Animal Research for Alzheimer Disease: Failures of Science and Ethics

This chapter addresses the epidemiology and current understanding of AD as a scientific and societal challenge, reviews the uses and results of animal research in basic science and drug development, and discusses risk factors and funding. Important follow-up topics, including current and in-development, human-relevant approaches for replacement of the failed animal research paradigm, deserve comparable treatment and hence are not addressed here. The reader is referred to the list of recommended readings at the end of the chapter for further discussion of these topics

Strategic aims for improving the regulatory assessment of Developmental Neurotoxicity (DNT) using non-animal methods

Currently, the identification of chemicals that have the potential to induce developmental neurotoxicity (DNT) is based on animal testing, since there are no regulatory accepted alternative methods for this purpose. Since at the regulatory level, systematic testing of DNT is not a standard requirement within the EU legislation of chemical safety assessment, DNT testing is only performed in higher tiered tests triggered based on structure activity relationships or evidence of neurotoxicity in systemic adult studies. However, these triggers are rarely used and in addition do not always serve as reliable indicators of DNT as they are observed in an adult rodent animal. Consequently, to date only a limited amount of chemicals (Grandjean and Landrigan, 2006; Smirnova et al., 2014), mainly pesticides (Bjørling-Poulsen et al., 2008) have been tested under US EPA (OPPTS 870.630) or OECD DNT TG 426. Therefore, there is the pressing need for developing alternative methodologies that can more rapidly and cost-effectively screen large numbers of chemicals for their potential to cause DNT. In this report we propose that in vitro studies could contribute to the identification of potential triggers for DNT evaluation since existing cellular models permit the evaluation of a chemical impact on key neurodevelopmental processes, mimicking different windows of human brain development, especially if human models derived from induced pluripotent stem cells are applied. Furthermore, the battery of currently available DNT alternative test methods anchored to critical neurodevelopmental processes and key events identified in DNT Adverse Outcome Pathways (AOPs) could be applied to generate in vitro data useful for various regulatory purposes. Incorporation of in vitro mechanistic information would increase scientific confidence in decision making, by decreasing uncertainty and leading to refinement of chemical grouping according to biological activity. In this report development of IATA (Integrated Approaches to Testing and Assessment) based on key neurodevelopmental processes and AOP-informed is proposed as a tool for not only speeding up chemical screening, but also providing mechanistic data in support of hazard assessment and in the evaluation of chemical mixtures. Such mechanistically informed IATA for DNT evaluation could be developed integrating various sources of information (e.g., non-testing methods, in vitro approaches, as well as in vivo animal and human data), contributing to screening for prioritization, hazard identification and characterization, and possibly safety assessment of chemicals, speeding up the evaluation of thousands of compounds present in industrial, agricultural and consumer products that lack safety data on DNT potential. It is planned that the data and knowledge generated from such testing will be fed into the development of an OECD guidance document on alternative approaches to DNT testing.JRC.F.3-Chemicals Safety and Alternative Method

The Three-Layer Concentric Model of Glioblastoma: Cancer Stem Cells, Microenvironmental Regulation, and Therapeutic Implications

Tumors arising in the central nervous system are thought to originate from a sub-population of cells named cancer stem cells (CSCs) or tumor initiating cells (TICs) that possess an immature phenotype, combined with self-renewal and chemotherapy resistance capacity. Moreover, in the last years, these cells have been identified in particular brain tumor niches fundamental for supporting their characteristics. In this paper, we report studies from many authors demonstrating that hypoxia or the so called “hypoxic niche” plays a crucial role in controlling CSC molecular and phenotypic profile. We recently investigated the relationship existing between Glioblastoma (GBM) stem cells and their niche, defining the theory of three-concentric layers model for GBM mass. According to this model, GBM stem cells reside preferentially within the hypoxic core of the tumour mass, while more differentiated cells are mainly localized along the peripheral and vascularized part of the tumour. This GBM model provides explanation of the effects mediated by the tumour microenvironment on the phenotypic and molecular regulation of GBM stem cells, describing their spatial distribution in the tumor bulk. Moreover, we discuss the possible clinical implications of the creation of this model for future GBM patient management and novel therapeutic strategies development

