newsletter october 2014 - NetSuite

Newsletter october 2014

Welcome to NANOSOLUTIONS!

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elcome to the NANOSOLUTIONS newsletter. In this newsletter we present an overview of the NANOSOLUTIONS project and explain in simple terms its main aims and objectives as a whole. This article also featured in Projects Magazine. We also speak to three work package leaders (Richard Handy, Dario Greco and Lang Tran) as a way of explaining the fundamental nature of the Engineered Nanomaterial Safety Classifier (ENM Safety Classifier), which is at the heart of the project. The three explain just what the classifier is, why it is needed and the experimental work taking place in the project that will ultimately deliver it.

As well as this, we will have progress updates from different work packages, including Materials (WP3), Life-cycle analysis (WP4), ENM bio-nanointeraction in biological media (WP5), Cell-models (WP6), and Disease models (WP8), and finally we will be taking a look at some of the upcoming NANOSOLUTIONS events on nanotechnology and nanosafety. Don’t forget to follow us on twitter @Nano_solutions and get in touch via our website for more debate, comments and the latest news about the progress of the project.

Links to the stories

Contact details

1. What is NANOSOLUTIONS?

PROJECT COORDINATOR Kai Savolainen Telephone: +358 40 742 0574 Email: [email protected] www.ttl.fi/en/Pages/default.aspx

2. Work package updates 3. The ENM Safety Classifier 4. Events diary

dissemination William Davis, IPL Telephone: +44 (0) 1172 033 120 Email: [email protected] www.ipl.eu.com

This project has received funding from the European Union’s Seventh Framework Programme for research, technological development and demonstration under grant agreement no 309329

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Newsletter october 2014 What is NANOSOLUTIONS? NANOSOLUTIONS seeks to identify and elaborate the characteristics of engineered nanomaterials (ENM) that determine their biological hazard potential. It will help develop a safety classification model for ENM based on an understanding of their interactions with living organisms, benefiting industry and enabling innovation Engineered nanomaterials (ENM) – defined as having at least one dimension ≤100nm – have attracted a great deal of interest during recent years, due to their many technologically interesting properties. The unique properties of ENM and their applications have given birth to immense technological and economic expectations for industries using ENM. However, some of these properties have given rise to concern that they may be harmful to humans. Currently, creating commercial products using ENM requires vigorous testing and there are many barriers to overcome.

The main objective of the project is to identify and elaborate those characteristics of ENM that determine their biological hazard potential Scientists, regulators, and the industrial representatives have now begun to investigate the features of ENM in order to be sure of their safe use in nanotechnologies (NT), i.e. technologies utilising ENM. The European Commission has also explored in-depth the characteristics of ENM and issued a document on ways to assure the safety of ENM. An effective test is required for these properties in order to ensure ENM are safe to use. While testing of individual applications of ENM is possible, it is expensive and timeconsuming and acts as a barrier to innovation. The NANOSOLUTIONS project, which began in April 2013, was created to develop a safety classification for engineered nanomaterials (ENM) based on an understanding of their interactions with living organisms at molecular, cellular and organism levels. Many important functions of living organisms take place at the nanoscale. The human body uses natural nanomaterials, such as proteins and other molecules, to control the body’s many systems and processes. The main objective of the project is to identify and elaborate those characteristics of ENM that determine their biological hazard potential. This potential includes the ability of ENM to induce damage at the cellular,

tissue, or organism levels by interacting with cellular structures leading to impairment of key cellular functions. These adverse effects may be mediated by ENM-induced alterations in gene expression and translation, but may involve also epigenetic transformation of genetic functions. The long-term goal is to create a set of biomarkers of ENM toxicity that are relevant in assessing and predicting the safety and toxicity of ENM across species. ENMorganism interaction is complex and depends not simply on the composition of the ENM core, but particularly on its physicochemical properties, which are largely governed by their surface properties. The overarching objective of this research is, therefore, to provide a means to develop an “ENM Safety Classifier” based on their material characteristics, using the understanding of ENM interactions with living organisms at the molecular, cellular and organism level acquired in this consortium. This will give scientists the ability to predict these harmful effects rather than simply describe them once they have occurred. The NANOSOLUTIONS ENM safety classification model will be of great benefit not only to industry, but also in enabling and speeding up innovation. By making the innovation cycle quicker and making it easier to develop commercially viable products that use ENM, NANOSOLUTIONS will deliver results critical for maintaining Europe’s position in the nanotechnology field. The project will contribute to testing procedures that will be shorter and more cost effective. The public will have greater confidence in the products produced using ENM, thus making them more commercially viable. Industry will be more inclined to use innovative technology in their product development if they can assure its safety and they know consumer concerns are abated. The work NANOSOLUTIONS does will be vital in bringing new materials and technologies to market. This article first appeared in Projects Magazine. The publication, which is published by Nanosolutions dissemination WP leader Insight, provides a platform for research dissemination and seeks to highlight the objectives and successes of European research efforts. Visit www.projectsmagazine.eu.com to subscribe free and follow the progress of Nanosolutions and many more FP7 and H2020 projects.

