The Virtual Cell Based Assay (VCBA) estimates time-dependent concentration of a test chemical in the cell and cell culture for a given in vitro system. The concentrations in the different compartments of the cell and test system are derived from ordinary differential equations, physicochemical parameters of the test chemical and properties of the cell line. The VCBA has been developed for a range of cell lines including BALB/c 3T3 cells, HepG2, HepaRG, lung A459 cells, and cardiomyocytes. The model can be used to design and refine in vitro experiments and extrapolate in vitro effective concentrations to in vivo doses that can be applied in risk assessment. In this paper, we first discuss potential applications of the VCBA: i) design of in vitro High Throughput Screening (HTS) experiments; ii) hazard identification (based on acute systemic toxicity); and iii) risk assessment. Further extension of the VCBA is discussed in the second part, exploring potential application to i) manufactured nanomaterials, ii) additional cell lines and endpoints, and considering iii) other opportunities.In order to replace the use of animals in toxicity testing, there is a need to predict  •VCBA as an alternative approach can be applied in the domain of nanotoxicology.•VCBA can support better testing strategies in acute toxicity.•Refinement of the VCBA taking into account biological oscillators could improve toxicity prediction.•Extensions of the VCBA can capture effects related to additional subcellular compartments. Reviews on the current status of alternative methods and their use in different sectors are given elsewhere  that these hazards will manifest themselves under defined conditions of exposure. While the management of chemicals involves policy decisions based on multiple considerations, the underpinning safety assessments are scientifically based. In particular, the risk assessment process consists of four steps: (i) hazard identification, (ii) hazard characterisation, (iii) exposure assessment, and (iv) risk characterisation. To determine the safety of chemicals, risk is evaluated as a function of both the exposure and hazard. Traditionally, hazard identification has been based on the use of animal testing, but these methods are gradually giving way to alternative  approaches for scientific, ethical and economic reasons. These approaches, typically lsewhere .in vivo toxic doses from concentrations that cause toxicological effects in relevant in vitro systems. The use of in vitro effect data  to predict in vivo toxicity presents two challenges: first, in analysing the results of in vitro experiments, since “nominal” concentrations do not represent the real concentration experienced by the cell . For example, in the case of caffeine, in the liver cell line HepaRG, the amount of test chemical that is freely available (dissolved) in medium after 24 h is 93% (results not shown). On the other hand Amiodarone shows an 85% and 5% affinity to lipids and plastic, respectively. In general the solubility, lipophilicity and volatility of the compound can influence the overall kinetics of the compound in an in vitro system. These uncertainties are chemical dependent and can be reduced by using a model that predicts the concentration of chemical in media, cell, plastic, etc. by considering only physicochemical properties of the test chemical and some parameters specific to the given cell line.In order to replace the use of animals in toxicity testing, there is a need to predict the cell ; and, sesessment . The usein vitro experiments) we have developed a Virtual Cell Based Assay (VCBA)To address the first of these challenges  and energy resources (lipids).3.A cell growth and division model that is based on a four stage based approach, with each stage corresponding to one of the four cell cycle phases: G1, S, G2 and M.4.A toxicity and effects model. The direct effects of a chemical concentration, C, on cell dynamics  are expressed by using the killing rate, kr, and the no effect concentration, NEC.As described in detail in this special issue , the VCBin vitro High Throughput Screening (HTS) experiments; ii) hazard identification (based on acute systemic toxicity); and iii) risk assessment. We also discuss the scientific and technical developments that will be needed to transform the VCBA from a research tool to one that is accessible to a broad community of safety assessors and decision makers. In this context, we consider the need to extend the applicability of the VCBA to simulate the fate and effects of a wider range of chemicals, including nanomaterials, which pose specific challenges due to the unique properties of substances at the nanoscale.In this paper, we discuss the current status of the VCBA and describe three applications that are already technically feasible, at least for selected compounds and cell types: i) the design of 2in vitro systems. A few of these studies also include a dynamic endpoint, such as cell viability. As can be seen, the models are based on both mammalian systems (primary cells and cell lines) as well as fish cell lines. Most studies have focused on single dose exposure scenario, with a few exceptions dealing with repeated exposure. Test compounds include pharmaceuticals, industrial chemicals, and pesticides. The The VCBA model consists of ordinary differential equations whose solution allows the estimation over time of the dissolved concentration of a chemical in cell culture as well as the internal concentration in the cells. The VCBA developed by 33.1High-throughput screening (HTS) has proven to be a useful technique used in drug design and may be applied in biological and chemical sciences . Using r1)Optimizing the efficiency of HT chemical-design methods ; for ins2)Extracting additional information from the HTS process. When screening multiple compounds applying HTS, this allows only one experimental design and does not allow modifications on a compound by compound basis which otherwise would be included to capture specific behaviour. However, this extraction can be obtained after the HTS is done, as the data of the HTS experiment allows the full parameterisation of the VCBA model for each compound which can then be used to model the compound specific exposure.3)in vitro kinetics.Helping the interpretation of unexpected False Negative effects within the experiment as this could be due to the The current VCBA can support HTS by:3.2in vitro experimental setup . Simulations of cell viability/cytotoxicity can be performed if the toxicodynamic part of the VCBA has been calibrated with in vitro experimental data. Information on the kinetics in an in vitro test system are highly relevant to refinement of any additional in vitro tests, and their application could help to better understanding and translating information to address acute oral toxicity  and 32 chemicals including polyaromatic hydrocarbons, cosmetics, and drugs  that span molecular weight from 71 to 822 g/mol and octanol-water partition coefficient from − 3.7 to 7.6. The VCBA simulates cell viability, in addition to the distribution of the chemical in an toxicity .in vivo endpoint routinely required for hazard identification and classification purposes under multiple pieces of EU legislation  in the test population within a 14 day observation period .Acute systemic oral toxicity represents a complex n period . The outin vivo tests do. The use of basal cytotoxicity to predict acute oral toxicity in vivo has been extensively studied (http://www.acutetox.eu/) evaluated the use of a battery of in vitro and in silico tests to predict acute oral systemic toxicity. While even the use of multiple, combined in vitro methods did not allow the correct identification of the different toxic classes according to CLP, some of the organ specific methods, such as the brain aggregate model, could be used to alert for specific target organ toxicities  assay, are useful in identifying non-classified (LD50 > 2000 mg/kg) substances according to CLP. Oral LD50 values can be derived from IC50 values that are produced in the in vitro 3T3 NRU experiments  assay. Simulations have been made for 32 compounds, after single exposure, for which available experimental data to perform downstream extrapolation were available . For eacin vitro set up. Knowing the time-dependent concentration of chemical at different levels of the in vitro test system can be of great help in detecting false negative predictions as there are cases in which a negative result is due to the chemical not reaching the target at a sufficient concentration due to its fate in the system . In this regard, and in the context of the published EURL ECVAM strategy on how to reduce, refine and ultimately avoid the use of animals for acute mammalian systemic toxicity testing  model. This may contribute to the chemical risk assessment process. As outlined above, many kinetic considerations have to be taken into account when extrapolating from an in vitro nominal concentration to an in vivo concentration. Thus, the kinetics of an in vitro test system has to be considered, as well as the kinetics of an in vivo exposure, when trying to translate an in vitro dose to an in vivo dose.For prediction and extrapolation of in vitro data, for the substance to be assessed or structurally similar substances. For example, the publicly available ToxCAST library , or whether further assessment is necessary, in which case it should also carefully be considered whether it would be possible to use any read-across assessment.The starting point in a completely animal-free risk assessment is to analyse expected exposure scenarios and predict the relevant dose and route of exposure. Then