Altex-Alternatives To Animal Experimentation最新文献

The webinar series and workshop titled “Trust Your Gut: Establishing Confidence in Gastrointestinal Models – An Overview of the State of the Science and Contexts of Use” was co-organized by NICEATM, NIEHS, FDA, EPA, CPSC, DoD, and the Johns Hopkins Center for Alternatives to Animal Testing (CAAT) and hosted at the National Institutes of Health in Bethesda, MD, USA on October 11-12, 2023. New approach methods (NAMs) for assessing issues of gastrointestinal tract (GIT)- related toxicity offer promise in addressing some of the limitations associated with animal-based assessments. GIT NAMs vary in complexity, from two-dimensional monolayer cell line-based systems to sophisticated 3-dimensional organoid systems derived from human primary cells. Despite advances in GIT NAMs, challenges remain in fully replicating the complex interactions and pro­cesses occurring within the human GIT. Presentations and discussions addressed regulatory needs, challenges, and innovations in incorporating NAMs into risk assessment frameworks; explored the state of the science in using NAMs for evaluating systemic toxicity, understanding absorption and pharmacokinetics, evaluating GIT toxicity, and assessing potential allergenicity; and discussed strengths, limitations, and data gaps of GIT NAMs as well as steps needed to establish confidence in these models for use in the regulatory setting.

Hazard assessment requires toxicity tests to allow deriving protective points of departure (PoDs) for risk assessment irrespective of a compound’s mode of action (MoA). The scope of in vitro test batteries (ivTB) needed to assess systemic toxicity is still unclear. We explored the protectiveness regarding systemic toxicity of an ivTB with a scope that was guided by previous findings from rodent studies, where examining six main targets, including liver and kidney, was sufficient to predict the guideline scope-based PoD with high probability. The ivTB comprises human in vitro models representing liver, kidney, lung, and the neuronal system covering transcriptome, mitochondrial dysfunction, and neuronal outgrowth. Additionally, 32 CALUXR- and 10 HepG2 BAC-GFP reporters cover a broad range of disturbance mechanisms. Eight compounds were chosen for causing adverse effects such as immunotoxicity or anemia in vivo, i.e., effects not directly covered by assays in the ivTB. PoDs derived from the ivTB and from oral repeated dose studies in rodents were extrapolated to maximum unbound plasma concentrations for comparison. The ivTB-based PoDs were one to five orders of magnitude lower than in vivo PoDs for six of eight compounds, implying that they were protective. The extent of in vitro response varied across test compounds. Especially for hematotoxic substances, the ivTB showed either no response or only cytotoxicity. Assays better capturing this type of hazard would be needed to complement the ivTB. This study highlights the potentially broad applicability of ivTBs for deriving protective PoDs of compounds with unknown MoA.

The adverse outcome pathways (AOPs) were developed to accelerate evidence-based chemical risk assessment by leveraging data from new approach methodologies. Thanks to their stressor-agnostic approach, AOPs were seen as instrumental in other fields. Here, we present AOPs that report non-chemical stressors along with the challenges encountered for their development. Challenges regarding AOPs linked to nanomaterials include non-specific molecular initiating events, limited understanding of nanomaterial biodistribution, and needs for adaptations of the in silico modeling and testing systems. Development of AOPs for radiation face challenges in how to incorporate ionizing events type, dose rate, energy deposition, and how to account for targeting multiple macromolecules. AOPs for COVID-19 required the inclusion of SARS-CoV-2-specific replicative steps to capture the essential events driving the disease. Developing AOPs to evaluate efficacy and toxicity of cell therapies necessitates addressing the cellular nature and the therapeutic function of the stressor. Finally, addressing toxicity of emerging biological stressors like microbial pesticides can learn from COVID-19 AOPs. We further discuss that the adaptations needed to expand AOP applicability beyond chemicals are mainly at the molecular and cellular levels while downstream key events at tissue or organ level, such as inflammation, are shared by many AOPs initiated by various stressors. In conclusion, although it is challenging to integrate non-chemical stressors within AOPs, this expands opportunities to account for real-world scenarios, to identify vulnerable individuals, and to bridge knowledge on mechanisms of adversity.