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Newsletter october 2014 NANOSOLUTIONS SEED A SEED is an interactive digital brochure that combines the very best of traditional publishing with the very latest digital publishing techniques, using a variety of media. The SEED and the SEED Research Library have been specifically designed for research projects to enable active dissemination. A SEED for Nanosolutions has been developed, which will grow and develop throughout the lifetime of the project. The SEED Library can be accessed at the below address, where you can join and choose to follow Nanosolutions. http://www.seedresearchlibrary.com/library_home

Work package updates WP3 - Materials Erik Larsen and Manuel Correia The first half of 2014 consisted of finishing the synthesis of the engineered nanomaterials (ENMs), which has now been completed. Present work, which is ongoing, involves the basic characterisation and development of dispersion protocols for the powder materials. Up until now, more than half of the ENMs have been characterised by basic techniques (basic characterisation,

e.g. dynamic light scattering) and dispersion protocols (standard operating procedures) have been sent to the toxicology partners for testing. The toxicology partners are now interacting with WP3 characterisation labs to perform assurance of the quality of the ENMs dispersions (quality assurance loop). WP3 will now concentrate on advanced characterisation of the ENMs in order to establish a set of ENM characteristics (e.g. particle size, aspect ratio, release of ions, surface characteristics) as input for the ENM safety Classifier.

WP3 group picture taken during WP3 meeting (Hotel Bristol, Frankfurt, 19th May 2014). Legend (from left to right): Alexei, Didier, Erik H, Manuel Correia, Richard Handy, Željka Krpetić, Guocheng Wang

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Newsletter october 2014 WP4 - Life-cycle analysis Socorro Vázquez-Campos WP4 is devoted to studying the transformation and release of engineered nanomaterials (ENMs) along their life cycle (production, manufacturing, use and disposal) when used in existing applications or industrial processes, and the assessment of the environmental and health impacts of these releases. Researchers have so far performed a selection of representative applications incorporating each ENM type to be studied during NANOSOLUTIONS. Additionally, a description of the life cycle stages of the ENMs depending on the application or consumer product where they are incorporated has been

carried out, with special relevance being given to the life cycle stages beyond the manufacturing stage. The life cycle stages that are most likely to result in the transformation of the ENMs and/or result in the release of ENMs have been identified, prioritising normal use conditions (releases generated in accidental scenarios have not been considered). On this basis, researchers in WP4 are currently designing and conducting experiments to investigate the release of ENMs from nanoproducts. The work being performed in WP4 will help to validate the NANOSOLUTIONS hazard tool by comparing physicochemical characterisation and release results of simulations with the predictions given by the tool.

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C A) TEM image of CuO nanoparticles; B) dispersion used to produce nano-enabled fabrics with antimicrobial properties; C) padding process in which the nanoparticles are deposited on the fabric surface

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Newsletter october 2014 WP5 - ENM bio-nanointeraction in biological media Marco Monopoli The overall objective of WP5 is to clarify the ‘biological identity’ of ENMs before and after they have entered into cells or organisms so that the relationship between the ENM surface (biomolecule corona composition) and mechanisms of toxicity (identified in WP7-10) can be understood. As soon as nanomaterials enter complex environments, such as tissue culture medium, blood plasma etc. they become surrounded by biomolecules with which they may strongly bind, forming a ‘corona’

that influence their biological fate, while their surface remains cloaked. WP5’s activities are focused on the study of these interactions in order to evaluate the behaviour of nanoparticles when engaging with living systems, cellular barriers and specific receptors. Firstly, the dispersion state of nanomaterials are characterised prior to and after the exposure to complex media. Secondly, methods have been developed to isolate and characterise the subsequently formed complexes with physicochemical and proteomics techniques to identify and understand the impact of the biomolecular corona.