to better understand possible effects and target organs, it is necessary to collect current knowledge and search for already existing animal and  library  is incluDepending on the hypothesis regarding target tissues, the VCBA could be adapted to predict expected adverse effects. The VCBA would then assist in predicting any expected adverse effect either a stand-alone assessment, or possibly to strengthen a read-across case.in vitro results to in vivo (human) effects, whole-body toxicokinetics  needs to be considered. Depending on the route of administration, oral, dermal or inhalation, the chemical first has to cross the relevant barrier to be absorbed into the blood stream if it is to have a systemic effect. This information can be provided by available QSAR models on absorption potential  modelling approach can be used to perform IVIVE (e.g. liver Cmax) with the VCBA model, which allows the application of both forward and reverse dosimetry. The forward approach means that when exposed to a given external dose, the joint model predicts the corresponding effect on liver cell viability. The backward approach can be used to estimate from a given cell viability the corresponding external dose. In addition, a user-friendly tool could be developed by implementing the PBK/D model into a KNIME workflow as currently done for the VCBA .The application of biokinetic/biodynamic models is an important means of extrapolating ituation . As suchrm IVIVE . Additiothe VCBA . This toWhen going through the different steps of the assessment, it is a necessary to accompany each step with an analysis of the uncertainty, to estimate the reliability in the final predicted point of departure.4While the previous section addressed applications that are already feasible with the current version of the VCBA, this section explores possibilities for future versions of the model.4.1e.g. flame retardants, cosmetics, food packaging, paints), since materials at the nanoscale exhibit novel physical and chemical properties which may enhance the effectiveness of the embedding products  are materials with one or more dimensions in the nanoscale,  cellular effects. As reported below, size distribution and surface characteristics of MNs play an important role in the fate of MNs in  density , and suc for MNs . Other ee of MNs : thus, iin vitro studies to generate relevant input parameters. The MN-specific fate processes are briefly explained below.However, for the VCBA to be applicable to MNs, additional fate processes need to be accounted for in the mathematical model, which also implies a need for targeted in vitro systems is affected mainly by the following fate processes: diffusion, gravitational settling, agglomeration, dissolution, and cellular uptake. These are affected not only by the characteristics of particles , but also for those of the surrounding medium . These particle-medium interactions, therefore, influence the fate and intracellular  concentration of MNs.The fate of MNs in The current version of the VCBA  takes in1)e.g.  and of properties of the fluid ; for particles including MNs size plays a role and passive diffusion can be calculated based on the Einstein-Stokes equation (e.g. .2)in vitro fate models applied to MNs There is evidence that size, surface charge, shape , surfaceFurthermore, considering the models specifically applied to MNs the following processes would need to be implemented in the VCBA:4.2in vivo systemic toxicity tests it would need to be extended to a broader range of cell lines and endpoints. This is discussed here in relation to acute systemic toxicity, as an endpoint that likely involves more than one target organ. It is worth noting that while this discussion is focused on the identification of other cell types to be implemented in the VCBA; there are of course other variable experimental parameters such as new medium composition and different types of plastic that may be considered in future versions of the VCBA.In its current form the VCBA has been implemented for single (and sometimes repeated) exposures in a limited number of cell lines. In order to use the VCBA to support the replacement of in vitro data for these cell lines in order to train the VCBA model. It has been suggested that acute systemic toxicity could be due to organ specific effects in addition to basal cytotoxic effects . Thus imin vitro test methods for systemic toxic endpoints. Each organ is comprised of several cell types that work together to make the organ function. In terms of modelling the function of the organ, is it then advisable to model only the cell type that makes up the largest portion of the organ? For example, the liver is comprised to 60% of hepatocytes when determined by absolute cell number, but make up 80% of the organ by volume. In some cases that might be the ideal cell type to model since its function might determine the overall function of the organ. However, for other organs/systems, such as the immune system it is not so clear which cell type would be most representative/most vulnerable to toxic insult/ or which feature should be modelled. For most of these organs, human and animal cells for at least one cell type, either primary or immortalised, are available commercially to be cultured and tested in the laboratory. For blood cells it is possible to obtain whole human blood from donors. In addition, advances in stem cell research have made the in vitro production of red blood cells from stem cells a possibility (in vitro data needs to be generated in order to train the model.sibility . The avain vitro test systems such as the multielectrode array (MEA) chips, which record electrical activity from cultured cortical neurons in the presence of test substances, could provide important functional information in this context  that should be investigated. While cytotoxicity still plays a role in target organ toxicity, it is also necessary to consider whether target organ specific mechanisms of toxicity play an additional (and possibly more important) role. For example, the neurons of the central and peripheral nervous system communicate through the generation and propagation of action potentials and the release of neurotransmitters. Toxicants can interfere with this basic function of the cell at several levels: the generation and propagation of the action potential is governed by the opening and closing of ion channels, with the ion balance across the membrane being restored by transporter enzymes; neurotransmitters are produced in the neuron and released from the nerve ending where they interact with their target receptors before they are cleared from the synaptic cleft. Interference with any of these steps would inhibit the normal functioning of the nervous system and thus could lead to death or severe adverse effects. Ongoing work is focused on trying to determine the mechanisms that lead to acute systemic toxicity. For example, information gained from  context . In term4.3i.e. the virtual cell, in larger scale models of organs and the human body; 5) incorporating biological oscillators; and 6) accounting for the influence of new medium composition and plastic/coating.1)e.g. via Quantitative Structure-Property Relationships (QSPR). In the worst case scenario, two different users that provide parameters for the same chemical obtained from different QSPRs could obtain different results. In order to tackle this issue, a node that will calculate the necessary parameters from just the name, CAS number, or SMILES codes of the test chemical will be added to the current workflow. The node will also allow the user to provide its own parameters if desired. The definition of model applicability domain (AD) is key for the use of QSARs and QSPRs as it determines whether the QSAR/QSPR is appropriate for the test chemical, and whether there is confidence in the resulting prediction. The fact that a physicochemical property is obtained from a QSPR for which the test chemical falls outside the AD does not necessarily imply that the value is wrong, but it means that the model was not optimized for such chemicals and that the quality of the prediction is uncertain. In order to gain confidence in the physicochemical predictions and the overall VCBA result simulation, the new implementation will also provide an alert indicating the test chemicals that fall outside the AD of the corresponding QSPR so that the user is aware of the increase of uncertainty of the result. Such an automation of the calculations will benefit the model in two ways: a) it will allow the model to be used in a batch mode, b) will reduce the uncertainties caused by the need to use third party software to obtain parameters. Thus, such a node will contribute to a better user experience and to increase the traceability of the model. The parameters needed to run the VCBA are the molar volume at its normal boiling point (Vb), Henry law constant (H), octanol-water partition coefficient (Kow), and the air and water degradation constants (KAW). The parts of the model that make use of these parameters can be found in another article that forms part of this special issue in vitro test systems and cells depends on additional physical processes. More generally, additional subcellular compartments could also be considered, for example, just as a generic mitochondrial compartment has been implemented, a generic lysosomal compartment could be added. This could be useful to capture the direct and indirect effects of chemicals, including MNs, on the lysosomes. Similarly, consideration could be given to adding compartments for the nucleus to simulate genotoxic effects, and the endoplasmic reticulum, to simulate for example effects