#	Pagina
attuale pagina	/open-h2020/projects/215979/results.html

Report

Teaser, summary, work performed and final results

Periodic Reporting for period 1 - PATROLS (Physiologically Anchored Tools for Realistic nanOmateriaL hazard aSsessment)

Teaser

Summary

Classical hazard testing strategies to define the human and environmental health impact of engineered nanomaterials (ENM) commonly apply unrealistic acute, high-dose exposures to models that do not reflect the in vivo environment. Furthermore, existing in vitro and in silico hazard detection methods are not highly predictive. Hence there is an urgent need to address these limitations by developing improved in vitro and in silico tools tailored to realistic ENM exposure scenarios. PATROLS therefore aims to establish and standardise a battery of next generation physiologically anchored, hazard assessment tools that more accurately predict adverse human health and environmental effects caused by long-term, low dose ENM exposure to support regulatory risk decision-making. The project objectives are to develop: 1) more realistic in vitro three-dimensional (3D) lung, gastrointestinal tract (GIT) and liver models for mechanism-based hazard assessment, which will be anchored against chronic in vivo exposure outcomes and will report on mechanistic endpoints linked to adverse outcome pathways (AOPs); 2) novel methods to evaluate long-term exposure hazard endpoints in ecologically relevant test systems and organisms, selected according to their position in the food chain; these endpoints will be based on key events associated with AOPs; 3) robust in silico methods for exposure and dosimetry modelling, as well as in vitro-to-in vivo extrapolation (IVIE) and hazard prediction; 4) improved understanding of ENM physico-chemical identity and dosimetry in physiologically relevant exposure scenarios dictated by the advanced human and environmental model development. Iterative training, testing, stakeholder feedback and pre-validation phases will be integral to establish robust, fit-for-purpose test systems. These objectives will be achieved by an international network of world-leading academic, governmental, industrial, SME, risk assessment agency and NGO partners. PATROLS will therefore provide an innovative and effective set of models, bioassays and computational tools to more reliably predict potential hazards resulting from realistic ENM exposures. These tools will minimise the necessity for animal testing to promote the 3Rs and will support for future nanosafety requirements.

Work performed

During this first project period a range of standardised approaches for the PATROLS consortium have been put in place. For example, a centralised database has been established for both new data collation and linkage to large legacy data-sets generated by recently completed European projects (WP6); this database is hosted on the eNanoMapper platform. Guidance documents have been established for ENM acquisition, handling, dispersion and endotoxin testing. Additionally, we have held two Standard Operating Procedure (SOP) workshops: a PATROLS-GRACIOUS-NanoSafety Cluster event on harmonization of Standard Operating Procedures and a PATROLS Ecotoxicity SOP workshop. The workshopsâ€™ outcome is an SOP template being used throughout the consortium, which has undergone review by OECD and ISO representatives to aid future adoption of the methods developed in PATROLS.

Key scientific and technical advances made to date include:
â€¢ Collated and tabulated the primary physico-chemical characteristics of the first ENM set to benchmark the new test systems being developed. Characterisation of the secondary ENM features under experimental conditions relevant to both human cell culture & environmental models is nearly completed (e.g. dissolution, redox potential, size distribution / agglomeration state).
â€¢ Completed a comprehensive review & database of all existing long-term in vivo oral and inhalation toxicity studies, for bench-marking of the advanced in vitro models under development. Effects after exposure, NOAEL/LOAELs, biodistribution and data gaps have been highlighted in this database. Additionally, further development of adverse outcome pathways (AOPs) to link in vitro with in vivo data are underway, to support identification of new predictive biomarkers later in the project for lung inflammation, fibrosis, cancer & mesothelioma; and liver inflammation, fibrosis & cancer.
â€¢ Designed the first generation lung, liver and gastro-intestinal (GIT) tract models, which are being tested with more realistic repeated ENM exposures. The first generation lung model is adapted for repeated aerosolisation exposures for upto 3-weeks. Two liver models systems have been developed; a commercially available InSphero system that supports repeated exposures for up to 3-weeks and a cell line based spheroid system facilitating genotoxicity testing which supports repeated exposures for up to 10-days. An intestinal triple culture has been established using enterocytes, goblet cells and macrophages. This GIT model has been characterised for barrier integrity (TEER, ZO-1/F-actin staining), cytokine release and mucus generation, and has been exposed repeatedly to low-dose ENM. Additionally, a prototype bioreactor has been developed and is currently being trialed which will facilitate both fluid flow and tissue flexing with the in vitro lung and GIT models.
â€¢ Long-term (multi-generation) exposure studies in have been conducted in several environmental organisms, where more significant toxic responses were observed when compared to acute exposures. A transgenic fish line for early signalling of ENM exposure has also been developed
â€¢ Established the PATROLS database and prepared two PATROLS MOdelling DAta (MODA) using templates originally developed by the European Materials Modelling Council (EMMC). This WP is also currently developing and implementing a QSAR and PBPK models for ENM.
â€¢ The PATROLS website and social media accounts (Twitter & LinkedIn) have been launched; strong connections to key stakeholders have been built through industrial events, integration into OECD activities and representation at high-profile events.

Final results

PATROLS is aimed at advancing the state-of-the-art in ENM human and environmental hazard assessment through the provision of improved and innovative tools that reduce the necessity for animal testing. The first period of the PATROLS project has been focused on development work to improve current hazard testing systems. This has involved adapting and further advancing cell based testing system by 1) enhancing their physiological relevance through increased biological complexity; and 2) extending their culture time whilst maintaining normal cell function and viability, to better support more realistic longer-term, repeated dosing scenarios. Ecological assays have also been improved to better support long-term ENM exposures, through development of novel systems to both support chronic environmental exposure and detect new hazard endpoints with greater sensitivity. Thus, PATROLS has already created advances in state-of-the-art ENM hazard testing, but to facilitate more demonstrable impact of the work in the final half of the project, the PATROLS partners have been pro-active in developing relationships and engaging with a range of stakeholders to raise awareness of the project. This has involved active interactions with stakeholders through dedicated workshops, webinars, distribution of e-newsletters, public engagement activities and transfer of knowledge generated within the project to on-going OECD Working Party on Manufactured Nanomaterials initiatives.