Case studies are useful for understanding the strengths and limitations of NAM across different decision contexts. Read-across (RAx), a method of filling a data gap for a target substance using existing data of similar substances, has been applied for many years in the screening-level risk assessment of industrial chemicals. By building the scientific confidence using NAM, RAx can become one of the foundations of the NGRA. This presentation will summarize the challenges to improving confidence in RAx and trends toward the use of NAM. For regulatory acceptance of RAx prediction results, it is necessary to scientifically justify the similarity hypothesis and provide the supporting information. It is required to verify how in vitro NAMs related to endpoints can improve the approach of analogue selection. Importance of these issues identified in case studies will be presented. In addition, the OECD (Q)SAR Assessment Framework is a systematic and harmonized framework for the regulatory assessment of (Q)SAR models and results based on single or multiple predictions. This guidance is useful for increasing confidence in NGRA when applying in silico NAMs. An overview of the guidance in the regulatory context will also be introduced.

    Download PDF (3015K)
  • Masashi HORIE
    Session ID: S5-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Approaches to evaluation for chemical properties efficiently have been developed internationally after the Strategic Approach to International Chemicals Management (SAICM) was adopted by United Nations Environment Programme (UNEP) in 2006. SAICM provides a policy framework to guide efforts to achieve the Johannesburg Plan of Implementation goal that, by 2020, chemicals will be produced and used in ways that minimize significant adverse impacts on the- environment and human health. The New Approach Methodologies (NAMs) are the technologies that have a great potential to increase quality and efficiency in chemical assessment. However, there remain difficulties to introduce NAMs into regulations. One of chemical management laws in Japan is Chemical Substances Control Law (CSCL). Some of NAMs such as quantitative structure-activity relationships (QSARs) model and read-across are utilized in evaluation for new chemicals and risk assessment for existing chemicals under CSCL. In addition, introduction of weight of evidence approaches under CSCL is being investigated. Internationally, application of NAMs is being discussed at OECD which is the center of the international discussions on the NAMs. Japan contributes to the discussion by providing the results of investigation in Japan. However, there are major challenges and necessary future efforts to expand the application of NAMs to regulations, such as improving reliability, accumulating practical use cases, and creating guidelines regarding NAMs.

    Download PDF (3014K)
  • Taku NISHIJO
    Session ID: S5-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Skin sensitization risk is an important toxicological endpoint as part of safety assessment of chemical substances. In place of conventional animal-based testing methods for identifying and characterizing skin sensitization hazards, New Approach Methodologies (NAMs) have been developed for risk assessment of chemical substances, and the Next Generation Risk Assessment (NGRA) using these methods is being actively developed.

    Several in chemico and in vitro testing methods have been developed for skin sensitization NAMs, focusing on key events in the early stages of sensitization, and have been adopted as OECD test guidelines. In addition, several in silico tools have been developed. More recently, the Defined Approaches for Skin Sensitization (DASS) have been proposed to cover the inherent complexity of processes underlying skin sensitization with high predictivity. Currently, the 2 out of 3 Defined Approach and Integrated Testing Strategy v1/v2 Defined Approaches are included in the OECD guidelines. This presentation will introduce these NAMs for skin sensitization, focusing on those newly adopted or updated in the OECD guidelines, and discuss their current status of regulatory use in the evaluation of skin sensitization of chemicals and cosmetic ingredients.

    In using NAMs as part of NGRA for skin sensitization, there are some points to be considered and cautions to be taken into account, such as the applicability domains or limitations of each NAMs. In this presentation, the issues of NAMs for the future development of NGRAs and the latest research topics that are currently being actively studied to solve these issues will be introduced.