on protein folding in vitro readouts) related to additional subcellular compartments. Another suggestion relates to the use of NEC and kr as the effect parameters. In the current version of the VCBA, the dose-response of a chemical is expressed as an exponential function of the exposure concentration (i.e. are “early” responders) and the differences in sensitivity follow a log-normal distribution. Thus, a possible extension would be to incorporate a probabilistic term to capture this natural variability in the susceptibility of cells to a given concentration.Possible extensions of the toxicity and effects model could be to capture effects (ntration . This as4)in vitro experiments are translated into different transporter and enzyme activities, which are thereafter distributed in the liver e.g. circadian clock, embryonic oscillators), and metabolic and biochemical oscillations in the order of minutes (glycolytic oscillations) or seconds . Oscillations are central to biology across different temporal and spatial scales and Oscillations in biological processes are periodic fluctuations ranging from hormonal oscillations with periods of days and months, to genetic fluctuations in the range of hours (6)in vitro system, the plates are increasingly being coated. Basic synthetic polymers, such as poly-d-lysine (PDL), have also been used as coatings to create a positive charge on polystyrene which, for some cell types, can enhance cell attachment, growth and differentiation, especially in serum-free and low serum conditions  full automation of the model; 2) extensions of the fate (kinetic) models; 3) modifications of the toxicity (dynamic) model, and 4) embedding the cellular component of the VCBA, 5in vitro HTS, adverse outcome pathways (AOPs) and a range of modern computational tools including the VCBA. The literature includes many variants of this emerging safety assessment framework  but also by integrating experimental and computable properties in the form of mathematically based phenomenological models. PBK and VCBA models are thus central to this paradigm shift, which is why progress in the scientific and technical development of these approaches needs to be continued, along with efforts directed at promoting their uptake and acceptance among scientist and regulators.Modern toxicity testing is moving away from its traditional reliance on the observation of apical effects in animal studies towards mechanism-based testing strategies that utilise techniques such as ramework . In the in vitro toxicity data, as well as the design (dosimetry) of in vitro experiments. As far as the authors are aware, these models have not yet been used to directly support a regulatory application, such as hazard characterisation, IVIVE, or risk assessment. This paper has identified some of the scientific and technical opportunities and challenges associated with possible applications of the VCBA in regulatory chemical risk assessment. While some of these challenges will require considerable further in vitro experimentation, it can be seen from the papers in this special issue that considerable progress has been made in the past five years, not only in developing the computational models and tools, but also in making them available and deployable in a user-friendly form.The VCBA, and VCBA-like models, are already being used as research tools to support the interpretation of Transparency document.Image 1


The World Health Organization (WHO) has ranked musculoskeletal diseases among the top ten leading causes of disability in the Middle East. The situation in the Middle East is unique as, although seventeen countries are geographical connected, there is considerable variability in the standard of rheumatology care and research between the countries. There are several factors contributing to this variability: allocated government resources to healthcare services, country demographics, the implemented healthcare system, the number of qualified rheumatologist, political stability, and the immigration to and from these countries. In Israel, despite being one of the most developed countries in the Middle East, there are still many clinically challenging issues frequently encountered, including that the number of board-certified rheumatologists is approximately 150, thus making the waiting time until an appointment often too long. In some areas of the country, the time until the initial visit to a rheumatologist might exceed 3 months. In addition, regarding inflammatory disorders, there are still formal barriers to the rapid use of biologicals; for instance, in order to be entitled to biological use, rheumatoid arthritis patients need to fail on three oral disease-modifying antireheumatic drugs (DMARDS), which includes methotrexate. These local guidelines are economically driven and have not been updated since 2003.Government expenditure on healthcare services influences the practice of rheumatology in the Middle East. In the less prosperous countries, there is limited access to rheumatologists and treatment. On the other hand, in some booming countries the government predominately funds rheumatology services or has fully implemented a health insurance system that provides the same coverage. This grants easy access to rheumatologists and treatment. The stable environment and work opportunities in these countries in the Middle East have attracted millions of immigrants . Hence, There is an opportunity for optimizing the access to rheumatology care in those countries. It is the role of the key opinion leaders in the Middle East to enlighten the decision makers of the growing burden of rheumatic diseases and the cost-effective outcomes of providing optimum rheumatology care to their nation in the long run.The postgraduate training in Western countries and the availability of training programs in different Middle East countries have helped in increasing the number of practicing rheumatologist in the last few decades. However, the current number is insufficient compared to developed countries. The ratio of rheumatologists per 100,000 population ranges from 0.3 to 0.89 . The lacGiven the current burden of musculoskeletal diseases and the expected increase in the near future, health authorities in the Middle East should include a rheumatology specialty in their workforce planning. Substantial measures should be directed to undergraduate and postgraduate training to attract students and newly qualified physicians to the rheumatology specialty.Ben Zvi et al.  from theThese authors have also collaborated with colleagues from Turkey and Italy  in definMoving on from FMF to other rheumatic diseases, Ofer-Shiber and Molad at the Rabin Medical Center in Petach-Tikva reported that a C-reactive peptide (CRP) level ≥0.9 mg/dl at diagnosis in patients with psoriatic arthritis predicted an earlier need for tumor necrosis factor (TNF) blockers in order to achieve disease control . The rolIn Tel-Aviv, at the Sourasky Medical Center, Ablin et al. reported that magnetic resonance imaging (MRI) findings consistent with sacroiliitis were found among a significant proportion of patients diagnosed with primary fibromyalgia . The samAt the Rambam Medical Center in Haifa, Balbir and collaborators from the EUSTAR group observed that, in patients with systemic sclerosis that present early after the onset of Raynaud’s phenomenon, the manifestation of 50% of all incident organs occurs within 2 years simultaneously, rather than sequentially .Zisman, from the Carmel Medical Center in Haifa, reported the risk of herpes zoster was significantly increased with age, treatment with steroids, and combination of antiTNF-α agents and c-DMARDs, but not with c-DMARDs or antiTNF-α therapy alone in patients with psoriatic arthritis . Mader eShaare-Zedek Medical Center researchers, in collaboration with the Hadassah Medical Center investigators, have demonstrated lower overall incidence rates of vasculitis among the Jewish population despite rising incidence rates of granulomatosis with polyangiitis and microscopic polyangiitis . In the Generally, there is quantitative and qualitative improvement in rheumatology research produced from these countries in the last five decades, as well as collaborative research work between countries within the region . We haveThe differences between Middle Eastern countries in the published research can be attributed to the number of rheumatologists and the number of academic institutes within a country, political stability, the availability of funds, research facilities, an appropriate environment to conduct research, and the incentives for research.In the last decade, governments in the Middle East have created research grants and awards to facilitate research. Pharmaceutical industries are vital partners in this transformation by spreading research skills among Middle East rheumatologists and providing funds for research. Furthermore, healthcare systems in Middle Eastern countries have an important role to play in promoting research culture, allocating time and incentive for those involved in research. Needless to say, it is important to mention that optimizing collaborative research projects between Middle Eastern countries will help in producing high-quality research from the region .The rheumatologists in the Middle East strive to improve the care of patients with rheumatic diseases despite the prevailing challenges in some countries. A few countries in the Middle East have contributed, and will continue to contribute, research and knowledge in the field of rheumatology. There is a great opportunity to improve research in some countries to reflect the high standard of care provided within the region.