The 4th Annual Forum on Endocrine Disrupters organized by the European Commission brought together the authors of this article around the topic: “From bench to validated test guidelines: (pre)val­idation of test methods”. Validation activities are meant to demonstrate the relevance and reliability of methods and approaches used in regulatory safety testing. These activities are essential to facil­itate regulatory use, still they are largely underfunded and unattractive to the scientific community. In the last decade, large amounts of funding have been invested in European research towards the development of approaches that can be used in regulatory decision-making, including for the identification of endocrine disrupters. There is a vast pool of candidate test methods for potential reg­ulatory applications, but most of them will not be used due to the absence of consideration of their relevance and reliability outside the method developer’s laboratory. This article explains the reasons why such a gap exists between the outputs of research projects and the uptake in a regulatory context. In parallel, there are also increasing expectations from the regulatory science community that validation becomes more efficient with respect to time and resources. This article shares some of the lessons learned and proposes paths forward for validation of new methods that are not intended as one-to-one replacements of animal studies. This includes submitting only mature methods for validation that were developed following good practices and good documentation, proposing a greater emphasis on well-documented transferability studies, and adopting a cost-sharing model among those who benefit from validated methods.

Disease models enable researchers to investigate, test, and identify therapeutic targets that would alter the patients’ disease condition and improve quality of life. Advances in genetic alteration and analytical techniques have enabled rapid devel­opment of disease models using preclinical animals and cell cultures. However, success rates of drug development remain low due to limited recapitulation of clinical pathophysiology by these models. To resolve this challenge, the pharmaceutical industry has explored microphysiological system (MPS) disease models, which are complex in vitro systems that include but are not limited to organ-on-a-chip, organoids, spheroids, and 3D bioengineered tissues (e.g., 3D printing, hydrogels). Capable of integrating key in vivo properties, such as disease-relevant human cells, multi-cellularity/dimensionality of organs, and/or well-controlled physical and molecular cues, MPS disease models are being developed for a variety of indications. With on-going qualifications or validations for wide adoption within the pharmaceutical industry, MPS disease models hold exciting potential to enable in-depth investigation of in vivo pathophysiology and enhance drug discovery and development processes. To introduce the present status of MPS disease models, this paper describes notable examples in six disease areas: cancer, liver/kidney diseases, respiratory diseases/COVID-19, neurodegenerative diseases, gastrointestinal diseases, and select rare diseases. Additionally, we describe current technical limitations and provide recommendations for future development that would expand application opportunities within the pharmaceutical industry.

A structurally diverse set of 147 per- and polyfluoroalkyl substances (PFAS) was screened in a panel of 12 human primary cell systems by measuring 148 biomarkers relevant to (patho)physiological pathways to inform hypotheses about potential mechanistic effects of data-poor PFAS in human model systems. This analysis focused on immunosuppressive activity, which was previously reported as an in vivo effect of perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS), by comparing PFAS responses to four pharmacological immunosuppressants. The PFOS response profile had little correlation with reference immunosuppressants, suggesting in vivo activity does not occur by similar mechanisms. The PFOA response profile did share features with the profile of dexamethasone, although some distinct features were lacking. Other PFAS, including 2,2,3,3-tetrafluoropropyl acrylate, demonstrated more similarity to the reference immunosuppressants but with additional activities not found in the reference immunosuppressive drugs. Correlation of PFAS profiles with a database of environmental chemical responses and pharmacological probes identified potential mechanisms of bioactivity for some PFAS, including responses similar to ubiquitin ligase inhibitors, deubiquitylating enzyme (DUB) inhibitors, and thioredoxin reductase inhibitors. Approximately 21% of the 147 PFAS with confirmed sample quality were bioactive at nominal testing concentrations in the 1-60 micromolar range in these human primary cell systems. These data provide new hypotheses for mechanisms of action for a subset of PFAS and may further aid in development of a PFAS categorization strategy useful in safety assessment.