    Download PDF (3019K)
  • Kouichi YOSHINARI
    Session ID: S5-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    There is a strong demand for the development of alternative methods to animal experiments in the safety evaluation of chemical substances, but it remains very difficult to develop such a method for repeated-dose toxicity and carcinogenicity tests. A method called read-across, which utilizes existing toxicity information, has been thus attracting attention. In this method, the toxicity of a target substance is expected from the toxicity information of “source substances” that have similar chemical and/or biological properties to the target substance. Therefore, the selection of appropriate source substances is very important, but the similarity judgment tends to be subjective, and the problem of reproducibility has been pointed out. Therefore, we are aiming to develop an objective read-across method to replace rat repeated-dose toxicity and rat carcinogenicity tests. In our studies, read-across is divided into several steps: For example, 1) definition of data set with substances having in vivo toxicity data, 2) selection of test substances, 3) determination of inter-substance distances using molecular descriptors, 4) selection of source substances based on the distance, 5) additional selection of source substances based on in vitro test results, and 6) toxicity evaluation of test substances by read-across. In this session, I will introduce our research strategy and recent findings and would like to discuss the (regulatory) science of read-across for the safety evaluation of chemical substances.

    Download PDF (3014K)
  • Shota UENO
    Session ID: S5-5
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    International Life Sciences Institute Japan (ILSI Japan) launched the Alternative Animal Testing Promotion (AAT) project in food sector (AAT project) in 2018. The project aims to establish more predictive evaluation approaches for humans that do not rely on animals, and to advance the sophistication and efficiency of food safety and functionality evaluation. In this project, food-related companies are participating and collaborating with academia to promote research to accelerate alternative approaches, to collect information on the latest technologies, and to disseminate information to build consensus. As research themes, we are working on the construction of a toxicity database of food ingredients for toxicity prediction by effectively utilizing existing toxicity test information, the construction of a technology for predicting food exposure in the body, and the development of animal-independent safety assessment strategies and case reports at the same time. In this session, we will present the current activities and challenges of the ILSI Japan AAT project on NAM-based assessment strategies in the food industry.

    Download PDF (3013K)
Symposium 6: Approach to Safety assessment of Extractables and Leachables in pharmaceutical products
  • Masayuki MISHIMA
    Session ID: S6-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Revised Food Sanitation Act enforced in 2020 restricts food contact materials to those in the positive list supported by safety information. No similar list is available for pharmaceutical products at present. A pharmaceutical product often needs its own packaging suitable for the dosage and administration. It is difficult to apply a blanket restriction to various pharmaceutical products. Contact surface arias of pharmaceuticals and containers are usually much smaller than those of foods, it is reasonable to think that contaminants from containers are less in pharmaceuticals than food. However, there is no doubt that patients are exposed to the chemicals that unintendedly come from production instruments and containers of pharmaceutical products. A strongly toxic chemical in container closure systems is a big issue that exists with ordinary products. Lubbers used for injection formulations may contains a nitrosamine polymerization agent. Nitrocellulose in blister package may react with amines in printing ink to form nitrosamines. Lubbers and plastics include various additives to give batter plasticity and durability, details of which are not disclosed to users. Toxicologists have to estimate and assess leachables based on information of extractables that are detected under controlled extraction conditions. There are many issues e.g. we can find no safety information of a leachable. Here, I’d like to discuss background of requirement of safety assessment on extractables and leachables, and how toxicologists can approach to the issues.

    Download PDF (3019K)
  • Akihiko HIROSE
    Session ID: S6-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    In the risk assessment of impurities in pharmaceutical products, a basic approach is to identify and quantify each impurity, and to assess the toxicity of each impurity substance, and establish quality standards based on acceptable intake levels such as PDE. In addition to side reactants and degradation products, impurities in raw materials and excipients, the leachables from container closure systems or equipment used in manufacturing may also be included as impurities in final products. In particular, most of leachables are industrial chemicals, for which little toxicity information is available, it is difficult to assess the toxicity of individual substances. Therefore, it is necessary to manage risks by setting control thresholds based on the TTC (Threshold of Toxicological Concern) approach, which enables comprehensive risk management for small amount of chemical exposure. The TTC approach has been applied not only to the threshold setting for genotoxic carcinogens, but also to the threshold setting for non-genotoxicity. The original non-genotoxic TTCs have been adopted in the evaluation of food flavors, and extended to apply the assessment of leachables from food containers and apparatus. However, the toxicity information used in threshold settings for such non-genotoxic effects is mostly based on toxicity results from oral exposure studies. Therefore, threshold settings essentially apply only to impurities in oral drug formulations. On the other hand, parenteral drug products are considered to have a higher risk of exposure to leachables from container closure systems and manufacturing facilities than oral drug products. I would like to discuss how toxicity data from oral exposure experiments should be applied to the assessment of extractables and leachables from intravenous administered drug products and inhaled drug products etc.