The Adverse Outcome Pathway (AOP) concept is a knowledge assembly and communication tool to facilitate the transparent translation of mechanistic information into outcomes meaningful to the regulatory assessment of chemicals. The AOP framework and associated knowledgebases (KBs) have received significant attention and use in the regulatory toxicology community. However, it is increasingly apparent that the potential stakeholder community for the AOP concept and AOP KBs is broader than scientists and regulators directly involved in chemical safety assessment. In this paper we identify and describe those stakeholders who currently—or in the future—could benefit from the application of the AOP framework and knowledge to specific problems. We also summarize the challenges faced in implementing pathway-based approaches such as the AOP framework in biological sciences, and provide a series of recommendations to meet critical needs to ensure further progression of the framework as a useful, sustainable and dependable tool supporting assessments of both human health and the environment. Although the AOP concept has the potential to significantly impact the organization and interpretation of biological information in a variety of disciplines/applications, this promise can only be fully realized through the active engagement of, and input from multiple stakeholders, requiring multi-pronged substantive long-term planning and strategies. Currently there is a need to determine the potential biological effects of tens of thousands of existing and new substances in a cost-effective and timely manner, but resources for chemical testing currently are either static or decreasing. Hence, there is increasing reliance on approaches to efficiently generate data concerning the possible biological activity of chemicals through use of in silico models, in vitro assays, and short-term in vivo tests emphasizing molecular and biochemical measures of effects . This crThe Adverse Outcome Pathway (AOP) conceptual framework was developed to serve as a knowledge assembly and communication tool to facilitate the transparent translation of mechanistic data into outcomes meaningful to chemical safety assessment . SpecifiAs the AOP framework has evolved in the context of regulatory toxicology, it has become recognized that the potential stakeholder community is broader than those involved in chemical safety assessment, as illustrated in Our aim with this paper is to present a strategic vision to help ensure that the AOP framework can realize its full potential for multiple stakeholder groups. The paper first details the unique features and qualities of the AOP approach and of the AOP framework, and goes on to profile a variety of stakeholders, describing how they stand to benefit from the further development of the AOP framework and its application to solving their problems. Our analysis includes consideration of the core obstacles to a full, long-term engagement of the stakeholder community with the framework. Finally, we conclude with a set of recommendations regarding the steps that need to be taken to address these challenges, and to ensure development of the AOP framework as a sustainable tool for the application of our scientific knowledge of living systems in numerous decision-making contexts.This paper stems from a Pellston workshop held April 2–6, 2017 in Cornwall, ON, Canada. The workshop was coordinated by the Society of Environmental Toxicology and Chemistry (SETAC), and financially supported by multiple governmental and business organizations. Pellston workshops typically assemble 30–40 (invited) experts on a given topic in the environmental sciences, who are charged with defining and proposing paths forward to address challenge(s) in the topic area of interest. This Pellston workshop focused on a number of issues associated with AOP science and implementation, including the development of a “road map” to promote sustainable use of the concept. Specific needs addressed at the workshop were identified through a global horizon scanning effort that gathered input from scientists and risk assessors/managers from throughout the world . In all,2Over the past several decades the global scientific academic, pharmaceutical, chemical and personal care product communities, as well as government agencies, have invested billions of (US) dollars generating biological information. The goal has been both to extend basic scientific knowledge of how biological processes causally unfold and to inform the development of less harmful chemicals and more effective disease diagnosis, prevention and treatment. Traditionally, this information has been deposited in scientific publications, but the sheer number of papers, compounded by the fact that the information is often conveyed in unstructured and inconsistent ways, makes it difficult to access, share and integrate. Initiatives to summarize and compile this information into centralized data repositories have been only partially successful, as they i) do not include all publications, ii) remain compartmentalized, and iii) use different ontologies. These disconnected databases have resulted in the next level of “silo-ed” information. The rise in bioinformatics, including collections of “omics” information and associated relational databases, has also produced a vast amount of potentially useful, but still ultimately difficult-to-access information. An additional complication is that lack of curation makes it difficult to assess the quality of much information .The goal of the AOP framework is to compile and synthesize this wealth of biological information such that it can be transparently and efficiently employed for decision-making. The AOP framework potentially goes beyond biological pathways, but will improve our understanding of chemical toxicity in relation to the ecological impacts of chemical contaminants in air, water, soil and food as well as impacts to human health. http://aopkb.org, accessed 20 Sep 2017). Other AOP-KB packages include Effectopedia, AOP Explorer, and the Intermediate Effects Data Base  the AOP knowledge itself, ii) the way the knowledge is assembled/treated and iii) the potential applications and impact the knowledge can have. The primary type of knowledge captured by the framework is obviously related to toxicological processes and mechanisms. However, in a description of an AOP there is also valuable information on test methods and models that can be used to measure or predict the KEs that comprise an AOP, and information on chemical initiators and their relevant chemical properties linked to the MIE. Thus when one describes the type of knowledge covered by the AOP framework, it is important to highlight the availability of information on methods and chemicals associated with measurement of the KEs.Some of the key attributes of the way knowledge is managed within the AOP framework are depicted in Clarity – the description of an AOP is highly structured and follows a particular format and well defined conventions and guidance , different hazard/risk assessment scenarios, varying amounts of available data and/or levels of data collection capabilities  and differing numbers of chemicals for which they are responsible (a few ranging up to thousands). Risk managers then have to use the output of a risk assessment to make decisions on whether measures should be taken as required by relevant laws and associated protection goals. Decisions might include restricting the use of a hazardous chemical or requiring its prior authorisation, and often include consideration of socio-economic consequences of intervention, which can be considerable. Accordingly, the utility and perception of value of the AOP framework will vary for different members of this stakeholder community. For example, assessors responsible for rapid processing of large numbers of chemicals with limited data may rely upon the AOP framework for the utilization of in silico and/or in vitro data for hazard profiling, which would serve as the basis for identifying chemicals of potential concern and prioritizing them for further testing or assessment. This scenario is exemplified by regulators involved in the US Environmental Protection Agency's (EPA)'s endocrine disruptor screening program, who use the AOP framework as a basis for prioritizing chemicals for possible in vivo testing based on their endocrine activity . Another3.2https://www.epa.gov/tsca-inventory, accessed 20 Sep 2017), REACH Regulation in the EU  through a better understanding of pathway plausibility and biological read-across for targeted data-gap filling, etc. . In the In the mid to longer term, the AOP approach has the potential to be used to help identify relevant pathways in the derivation of biological pathway altering doses  or for h3.3The pharmaceutical (both human and veterinary) and agrochemical industries are primarily focussed on developing novel chemical entities for the purpose of medicinal and agricultural usage. Here a requirement is to efficiently identify compounds that may be harmful to human, animal health or the environment. The motivations are significant, as a number of chemicals used, for example, in commonly prescribed medications may be hazardous to the environment  they have vocational motivations as scientists who aspire to advancing knowledge in their field and teaching students; and ii) they have career motivations. For the latter, publications are a main indicator of scientific productivity and performance, and necessary for career progression in terms of positions, resources, and promotions. While there is some variability depending on country and institutions, in general there is increasing pressure on academics to demonstrate the impact of their research beyond academia. If the importance of impact on policy-making and industrial innovation continue to grow, it could provide an important incentive to contribute to AOP development.Academic scientists are in a critical position to make a significant contribution to the uptake and establishment of the AOP framework through training new generations of researchers and graduates who will be employed in industry, policy and regulation. For example, academics are able to promote AOPs via course curricula and incorporating the AOP-KB and applications in toxicology and pharmacology courses. As