A Survey on Monitoring Innovation and Societal Impact of EU-funded Research: Factual Summary Report

The Joint Research Centre (JRC) of the European Commission (EC), in collaboration with the EC Directorate General for Research and Innovation initiated an activity to define suitable indicators to retrospectively assess the impact of European Union (EU)-funded research. To this aim, the JRC conducted a survey addressed to current and former participants of EC-funded research projects in the fields of Alzheimer’s disease and other dementias, breast cancer, and prostate cancer. This summary report provides a brief factual overview of the replies received, with information on the respondents as well as the number of responses and range of opinions. The replies gathered through this survey will help the European Commission assess how EU-funded research activities have contributed to innovation and impact.JRC.F.3-Chemicals Safety and Alternative Method

Molecular Mechanisms of HIF-1α Modulation Induced by Oxygen Tension and BMP2 in Glioblastoma Derived Cells

BACKGROUND: Glioblastoma multiforme (GBM) is one of most common and still poorly treated primary brain tumors. In search for new therapeutic approaches, Bone Morphogenetic Proteins (BMPs) induce astroglial commitment in GBM-derived cells in vitro. However, we recently suggested that hypoxia, which is characteristic of the brain niche where GBM reside, strongly counter-acts BMP effects. It seems apparent that a more complete understanding of the biology of GBM cells is needed, in particular considering the role played by hypoxia as a signaling pathways regulator. HIF-1alpha is controlled at the transcriptional and translational level by mTOR and, alike BMP, also mTOR pathway modulates glial differentiation in central nervous system (CNS) stem cells. METHODOLOGY/PRINCIPAL FINDINGS: Here, we investigate the role of mTOR signaling in the regulation of HIF-1alpha stability in primary GBM-derived cells maintained under hypoxia (2% oxygen). We found that GBM cells, when acutely exposed to high oxygen tension, undergo Akt/mTOR pathway activation and that BMP2 acts in an analogous way. Importantly, repression of Akt/mTOR signaling is maintained by HIF-1alpha through REDD1 upregulation. On the other hand, BMP2 counter-acts HIF-1alpha stability by modulating intracellular succinate and by controlling proline hydroxylase 2 (PHD2) protein through inhibition of FKBP38, a PHD2 protein regulator. CONCLUSIONS/SIGNIFICANCE: In this study we elucidate the molecular mechanisms by which two pro-differentiating stimuli, BMP2 and acute high oxygen exposure, control HIF-1alpha stability. We previously reported that both these stimuli, by inducing astroglial differentiation, affect GBM cells growth. We also found differences in high oxygen and BMP2 sensitivity between GBM cells and normal cells that should be further investigated to better define tumor cell biology

Development of a pluripotent stem cell derived neuronal model to identify chemically induced pathway perturbations in relation to neurotoxicity: Effects of CREB pathway inhibition

JRC.I.5-Systems Toxicolog

In Vitro–In Vivo Extrapolation by Physiologically Based Kinetic Modeling: Experience With Three Case Studies and Lessons Learned

Physiologically based kinetic (PBK) modeling has been increasingly used since the beginning of the 21st century to support dose selection to be used in preclinical and clinical safety studies in the pharmaceutical sector. For chemical safety assessment, the use of PBK has also found interest, however, to a smaller extent, although an internationally agreed document was published already in 2010 (IPCS/WHO), but at that time, PBK modeling was based mostly on in vivo data as the example in the IPCS/WHO document indicates. Recently, the OECD has published a guidance document which set standards on how to characterize, validate, and report PBK models for regulatory purposes. In the past few years, we gained experience on using in vitro data for performing quantitative in vitro–in vivo extrapolation (QIVIVE), in which biokinetic data play a crucial role to obtain a realistic estimation of human exposure. In addition, pharmaco-/toxicodynamic aspects have been introduced into the approach. Here, three examples with different drugs/chemicals are described, in which different approaches have been applied. The lessons we learned from the exercise are as follows: 1) in vitro conditions should be considered and compared to the in vivo situation, particularly for protein binding; 2) in vitro inhibition of metabolizing enzymes by the formed metabolites should be taken into consideration; and 3) it is important to extrapolate from the in vitro measured intracellular concentration and not from the nominal concentration to the tissue/organ concentration to come up with an appropriate QIVIVE for the relevant adverse effects