    Download PDF (3015K)
  • Kazusei KOMATSU
    Session ID: S6-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    The ICH Q3E guideline is under development to promote international harmonization regarding the safety assessment and control of E&L, with reference to several documents that have been presented mainly by authorities and industries in Europe and the United States.

    Step 2b draft guideline for consultation is expected to be available in November 2023.

    Although, related guidelines in Japan have now been limited, it would be better to initiate preparation for future recommendations.

    Current E&L assessment and control processes generally include: 1) obtaining information for Material and Component selection, 2) gathering structure and safety assessment information for substances from Extractables studies, and 3) testing and corresponding control of leachables.

    Meanwhile, according to a part of the Concept Paper of the ICH Q3E guideline, there are at least following considerations as well as harmonization expectations related to quality control: 1) process and address material and component selection and characterization, 2) the conduct of E(&L) studies, 3) the design of an E&L control strategy based on science and risk-based principles.

    In this presentation, such points are planned to be picked up and explained to the extent possible from the scene of guideline development.

    Download PDF (3013K)
  • Yusuke NOMURA, Yusuke OKAMOTO, Chie HASEGAWA, Yuji HAISHIMA, Eiichi YA ...
    Session ID: S6-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    International standards such as ISO is positively adopted in worldwide regulations of medical devices. Medical devices that come into contact with the human body must be evaluated for biological safety according to the ISO 10993 series. ISO 10993-1, " Evaluation and testing within a risk management process" requires the collection of information on the chemical properties of medical materials. The concept of toxicological threshold of concern (TTC) may be able to promote animal welfare and reduce the testing costs. Safety assessment methods using chemical analysis are expected to be increasingly used in the near future. The U.S. Food and Drug Administration is actively promoting the use of chemical analysis. In Japan, some medical device manufacturers and contract research organizations are attempting to use chemical analysis. Chemical analysis is very useful for assessing the amount of elution from medical devices and exposure amounts to human when the analysis target chemical is known. On the other hand, the information available for chemical analysis is limited because raw material suppliers do not disclose the full composition including impurities. Therefore, the TTC approach can be used for a comprehensive assessment of medical devices. However, there are several problems in the current extractables and leachables analysis used for the approach. In this presentation, we propose a strategic analytical package that is available as an alternative for genotoxicity and sensitization testing according to the TTC and the dermal sensitization threshold approaches.

    Download PDF (3019K)
Symposium 7: Current status and perspective for Cutting-Edge Technologies for accelarating the research of toxicological mechanism
  • Ikuro SUZUKI
    Session ID: S7-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Due to the international trend towards the abolition of animal testing and the enactment of the FDA Modernization Act 2.0, there is a growing expectation for in vitro testing methods. One of these methods is the microelectrode array (MEA) measurement, which can non-invasively measure in vitro neural activity and is used as one of the toxicity evaluation methods for compounds. The OECD has created guidance on this evaluation method. In this presentation, I will introduce the current state of compound toxicity evaluation and action mechanism prediction using MEA in both central and peripheral nerves. Specifically, we will present a toxicity evaluation method that can be extrapolated to in vivo situations from waveform data obtained from human iPS cell-derived central nerves, sensory neurons, and brain organoids, as well as a mechanism prediction method using deep learning. Additionally, we will introduce our efforts to predict the mechanism of action through detailed analysis at the single-cell level of big data obtained by "field potential imaging" using a 236,880-electrode CMOS-MEA.