exemplified in a teaching monograph by www.eu-toxrisk.eu). The project aims to employ non-animal methods to develop new approaches to assessing the safety of chemicals in a variety of sectors, addressing in particular their potential chronic, developmental and reproductive effects.An example of engagement of academic stakeholders with the AOP framework is the large European research consortium project SOLUTIONS , which h3.7NGOs give voice to concerns emerging from civil society and to the interests of those who are affected by chemical safety assessments and resulting decisions. Apart from this common goal, individual NGOs typically have differing, sometimes contrasting interests, and therefore have different incentives for engagement. Despite this, they are potential users of AOP knowledge and participants in developing and using the AOP framework.Engagement of the animal welfare NGOs stems from the desire to move away from the use of animals for chemical evaluation and biological research . A compahttp://endocrinedisruption.org/, last accessed 20 Sep 2017). Thus far, there has been only a somewhat limited engagement of environmental NGOs, with a few notable exceptions. Specifically, since pathway-based information is used in research into the effects of pesticides on the environment  see  below.4.2While we take the view that the AOP framework is the most promising and most realistic answer to the challenges of chemical safety testing, it is necessary to highlight possible risks and burdens, so as to address them.Policy-oriented organizations (like OECD) play a critical role in sustaining the credibility of the AOP framework and in providing assurance that AOPs can improve decision making. However, there are significant reputational risks, should there be non-optimal decision-making based on AOP information or knowledge. This risk increases as the popularity of the framework and the number of AOPs grow and the quality control process becomes itself increasingly crowdsourced and not limited to a tight community of highly dedicated scientists, as currently is the case.Governmental and industrial organizations involved in safety assessment are key players in securing the actual usefulness and hence the ultimate success of the AOP framework. But they are also those facing the most serious risks in adopting them, since they could be held accountable for harm to consumers, the environment or business interests. There needs to be confidence that decisions based on AOP considerations are as good as conventional approaches.Serious obstacles may come from lack of adequate incentives for developing and adopting new testing methods. For those in industry, the pressures to deliver assessments to an existing regulatory standard mean that there are few resources to consider alternative and new approaches. With significant initial development costs required to demonstrate robustness of a new approach, both in terms of intellectual and financial input, this means that often corporations are not in a position to support significant changes in an approach until it has already been sufficiently vetted by other organizations. Regulatory agencies are also resource- and time-strapped, such that adding another source of information to be weighed against other types of scientific evidence can make assessments more complicated.4.3AOPs can serve a number of functions ranging from, at their most basic, assembling available data as a basis for understanding what is and what is not known about a given pathway, to relatively complete, quantitative depictions of a pathway that could be applied to risk assessment. Accordingly, From a science point of view, the issue of quality control is tightly interconnected with that of publication and peer review: the “gold standard” for quality assurance of research results. Accordingly, the AOP community has established a multi-level system of reviewing well-developed AOPs, internally and externally. For example, OECD-endorsed AOPs have a level of peer review equivalent or superior to that of a published paper. OECD endorsement is recognized as bringing with it an important reputational added value, especially by the regulatory community. Career wise for the academic community, there is still a preference for publishing in known, and high impact journals, even though they may recognise that the level of peer review is equivalent. The review process is also an important way of building the AOP community, as it draws in more contributors than those who develop and upload AOPs. However, there is a question whether such a work-intensive process can be sustained in the long term, especially in consideration of the lack of incentives for some of the reviewers. An increasing number of AOPs uploaded onto the AOP-KB would also make greater demands on the pool of qualified, often over-committed reviewers.4.4The AOP framework originally arose from the discipline of ecotoxicology to support regulatory decision-making . This meSpecifically, AOPs can be assembled into ‘non-linear’ networks which at times can be a better reflection of reality and thus be a focus of application . It mustAn important challenge in stakeholders dialogue arises because of conflicting perceptions of the utility associated with different levels of detail included in the AOP-KB. The tension between development and communication in the AOP-KB is particularly prominent for different stakeholder communities reflecting “users” as opposed to “developers”, with the former often requiring less complexity than the latter.4.5There are legal issues around intellectual property rights (IPR) in the production and dissemination of AOPs that deserve attention and action. Since many scientists need first to publish the results of their research in academic journals, steps should be taken in order to enable scientists to reuse their work for contributing to or authoring AOPs without infringing on copyright ownership. Conversely, there is a need to enable researchers to reuse parts of their research that have been uploaded on the AOP-KB in publications without violating anti-plagiarism rules. In addition, AOPs published by the OECD do not specify what kind of copyright, if any, protects the intellectual work and which uses are authorised. This is striking when intellectual work freely accessible through the Internet often comes with clearly specified licences. The same applies to what is available through the AOP Wiki, which by its nature as a wiki platform, is often associated with open access and free sharing or reuse of content. Authors need to have a clear perception of the kind of IPR protection available for work shared through the AOP-KB.AOPs are not only a tool for systematically organizing and curating scientific knowledge and indicating knowledge gaps that need to be filled, but have been promoted and are perceived as an important instrument for regulators dealing with protecting the environment and human health. As such AOPs may be used in regulatory contexts where the stakes and uncertainty are high, fierce controversies may arise and stakeholders may become polarized, as exemplified by recent activism around endocrine-disrupting chemicals in the EU .An obvious ethical issue that AOP usage brings to the fore is the widespread desire to reduce or ultimately replace animal testing for chemical safety assessment. As noted above, this is a strong motivator for animal welfare NGO stakeholders, but it may also result in tensions with other ethical priorities, for example, associated with perceptions regarding the role of animal tests in the effective protection of human health and the environment. The interplay and potential conflicts between these ethical priorities needs further analysis.https://ec.europa.eu/programmes/horizon2020/en/h2020-section/science-and-society, accessed 20 Sep 2017). A robust consensus about the appropriate conditions and domains of application of AOPs is therefore needed. In other words, different stakeholders need to agree on when  AOPs are fit for what purpose  handles several practical logistics associated with the wiki module of the AOP-KB, including funding an open-access server, and providing support to ensure that information supporting new or existing AOPs is entered into the system in a manner consistent with established OECD guidelines. However, the SAAOP is comprised of a comparatively small group of people, none of whom are primarily dedicated to AOP-associated governance or coordination. This is not a sustainable long-term situation.As the AOP framework continues to evolve, governance and coordination of the effort in the future would ideally involve all stakeholders as much as possible. Objectives of overall governance arguably would be best achieved through an impartial organization that broadly represents the various stakeholder groups. This could be accomplished through a dedicated professional society charged with coordinating advances in AOP knowledge and communication for the common good of all. Currently, the Society for Advancement of AOPs  has come via resources provided from a relatively limited number of stakeholder groups, principally the US EPA and the European Commission, often under the auspices of the OECD. Since there is no assurance of continuing availability of resources from these organizations, the development of a stable, long-term “business plan” for provision of resources to cover these direct costs including dedicated personnel needs to be defined. Additionally, there needs to be a formal vision to attract and involve other institutional investors.5Many challenges that have been discussed in the previous section can be addressed in several ways. Solutions are proposed in this section; however, there is not always a one-to-one mapping between challenges and solutions. Instead, there are cross-cutting solutions to meet multiple challenges .5.1https://www.ncbi.nlm.nih.gov/geo/, accessed 20 Sep 2017). For example, those involved in AOP governance could discuss the possibility of making deposits into the AOP-KB, or using the AOP framework standardized format for depositing in other databases mandatory with the publication of AOPs in journals such as Environmental Science and Technology, Environmental Toxicology and Chemistry and Toxicological Sciences, all of which publish AOP-oriented papers. An