Possible implications for SARS-CoV-2 impact on brain development

Funding Information: The authors would like to thank Dr. Marc Peschanski (I-Stem, Évry, France) for providing IMR90-hiPSCs, and Dr. Anna Navarro Cuenca for providing the license for the use of BioRender.com. Publisher Copyright: © 2022The possible neurodevelopmental consequences of SARS-CoV-2 infection are presently unknown. In utero exposure to SARS-CoV-2 has been hypothesized to affect the developing brain, possibly disrupting neurodevelopment of children. Spike protein interactors, such as ACE2, have been found expressed in the fetal brain, and could play a role in potential SARS-CoV-2 fetal brain pathogenesis. Apart from the possible direct involvement of SARS-CoV-2 or its specific viral components in the occurrence of neurological and neurodevelopmental manifestations, we recently reported the presence of toxin-like peptides in plasma, urine and fecal samples specifically from COVID-19 patients. In this study, we investigated the possible neurotoxic effects elicited upon 72-hour exposure to human relevant levels of recombinant spike protein, toxin-like peptides found in COVID-19 patients, as well as a combination of both in 3D human iPSC-derived neural stem cells differentiated for either 2 weeks (short-term) or 8 weeks (long-term, 2 weeks in suspension + 6 weeks on MEA) towards neurons/glia. Whole transcriptome and qPCR analysis revealed that spike protein and toxin-like peptides at non-cytotoxic concentrations differentially perturb the expression of SPHK1, ELN, GASK1B, HEY1, UTS2, ACE2 and some neuronal-, glia- and NSC-related genes critical during brain development. Additionally, exposure to spike protein caused a decrease of spontaneous electrical activity after two days in long-term differentiated cultures. The perturbations of these neurodevelopmental endpoints are discussed in the context of recent knowledge about the key events described in Adverse Outcome Pathways relevant to COVID-19, gathered in the context of the CIAO project (https://www.ciao-covid.net/).publishersversionpublishe

The use of plant-derived bioactive compounds to target cancer stem cells and modulate tumor microenvironment

In the last decades cancer has been considered as an epigenetic dysfunction, given the profound role of diet and lifestyle in cancer prevention and the determination of cancer risk. A plethora of recent publications have addressed the specific role of several environmental factors, such as nutritional habits, behavior, stress and toxins in the regulation of the physiological and cancer epigenome. In particular, plant-derived bioactive nutrients have been seen to positively affect normal cell growth, proliferation and differentiation and also to revert cancer related epigenetic dysfunctions, reducing tumorigenesis, preventing metastasis and/or increasing chemo and radiotherapy efficacy. Moreover, virtually all cancer types are characterized by the presence of cancer stem cell (CSC) subpopulations, residing in specific hypoxic and acidic microenvironments, or niches, and these cells are currently considered responsible for tumor resistance to therapy and tumor relapse. Modern anti-cancer strategies should be designed to selectively target CSCs and modulate the hypoxic and acidic tumor microenvironment, and, to this end, natural bioactive components seem to play a role. This review aims to discuss the effects elicited by plant-derived bioactive nutrients in the regulation of CSC self-renewal, cancer metabolism and tumor microenvironment

Role of plant-based diets in the prevention and regression of metabolic syndrome and neurodegenerative diseases

Plant-based diets are known to preserve body tissues from oxidative stress and inflammation, both hallmarks of chronic-degenerative diseases. In particular, plant-derived foods, such as legumes, represent a natural source of bioactive nutrients known to contribute to the prevention and amelioration of insulin resistance, dyslipidemia, hypertension and impaired glucose metabolism, all factors implicated in metabolic syndrome (MetS), but also osteoporosis, neurodegeneration and some types of cancers. Here we revise recent literature on the role of plant-based diets, plant-foods and specific plant-nutrients in the prevention and regression of MetS and neurodegenerative diseases. We describe some of the molecular mechanisms underlying these protective effects, highlighting the role of diet in the control of hyper-homocysteinemia and insulin resistance, often implicated in the etiology of both metabolic and neurodegenerative syndromes