    Download PDF (3013K)
  • Tadahiro SHINOZAWA
    Session ID: S7-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    In recent years, innovative technologies beyond the conventional have begun to be developed and applied to drug discovery research. With innovations in iPS cell technology, gene editing and Omics-related technologies, the granularity of data has become finer than before, and it was realized that a huge amount of data can be handled in the safety evaluation process with multiple readouts. For example, according to a survey by a pharmaceutical company in the EU, high content analysis technology that can analyze the health status of cells from multiple angles is understanded as a game changer in several companies in 2015. It has been used as a practical tool by almost companies in 2020. On the other hand, retrospective analysis using biobanks not only contributes to the elucidation of disease mechanisms in human but is beginning to be applied to drug safety research. Using knowledge management databases integrating the results from complex searches with automated processing will help to handle the big data volumes. Thus, by leveraging knowledge management to accumulates various in vitro data and in vivo data and integrates clinical information, we may be able to predict drug safety more accurately in pre-clinical stage. Finally, machine learning and AI with the database could help to develop hypotheses of the toxicological mechanism for early candidates. In this presentation, I try to introduce examples of various in vitro data accumulation in our company and discuss the future potential and challenges for using AI in investigative toxicology.

    Download PDF (3016K)
  • Ichiji NAMATAME
    Session ID: S7-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Induced pluripotent stem cells (iPSC) can differentiate into any cell type, making them promising for cell therapy applications, as well as for cell-based assays in in vitro pharmacological evaluation, including evaluating the safety and toxicity of drug candidates. However, the maintenance and differentiation of iPSC cultures requires high technical expertise. Cell-based assays involve complex processes such as long-term culture, real-time observation of live cells, and detection of many cellular events. To facilitate these processes, Astellas has developed Mahol-A-Ba, an automated drug discovery platform. It consists of Maholo, a dual-arm robot, and Screening station, an automated experiment system. Mahol-A-Ba quantifies the advanced experimental techniques required for iPSC experimentation and translates them into robot behavior; automates multi-sample processing of multiple processes; improves reproducibility by limiting human error; and saves researchers valuable time otherwise spent on tedious experiment. In addition, by incorporating AI image analysis technology, Mahol-A-Ba allows high-quality pharmacological evaluation by tracking and quantifying minute changes that were previously only evaluable with a qualitative score. Furthermore, Mahol-A-Ba can perform experiments and analyses remotely. In the future, this platform is expected to harmonize the activities of researchers, robots, and AI in a highly orchestrated manner; promote global research collaboration; and accelerate the discovery of new drug candidates.

    Download PDF (3020K)
  • Hiroki DANNO
    Session ID: S7-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Our single-cell transcriptome technology was ranked as No.1 on both accuracy scores and overall scores in an international benchmarking of the Human Cell Atlas project, where 25 research teams participated from 7 countries and the results were published in the Nature Biotechnology (Mereu et al. 2020). We also developed high-throughput bulk transcriptome technology which can analyze a large variety of bulk samples simultaneously at less than a tenth the cost of existing technology such as microarray or RNA sequencing. Whole transcriptome based phenotypic drug discovery become more important to analyze drug efficacy to cells on genome-wide basis recently. As it can detect influences of drugs on pathways in cells comprehensively, we can select compounds with novel mechanism while evaluating drug efficacy and toxicity, as well as extrapolating drug target pathways and mechanisms. In 2018, previous study reported whole transcriptome-based drug screening technology, “DRUG-seq” in the Nature Communications (Ye et al. 2018). As a demonstration, we compared gene detection capability of our bulk transcriptome technology with that of “DRUG-seq”, by conducting comparable experiments under the same conditions. While the DRUG-seq detected 10,000 genes by sequencing of 2.17 million reads, our technology detected almost the same number of genes by sequencing only 0.23 million reads. It showed our bulk technology can analyze 10 times more compounds in 1 run of the next generation sequencer.