example of how this can be done is illustrated by The AOP framework is part of a rapidly changing publishing infrastructure that is affecting almost all sciences . For exaMolecular Ecology Resources 1755-0998, accessed 20 Sep 2017) and Scientific Data  in which the authors publish their work by depositing the research data following a stringent format requirement, and provide detailed descriptions of methods used for data collection, quality control and assurance, processing and analyses. These database-only journals provide a new framework for data sharing, thereby accelerating the pace of scientific discovery. Such journals could be sponsored by technical societies whose membership includes large numbers of AOP stakeholders, such as the Society of Environmental Toxicology and Chemistry (SETAC) or the Society of Toxicology (SOT).Other possibilities in next generation publishing could include developing a database-only journal to complement the AOP-KB. There is an increasing trend towards database-only journals such as Finally, it also might be possible to establish a journal whose main focus is AOPs, though this could be perceived as a relatively narrow field of science thus limiting the number of contributors and hence restricting its growth. Since the development of any new journal would take time and resources, and obtaining an ISI Impact Factor can take at least 3–5 years, this option could discourage academics from submitting to an AOP-only oriented journal during its early development phase.For any of these solutions, it is paramount that the data format be compatible and interoperable, so that information can be added onto the AOP-KB with minimum effort. Steps to ensure this are being undertaken by the OECD.5.2SAAOP.org, but other centralized repositories for this type of information are needed.Outreach to various stakeholder groups requires different strategies and educational materials. Some common and overlapping characteristics of stakeholders that will affect communication strategies include whether they are AOP developers, users of AOP-based information, or simply interested in understanding the concepts and the science behind AOPs and their use. A list of existing guidance documents and training presentations is available from To date, most outreach has focused on scientific training of current and potential AOP developers, and a number of materials have been created for this purpose  DevelopeFinally, as university teaching evolves from learning content to mastering problem-solving tools, the AOP concept offers an opportunity for a structured approach to environmental toxicology and ecotoxicology. Curricula may include practical courses on the development of AOPs as team-building literature review courses or may apply the conceptual framework of logic and uniform structuring of information. New venues for spreading of AOP information that remain to be explored are the emerging massive open online course (MOOC) that range from free availability to formal inclusion into universities' curricula . At writ5.3Different stakeholder communities have different needs regarding how much information they require, how they prefer it presented, and how they most productively could interact with the AOP-KB and approach. Research shows that virtual collaborative environments require significant buy-in from the communities for which they are designed, but this depends on a number of factors, not least of which is the active participation of the communities in designing and developing the interface of the knowledgebase . The ‘bu5.4Although AOP development principles are well described in guidance documents  and in tCase studies are also important to illustrate the application of AOPs in different chemical risk assessment contexts  and are translational surveillance exercise: an active monitoring of decisions made on the basis of AOPs. The goal would be to promptly detect and flag up inappropriate uses that can have negative consequences for safety and bring the AOP framework into disrepute. Moreover, this would provide very useful feedback for improving the communication strategy of the AOP-KB.AOPs at different stages of development and backed by different levels of evidence are fit for different purposes . For ins5.5Various alternatives are possible in terms of governance. While the role of the SAAOP could be expanded to support a broader stakeholder community, an alternative option might be to engage other professional societies with a stake in AOPs. For example, the SETAC and SOT are well-established scientific organizations representing a diverse membership, many of whom already are stakeholders in the AOP community. There would be a variety of mutual benefits to establishing a governance and coordination body for AOP-oriented activities through a multipartite SETAC/SOT consortium of some type.To ensure sustainability and advancement of the AOP framework, adequate, ongoing financial support is a prerequisite for continuity and growth in the future. Currently, the AOP-KB (largely the AOP-Wiki) platform is supported by membership fees (from the SAAOP). While the OECD has funded the recent developments of Effectopedia and the e.AOP.Portal, the technical support and input for development of the AOP-Wiki are primarily contributed by volunteer experts. The current, very restricted, funding model will hinder the further growth of the AOP-KB, and even threatens its sustainability in its current form.Continuous funding would be required to expand the IT hardware , and human resources, such as full-time personnel, to upgrade software, enrich bioinformatics analyses, maintain the online system and coordinate/organize promotion and outreach activities with a view to supporting further growth of the AOP platform.only by industry or depending too much on the contributions of one or a few donors would be inappropriate and undermine independence. Consequently, it would be important that contributions supporting AOP development/system maintenance come from a variety of contributors from different sectors, and that none can exercise undue pressure on account of their relative financial weight.A critical role of a governance unit focused on AOPs would be development of a formal Business Plan, for example, a multi-year AOP Development Plan (ADP), presenting a sustainable financial budget and expense projections alongside with deliverables supporting the various stakeholder communities. Based on this ADP, a governance unit should be able to solicit resources from various potential funding agencies, charitable trusts and donors. Our vision is that this unit would be a non-profit NGO (consistent with the current SAAOP model), which in order to avoid potential conflicts of interest might impose a limitation on sources of funding. For instance, to have a governance body supported https://www.ncbi.nlm.nih.gov/genbank/, last accessed 20 Sep 2017), which is an open access, annotated database containing nucleotide sequences and their protein translations, could serve as a model for AOP governance/support. Initial funding from among others the National Institutes of Health, allowed for further collaboration with private enterprise. Early successes .Following the GenBank example, an AOP governance body could initially seek resources from multiple sources representing the whole range of stakeholders and geographies. Since the AOP platform is an international project, the funding may come from various countries. We recommend that a nascent AOP Consortium consider development of partnerships with the United Nations Environment Program (UNEP) under their program for the Strategic Approach to International Chemicals Management (SAICM). SAICM aims to promote chemical safety around the world, and provides funding to support relevant projects. While funding is primarily directed to developing countries, SETAC's long term history of collaboration with SAICM could provide a good basis for exploring the possibility of getting support for the AOP-KB in the context of the development of the “The Strategic Approach and sound management of chemicals and waste beyond 2020” (s) recently sponsored a call for proposals on the development of an AOP for cardiotoxicity under their strategic award funding scheme . In the US, one could envision that various units within the National Institute of Health could include similar disease or pathway-specific calls, or include contribution of information to the AOP Wiki as a requirement for funding, in the same way as the EU's Horizon 2020 programme has provided funding to the EU-ToxRisk consortium, (which includes a specific objective to develop AOPs for the KB).Funding agencies at country or regional levels also may be interested in supporting more targeted AOP endeavours. For example, the UK-based National Centre for the Replacement, Refinement and Reduction of Animals in Research  to develop a sustainable platform for the derivation of AOPs, ii) to engage and encourage stakeholders in developing and using AOPs, and iii) to embed AOPs and AOP thinking into decision-making we propose some initial “Impact Indicators” to measure success . Additio6Toxicological knowledge and regulation are currently at a turning point; historical approaches to regulatory toxicology are no longer scientifically, economically or socially robust enough to deal with the challenges of assessing chemicals that affect the environment, human and animal health. The AOP framework and associated knowledge has the potential to make a significant contribution to addressing these challenges, as well as parallel demands in biomedical fields. Realizing this potential is a two-pronged process: the first is to fully incorporate the AOP framework into toxicological research and related scientific fields; the second is to populate the AOP-KB with AOPs, making the constantly evolving knowledge on AOPs consistently organised and curated, accessible, shareable, and fit for purpose in regulatory and other decision-making contexts. In this review, we have identified stakeholders who both stand to benefit and to make significant contributions to the AOP approach, discussed the main challenges to full involvement with the approach, and made far-reaching suggestions for a proactive, cohesive and targeted approach to fully realizing the potential of the AOP approach.