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A) WP5 laboratory work B) Marco Monopoli and C) Željka Krpetić

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Newsletter october 2014 WP6 – Cell models Bengt Fadeel and Lucian Farcal WP6 works on the application of in vitro methods for the assessment of nanomaterial (NM) immunotoxicity and genotoxicity. The hypothesis is that refined in vitro assays can replace in vivo testing of NMs. To achieve this goal, WP6 partners selected and refined the SOPs for the in vitro methods to be used within the project. The final goal is to select and adapt a set of in vitro assays for High-Throughput Screening (HTS). In addition, WP6 partners will also deliver samples of NM-exposed cells to WP10 for further analysis. The data generated will ultimately be used to develop the Nanosafety Classifier.

production in lungs which was supported by elevated expression of pro-allergic cytokines such as IL-13 and IL-5. Exploration of the early events by transcriptomics analysis revealed that a 4-h exposure to rod-like MWCNT caused drastic up-regulation of genes involved in innate immunity and cytokine/chemokine pathways. These data indicate that inhalation of certain types of ENM is able to induce an allergic airway inflammationlike reaction.

A first set of experimental data on immunotoxicity and genotoxicity of CuO NMs with different surface properties were recently generated in WP6. The preliminary results suggested that the toxicity of CuO NMs is dependent upon the surface functional group.

The ENM Safety Classifier

The cytotoxicity effects were confirmed in different cellular models (i.e. macrophages vs. lung cells) and were also correlated with the level of intracellular uptake for some cell types. Further experimental work focuses on the elucidation of toxicity effects of the entire panel of NMs in NANOSOLUTIONS using various in vitro methodologies that may have a good potential to be adapted for HTS. WP8 - Disease Models Fritz Krombach Inhalation of rigid multi-wall carbon nanotubes induces allergic asthma-like symptoms in mice The overall goal of WP8 is to identify the key physicochemical properties and surface modifications of engineered nanomaterials (ENM) that control their fate and biological and toxic effects in cells and tissues from susceptible individuals. For this goal, WP8 partners will use in vivo murine cardiovascular disease and asthma models as well as in vitro cultured human and murine endothelial cells or a 3D airway model with cells from healthy or diseased human donors. Finnish Institute of Occupational Health conducted a preliminary study in which pulmonary effects of inhaled rigid and tangled multi-walled carbon nanotubes (MWCNT) were investigated in healthy mice using two differently shaped materials. Results showed that after 4-day exposure, rod-like MWCNT, but not tangled MWCNT, induced eosinophilia and increased mucus

The knowledge gained from this study will lead way in exploring effects of ENM on pre-existing allergic airway inflammation for which the murine allergen-induced asthma model will be used. Current legislation that governs the safety classification of engineered nanomaterials (ENM) in Europe is complex. Each ENM has to be treated as a separate chemical and be individually tested to assess its safety characteristics. This means developing new materials using different nanoparticles is both time-consuming and expensive. If you take an element and add something to it, it changes its chemical composition and it becomes a ‘new’ material. In the existing legislative framework, each time a small modification is made to a nanomaterial it becomes a new material, and each new material needs testing, which is time-consuming and costly. Consequently, there is a real need for ways to reduce the amount of individual testing of ENM that is currently occurring, as this will help to reduce the cost of developing ENM-based products and speed up innovation.

If you take an element and add something to it, it changes its chemical composition and it becomes a ‘new’ material The main innovation of the NANOSOLUTIONS project will be the development of the engineered nanomaterial (ENM) safety classifier. This novel hazard profiling principle will provide a basis for understanding and defining the toxic potential of all types of ENM. It will be used by companies that manufacture ENM and by a regulatory community to manage, reduce uncertainty, and clarify the current debate, since it will provide the potential to effectively “de-classify” many types of ENM in many applications in terms of safety risks.