    Download PDF (3020K)
  • Yoshiyuki ARATA
    Session ID: S7-5
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Cancer immunotherapy is a rapidly growing field in the treatment of cancer and holds great promise for improving patient outcomes. One of the major challenges in the development of T-cell engagers, a type of immunotherapy that redirects T-cells to attack cancer cells drugs, is the risk of on-target/off-tumor toxicity. This occurs when the immune system mistakenly attacks healthy tissues due to expression of target antigen on healthy tissues as well as tumor, leading to severe side effects, including tissue injury and cytokine release syndrome.

    In this study, we aimed to extrapolate on-target/off-tumor toxicity by T-cell engager from cynomolgus monkeys to humans, using cyno- and human-derived organoids. For the in vitro assessment, we employed a co-culture system of peripheral blood mononuclear cells (PBMCs) and organoids, which are three-dimensional cell cultures that mimic the structure and target expression of organs. The in vitro data reflected intestinal toxicity observed in in vivo toxicity study with cynomolgus monkeys. Comparing the in vitro result using cyno- and human-derived organoids, we discussed the extrapolation from the preclinical data in cynomolgus monkeys to humans.

    This tool helps researchers to better understand the mechanisms of on-target/off-tumor toxicity and extrapolate the results of animal studies to human prediction, thus can be a valuable approach for the safety assessment of T-cell engagers.

    Download PDF (3014K)
Symposium 8: Elucidation and Detection of Novel Toxicity Mechanisms via Chemical Adduct Formation
  • Takashi UEHARA
    Session ID: S8-1
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    It has been pointed out that dysfunction of the epigenome results in various diseases. However, the mechanism by which the epigenome is regulated remains unclear. We focused on DNA methyltransferase (DNMT), which catalyzes cytosine methylation in the genome. Initially, we investigated the effect of nitric oxide (NO) on DNMT. NO oxidatively modifies (S-nitrosylates) the thiol portion of cysteine residues in various proteins and modulates its enzymatic activity. We examined whether NO induces DNMT S-nitrosylation in vitro and in vivo. S-Nitrosylation of DNMT (SNO-DNMT) formation was observed by treatment with NO donor in a concentration- and time-dependent manner. In addition, the enzymatic activity of DNMT was significantly reduced by exposure to NO. This inhibition resulted in genomic DNA demethylation, which in turn led to various genes expression. This phenomenon may be involved in NO-mediated pathogenesis. We then screened for environmental chemicals with similar effects to NO. Interestingly, some environmental electrophiles inhibited DNMT enzymatic activity via covalent modifications. We will introduce the recent findings on the properties and functions of those compounds.

    Download PDF (3018K)
  • Akihiro ITO
    Session ID: S8-2
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Epigenetic regulation of gene expression determines cell fate and is involved in various life phenomena. On the other hand, this epigenetic genetic information can be influenced by diverse factors such as environment, and aberrant epigenetic changes are associated with various diseases, including cancers. Lysine acylation of histones is one of the molecular bases of epigenetic regulation. Some of these lysine acylations are caused by the addition of endogenous carboxylic acids such as fatty acids via acyl-CoA. We consume a variety of compounds daily through our diet, including carboxylic acids. These unintentionally ingested environment-derived carboxylic acids may modify histones and alter gene expression. Indeed, we found novel histone modifications induced by environment-derived carboxylic acids. In this presentation, we would like to introduce the hidden functions of environmental chemicals, focusing on the regulation of gene expression by histone modifications derived from sorbic acid, a food additive.