Taenia solium diagnostics, several TPPs were initiated to address diagnostic needs for different stages in the parasite’s transmission . Following the meeting, draft TPPs were completed and distributed for consultation to 100 people/organizations, including experts in parasitology, human and pig cysticercosis, diagnostic researchers and manufacturers, international organizations working with neglected or zoonotic diseases, Ministries of Health and Ministries of Livestock in some of the endemic countries, WHO regional offices and other interested parties. There were 53 respondents. All comments and feedback received were considered and discussions were held with different experts according to their area of expertise. The comments were consolidated and final TPPs are presented here. They are considered to be live documents which are likely to undergo review and updating in the future when new knowledge and technologies become available.Target Product Profiles (TPPs) are process tools providing product requirements to guide researchers, developers and manufacturers in their efforts to develop effective and useful products such as biologicals, drugs or diagnostics. During a WHO Stakeholders Meeting on  Taenia solium  presents many challenges. The current paper describes the Optimal and Minimal attributes of the diagnostic tests that would be fit for a variety of diagnostic purposes, presented in the form of Target Product Profiles (TPPs). Four TPPs were drafted, distributed for consultation, and the feedback received was consolidated to produce the final versions presented here.Diagnosing the different manifestations of  Taenia solium. NCC is endemic in many low- and middle-income countries [Neurocysticercosis (NCC) is a Neglected Tropical Disease caused by infection with the cestode parasite ountries  and is tountries . In endeountries .T. solium involves two hosts. Humans harbour the sexually reproducing mature tapeworm in the small intestine (taeniasis). Eggs released with the faeces are infectious if eaten by pigs in which the larval parasite (cysticercus) encysts in the muscle tissues and brain, causing porcine cysticercosis. Ingestion of poorly cooked, infected pig meat by humans leads to growth of the tapeworm, completing the lifecycle. Eggs in the faeces of a patient infected with the adult tapeworm are also infectious if they are ingested by humans, in which case the cysticerci may lodge in muscle and other tissues, including the central nervous system causing NCC.The lifecycle of Symptoms of NCC vary depending, among other things, on the number, size and location of the cysts. They include headache, blindness, convulsions, epilepsy and death. The disease usually affects very poor communities, where pigs live in close contact with humans, and hygiene, sanitation and education are limited. The disease has a very strong social impact; epilepsy sufferers are often stigmatised, especially if sufferers are women and girls, as it is linked with witchcraft.Diagnosis of NCC is complicated, and neuroimaging is frequently required for a definite diagnosis , 5. CurrThere are several options for NCC disease control at population level, aimed at breaking the parasite’s transmission cycle. They include identification and treatment, or mass treatment for taeniasis (infection in humans with the adult tapeworm), chemotherapeutic treatment of porcine cysticercosis, pig vaccination, improvements in pig housing and management, meat inspection, improved human hygiene and sanitation, health education, etc. . EffortsT. solium is a need for better diagnostic tools to monitor the outcome of control efforts [Taenia solium, and generally for programs addressing neglected tropical diseases targeted for control and elimination [One of the challenges faced by efforts to control  efforts . Evaluat efforts . For Taemination , it is cT. solium infections, to coordinate and harmonize the search for appropriate diagnostic tests and to overcome the challenges in doing so. During the meeting, diagnostic priorities for different stages of the parasite’s transmission  were defined, and it was decided to initiate several Target Product Profiles (TPPs). An important step early in the process of designing diagnostic tools is defining their essential features, to allow researchers and diagnostic manufacturers to develop solutions that meet the needs in specific settings and for defined purposes. A TPP can be described as an important strategic document doing exactly this—describing key characteristics of a product. TPPs are used by the pharmaceutical and diagnostic industries, but also by organizations working on developing tools for the control of neglected diseases. Here we describe the development of four TTPs for T. solium taeniasis/cysticercosis.In December 2015 the World Health Organization (WHO) Department of Control of Neglected Tropical Diseases (WHO/NTD) and the Special Programme for Research and Training in Tropical Diseases (WHO/TDR) convened a stakeholder meeting  to discuT. solium Taeniasis/Cysticercosis Diagnostic Tools held at WHO headquarters, Geneva in December 2015, four TPPs were prioritised and progressed, and final drafts were elaborated. The attributes selected were based on previously published diagnostic TPPs produced by organizations such as FIND (www.finddx.org) and PATH (www.path.org), as well as the attributes suggested by the WHO [Following the Stakeholder Meeting on  the WHO . Two TPPT. solium: A Point-of-care test that could be used for surveillance, could be used at the initial stages of a control program, as well as for Diagnosis & Treatment.Taeniasis in humans caused by T. solium: A specific test (various platforms). Similar to number 1, but a test that could be used at later stages of a control program, when specificity and sensitivity are critical. This test could also be used for Diagnosis & Treatment.Taeniasis in humans caused by Human NCC: A Point-of-care test, to identify symptomatic patients with viable cysts that need to be referred for confirmation by brain imaging.Porcine cysticercosis (various platforms) test to be used to monitor control programs.TPPs for other use cases, such as screening of human populations, or detection of positive pigs through the pork chain, were not prioritised at this point in time.T. solium endemic regions [The draft TPPs were circulated to an extensive group of stakeholders for consultation. These included a wide range of experts in parasitology, human and pig cysticercosis, diagnostic experts, diagnostic manufacturers, organizations working with neglected diseases, organizations working with zoonotic diseases, some Ministries of Health and Directors of Veterinary services, WHO regional offices, as well as other interested parties. Efforts were made to ensure individuals and/or organizations from all  regions  were incAll comments and suggestions received were considered. Clarifications and follow up discussions were held with almost all contributors according to their area of expertise, to reach a consensus or arrive at a decision in relation to the various aspects of the final TPPs. In many cases, compromises needed to be made in order to keep the TPP attributes realistic and to account for the different opinions and needs as best as possible.A total of 53 replies were received: 13 from Africa, 13 from the Americas, 9 from Asia-Pacific, 11 from Europe, and 7 from International Organizations . Six of The final TPPs are presented in Tables Many constructive comments and suggestions were received and incorporated into the final versions. An alternate approach could have involved holding a consensus meeting of experts, however due to limited resources this was not deemed to be a viable option. The consultation via e-mail provided the opportunity for feedback to be extended to an extensive group of contributors who otherwise might not have been able to participate.The response rate was fifty-three per cent. This was considered to be more than satisfactory as the number and type of invitees was extensive for the size of the taeniasis/cysticercosis field globally. Constructive contributions came from a variety of individuals/organizations and included respondents from Africa, Asia-Pacific and the Americas . The sixT. solium taeniasis lack either sensitivity, specificity, or both (10). Current methods include faecal microscopy to identify parasite eggs, detection of coproantigens or copro-DNA, and