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Newsletter october 2014 The development of the ENM Safety Classifier spans a number of different disciplines. Here, we hear the perspective of three different work package leaders, who explain just what the classifier is and the experimentation work that is needed to develop it. Richard Handy

experiments will be used in the molecular biology and systems biology approaches being used in WP10 (Omics Methodologies) and WP11 (Systems biology analysis), the Nanosolutions project to provide the information needed for the ENM Safety Classifier. The WP7 team will be exposing a variety of organisms to the nanomaterials, with the test methods varying from organism to organism. With the fresh water marine organisms, for example, the nanomaterial is added to the water and a short-term study over a couple of weeks is made, during which the physiological effects on the organisms are monitored. Subtle changes are observed, as the team is not trying to produce overt toxicity, but instead trying to understand the organism response to the material.

The project is working with a completely new data set of organisms in order to ensure the quality of the data used in the systems biology models

Richard Handy (Cross species models, WP7) A fundamental part of developing a classification system for nanomaterial safety is to identify the toxic effects that different ENMs have across a wide range of organisms, from microbes to mammals, and on different body systems. These effects are assessed by their overall magnitude of toxicity – low toxicity, toxic, very toxic - using detailed experimentation. There are two main objectives. Firstly, the traditional hazard classification approach will provide both regulators and industry with information in a format they can use now, within the current regulatory process, meaning the experiments will have an immediate impact. Secondly, the samples from the

We will collect a high standard data set from these experiments so that we have the exact chemistry of each nanomaterial, as well as the environment (e.g. the bodily fluids), to which they are being exposed. The biological responses of the organisms at different levels, from molecular through to organism biology, will all be put into a systems biology model, where we are able to predict the biological hazard from the chemistry and behaviour of the nanomaterial. We will also be able to do the reverse, so from a known biological hazard, we will be able to work out what type of material we would need to generate that or to remove that hazard. The project is working with a completely new data set of organisms in order to ensure the quality of the data used in the systems biology models. This ensures that the model will be robust and the data will all be connected in some way. The molecular biology data will come from the same tissue as the histology and biochemistry data, and all will have been exposed to the same material. We are trying to remove all those methodology biases by doing original experiments in a very precise way that allows us to make a good model.

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Newsletter october 2014 Dario Greco

Dario Greco (Systems biology analysis, WP11) One of the main goals of the NANOSOLUTIONS consortium is to create an ENM classifier - a model that can predict the effect of any given nanomaterial. This is being done through what is known as machine learning; we are essentially training a computer programme to know what is dangerous and what is not. The more data we input into the computer, the better the programme becomes at recognising overlying patterns, after which it can extrapolate these patterns to create general rules that will allow it to predict how unknown nanomaterials will behave.

The problem lies in the fact that with such a huge amount of data, it is currently impossible to explore all the possible combinations. Using a computer that could evaluate each combination in one second with just 10,000 features, it would take 1079 years to go through them all. No computer powerful enough exists yet to carry out the task in a feasible time, and so a more holistic approach is needed to explore the vast solution space. A genetic algorithm is a special type of solution space evaluation algorithm that mimics natural selection, the key mechanism behind Charles Darwin’s theory of evolution. In terms of NANOSOLUTIONS, this involves grouping together features into, for example, 1000 random groups, each of which represents a solution. The predictive power of each of these solutions is then calculated, after which the least effective solutions are disposed of. The more effective solutions are then “mated” with each other in order to produce a new population of daughter solutions (much like the genetic rearrangement that occurs in natural populations), but this time drawing from a narrower and more powerful selection of features. This process is then repeated a number of times until only the most relevant features remain.

A number of the work packages in the NANOSOLUTIONS consortium are dedicated to gathering data on the biological effects of around 40 nanomaterials when they come into contact with living cells and organisms. This data is then collated in work package 11 and fed into a computer, which will then use this information so that when it is shown a new, unknown material, it will be capable of making an accurate prediction as to whether it is safe or not.

We know that we won’t be able to explore all solutions using our genetic algorithm, but we also know that this is probably the most effective way of finding the closest we can get to an ideal solution. Additionally to mating the best solutions, at each generation there will be other events such as substitutions, deletions, or insertions of new features, parallel evolving populations of smaller individual solutions mimicking viral infections, as well as local search operators that will work on optimising the best solution at each generation. Mimicking the evolution of natural populations will allow for the evaluation of many solutions, helping to explore the data as much as possible.