    Download PDF (3012K)
  • Naoshi DOHMAE
    Session ID: S8-3
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    “Exposome” can be defined as the totality of environmental factors to which an individual in a lifetime, and these environmental factors have a greater impact on humans than genetic factors, contributing to diseases such as cancer. Although there is currently no known method to accurately identify the exposome, we believe that interactions between proteins and environmental substances are essential for environmental substances to trigger biological reactions, and we have proposed the "adduct exposome" to examine protein modifications by environmental substances. Currently, DDA (data dependent aquisition, also called shotgun analysis) is the mainstream proteomics method to comprehensively examine protein modifications, and we are attempting to elucidate the adduct exosome based on label-free quantification (LFQ) using this method. We have developed a method for proteomic analysis by adding various compounds to cells. In collaboration with Prof. Akihiro Ito and colleagues at Tokyo University of Pharmacy and Life Sciences, we added food additives to cells and searched for histone modifications in cells. They found that many organic acid compounds exhibit covalent binding to histones, which may regulate gene expression in an epigenetic manner. We will continue to investigate various compounds and protein modifications to study the relationship between compound exposure and biological reactions such as aging, addiction, and disease.

    Download PDF (3013K)
  • Kosuke DODO
    Session ID: S8-4
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    We previously developed Alkyne-tag Raman Imaging (ATRI) method, in which alkyne-tagged small molecules are visualized in live cells by using Raman microscope. Alkyne tag is compact and shows a strong Raman signal in the cellular silent region where no endogenous biomolecules have Raman signals. We applied this method for the analysis of adduct-forming compounds. In addition, we succeeded in the improvement of Raman signal of alkyne tag by using Surface Enhanced Raman Scattering (SERS) on metal nanoparticles, and it realized the analysis of protein adducts and cellular uptake. Furthermore, we recently investigated Raman imaging using deuterium, which is more compact than alkyne. In this presentation, the detection of adduct-forming chemicals by Raman spectroscopy will be presented.

    Reference 1) H. Yamakoshi, K. Dodo, M. Okada, J. Ando, A. Palonpon, K. Fujita, S. Kawata, M. Sodeoka, J. Am. Chem. Soc. 2011, 133, 6102. 2) H. Yamakoshi, K. Dodo, A. F. Palonpon, J. Ando, K. Fujita, S. Kawata, M. Sodeoka, J. Am. Chem. Soc. 2012, 134, 20681. 3) J. Ando, M. Asanuma, K. Dodo, H. Yamakoshi, S. Kawata, K. Fujita, M. Sodeoka, J. Am. Chem. Soc. 2016, 138, 13901. 4) K. Koike, K. Bando, J. Ando, H. Yamakoshi, N. Terayama, K. Dodo, N. I. Smith, M. Sodeoka, K. Fujita, ACS Nano, 2020, 14, 15032. 5) K. Dodo, A. Sato, Y. Tamura, S. Egoshi, K. Fujiwara, K. Oonuma, S. Nakao, N. Terayama, M. Sodeoka, Chem. Commun., 2021, 57, 2180. 6) K. Dodo, K. Fujita, M. Sodeoka, J. Am. Chem. Soc., 2022, 144, 19651.

    Download PDF (3033K)
  • Koji UCHIDA
    Session ID: S8-5
    Published: 2023
    Released on J-STAGE: March 08, 2024
    CONFERENCE PROCEEDINGS FREE ACCESS

    Protein structures are extremely diverse. When subjected to post-translational modifications by some active species, their structures and functions change according to the surrounding environment, leading to the expression of functions, such as biological responses. Such cases are found in host defense reactions, such as detoxification and innate immunity. Therefore, chemical protein modification not only changes the protein structure, but also imparts new functionality to the protein in an adduct-specific manner, which can be used as a signal. On the other hand, recent studies have shown that not only endogenous short-lived active species derived from fatty acids and carbohydrates, but also everyday common compounds, such as vegetable food ingredients, act on proteins and that the modified structures formed on the proteins also interacts with non-immune cells and proteins. Based on this background, it is important to comprehensively analyze the characteristic patterns of chemical modification ("modification signatures") of biological components, such as proteins and lipids, and to understand the mechanisms of biological responses to them. This will bring new changes and conversions to the biology of chemical modification, which has been regarded as a mere “injury” until now. In this symposium, I would like to introduce recent findings on the mechanisms of biological responses brought about by the formation of modified structures caused by oxidation.

    Download PDF (3014K)
feedback
Top