The real challenge lies in identifying the features of the animals, cells and nanomaterials that are useful in predicting the behaviour of the nanomaterials. There are hundreds of thousands, even millions, of chemical and physical features in living organisms that could potentially be useful to us, but we also know that the vast majority of them will not be. Our challenge is to mine these useful pieces of information out of a huge mass of data. In technical terms, this is a feature selection problem; we need to select the smallest number of features that can give us the most powerful information for accurate classification.

The algorithm will only need a couple of weeks computing time using a high-end desktop computer for an accurate predicting tool to be produced. However, the way the algorithm is designed will allow it to continue to evolve and become more accurate as new data becomes available. From a computational point of view, we would like the ENM Safety Classifier to also be a software package that can be used by future projects and data generators. They will be able to input new data so that the Classifier can continue to evolve. That is the beauty of using a dynamic system – it can always carry on learning when presented with new data.

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Newsletter october 2014 also maintain an awareness of the composition of the materials. That is something that takes a long time to work out.

Lang Tran

Lang Tran (Safety Classification,WP12) The development and application of ENMs promises many benefits to both society and the global economy. However, to ensure responsible development of this emerging technology, governance must be put into place to ensure that any potential risk posed by ENMs are fully understood and controlled. The regulatory landscape is constantly evolving for all substances and products. However, in the case of ENMs, challenges are greater because on the nanoscale, properties of a material relevant to its safety and effectiveness may differ from those on the bulk scale. The idea for NANOSOLUTIONS is that it offers a solution to the needs of industry operating within necessary regulatory circles to be able to classify the hazards of nanomaterials. Classification simplifies and rationalises the description of nanomaterials. Different materials can be put into different boxes depending on their physical and chemical characteristics in order to facilitate their management. If this doesn’t exist, each nanomaterial has to be dealt with on a case-by-case basis. The challenge for us is to do two things: not only will we classify the materials as being safe or not, but

WP12 will develop an ENM Safety Classifier that can be used for the reliable assessment of ENM safety. Based upon the resulting design and data requirements of the model, specification for a high throughput system for future testing and analysis of further refined or new ENMs will also be produced. The development and design of this will depend upon access to large volumes of data and information being retrieved and abstracted from the data repository in the systems biology work package (WP11), in which all the data produced by the partners, including the characterisation of engineered nanomaterials (ENM) and the study of their effect on cell and animal models by a number of in vitro and in vivo experiments, will be systematically collected, organised and analysed. There are a lot of materials that can be described as hazardous. But, if there is no exposure to humans, they can be safe; there will be no risk. But we are not doing a risk classification. We are doing a hazard classification, and in order to do that we need a lot of toxicology data. Data mining and neural networking will be used to find the traits that link the physical and chemical characteristics of the nanomaterials with these toxicology results. Those correlations will provide the basis of the Classifier. At present, the data that will be used for the algorithms that will provide the backbone of the classifier is still being collected. Once this is done, the Classifier will be presented to industrial experts in the form of a prototype computer programme. There are a lot people from industry, academia and regulatory bodies who have an interest in the production of nanomaterials, and so we will bring these people together for a workshop where we can demonstrate how the classifier works and also get feedback from them, so that we can improve on it and also demonstrate that we have a viable solution to their needs. Nanomaterials are being used in an increasing variety of industries, and it won’t be long before they are incorporated into the mainstream. 20 years ago we were talking about micro-technology. Now we just call it technology. We are now going through the same cycle with nanotechnology, and if the NANOSOLUTIONS classifier helps to speed up the process by giving people confidence in the safety of these materials, we will have achieved our goal.

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Newsletter october 2014 Events diary SENN2015 – 12-15 April 2015 The 2nd “International Congress on Safety of Engineered Nanoparticles and Nanotechnologies” SENN2015 will be hosted by the Finnish institute of Occupational Health in Helsinki, Finland, on 1215th April 2015. SENN2015 is a great forum for reporting and sharing the latest knowledge on the safety of engineered nanomaterials and nanotechnologies. The congress is recommended for those with an interest in:

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Nanomaterial identification and classification Exposure, transformation and life cycle Hazard mechanisms Risk assessment and management

The abstract submission deadline for SENN2015 is 31 October 2014. For more details on how to register to attend this exciting and productive confgress please visit the website. www.ttl.fi/senn2015

Systems Biology in Nanosafety Research Workshop – 09-10 November 2015

This project has received funding from the European Union’s Seventh Framework Programme for research, technological development and demonstration under grant agreement no 309329

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