detection of specific antibodies in serum. Faecal microscopy fails to identify pre-patent infections and is not species specific, antibody tests continue to test positive after treatment and currently available nucleic acid amplification tests (NAAT) are not suitable for application at point-of-care. Specificity of some tests might be high, such as the coproantigen test described by Guezala et al [Currently available methods for the diagnosis of la et al , howeverT. solium taeniasis. The rationale for having two tests was that they differ fundamentally in their intended use  and this connotes important differences in some attributes, such as the use case (attribute 1.2), location of use (attribute 1.5), as well as test specificity (attributes 3.1 and 3.4). A point-of-care antigen (Ag) test (TPP1) was recommended for use at the initial stages of an intervention when specificity for T. solium would not be critical. The second test, a more specific test (TPP2), was formulated for use in the later stages of an intervention in order to monitor the effectiveness of intervention procedures. If necessary, this second test could be performed in a specialist laboratory (does not need to be a point-of-care test). At the later stages of a control intervention the prevalence of taeniasis would be expected to be very low  is linked to the use case (attribute 1.2) and specificity (attribute 3.4) amongst others.The draft circulated TPPs were expecting all the tests to be species specific for The draft circulated for the Optimal attributes for a point-of-care test (TPP1) was based on a lateral flow assay. In response to the feedback received, the format (attribute 2.1) was changed to a Rapid Diagnostic Test, multiple formats allowed, in order to provide greater flexibility in relation to the technologies that could be adopted.A number of comments to the draft TPPs concerned the time period after a patient was successfully treated for taeniasis before the test should be expected to revert to being negative. A 6-day period was chosen as being realistic based on the performance of the current coproAg test .T. solium taeniasis, it was considered unusual to have children younger than 2 years of age infected. Ethical implications of testing young children, including the feasibility of treatment, need to be considered when using the test in the field.Several respondents questioned the age at which humans can be infected with taeniasis. A consensus was reached that, although young children have been occasionally found with Both the sensitivity and specificity for the taeniasis test TPPs (attributes 3.3 and 3.4) circulated as drafts were altered in the final versions based particularly on feedback that highlighted a need for very high test specificity for monitoring control programs. Specificity was prioritized for all taeniasis tests, except for the Minimal attributes point-of-care test, in which sensitivity was prioritized considering that the test would be used for surveillance, and not for monitoring an intervention.T. solium as much of the published information refers to taeniasis without defining the species. However, the starting prevalence of T. solium taeniasis is expected to be around 1–2%, and up to approx. 5% [While sensitivity and specificity are inherent test characteristics, they should be defined in the TPP taking into account the expected prevalence of the condition to be diagnosed in the conditions of use, which will determine the predictive value of a positive (PPV) and a negative (NPV) test. There is limited data about the prevalence of prox. 5% –19 althoprox. 5% . Within T. solium taeniasis [The reference test (attribute 3.6) selected for aeniasis  was chosThe relevant range of cost , was based on what was considered realistic based on the feedback provided, but no real cost analysis was done.Current tests for the diagnosis of NCC include antibody detection  and antigen detection (ELISA) tests. A challenge with the currently available tests is that a proportion of people living in endemic areas might test transiently positive on either antibody or antigen detection tests, apparently in the absence of mature or clinically-relevant cysticercosis –25. AntiComments were received and discussions held on the need to include also detection of antibodies, in order to detect patients with only non-viable/calcified cysts. Viable cysts degenerate over time, and some may eventually be calcified , 26. In The analytical sensitivity/limit of detection  was a frequent topic of discussion. A consensus emerged that the Optimal attribute should specify to detect patients with a single intracranial cysticercus, including both intraparenchymal and extra-parenchymal cysts. However, several opinions were received regarding the Minimal requirements. Eventually a compromise was reached to prioritise the detection of high-risk patients, defined as patients with 5 or more parenchymal cysticerci, or with a single ventricular or subarachnoid cysticercus.Respondents considered that it would be useful if a test could be used to monitor the effectiveness of anthelmintic treatment of NCC, particularly in specific scenarios such as patients with subarachnoid NCC . Hence tT. solium endemic areas. These responses could potentially be due to people being exposed to T. solium eggs, with the parasite possibly undergoing early development sufficient to induce a detectable serologic response, but not completing development and hence not causing detectable or symptomatic NCC. However, the precise cause of these transient responses has not been determined and it could be due to some cause unrelated to T. solium. This is particularly the case for the currently used test for circulating antigen which is known not to be specific for T. solium. In the case of the Minimal attributes for NCC, the existence of transiently positive reactions is included and, based upon the feedback that was received, a consensus was reached that a positive test would require to be repeated with a second sample collected after three months in order to confirm a case of persistent NCC. However, depending on the history and severity of the signs, the physician might decide to refer the patient immediately without repeat testing.An important and complex issue was how to deal with the existence of cases where transient serologically positive responses occur (both for antibody and antigen) in people living in Optimal sensitivity and specificity was reviewed, increasing the sensitivity and decreasing the specificity suggested in the draft, based on the need to detect the patients to be referred, but also accounting that the test would be used in a pre-selected population (symptomatic patients), so the test would still have a high positive predictive value.The definitive method for the diagnosis of porcine cysticercosis is detection of cysts with a full carcass muscle and brain dissection; this is both time consuming and expensive, particularly in the case of light infections. Serological tests (both for circulating antigen and antibody) are available but specificity can be problematic when used for monitoring a control program. It is important that tests do not cross react with other taeniid cestode parasites to which pigs are likely to be exposed. Additionally, many pigs in endemic areas that are serologically positive have no cysts at necropsy –31. It iT. solium, with some attributes modified to fit the use case. The occurrence of transient positive responses in animals that do not have mature cysts were not considered acceptable, unless it could be demonstrated unequivocally that these responses only occurred in animals exposed to T. solium and are not induced by any other cause such as exposure to any other parasites.The Minimal attributes were based on a currently available commercial antigen detection test for Discussions were held with a number of experts on the period over which the test would revert to being negative after a successful chemotherapy. The drug most commonly used to treat porcine cysticercosis is oxfendazole , 33. TheT. solium circulating antigens, however there are few data on serological responses in animals having only brain cysts. With these considerations in mind, in order to provide flexibility in the development of diagnostics for porcine cysticercosis the TPP’s Minimal indication for treated animals (attribute 1.1) was worded in a way such that it would not be restricted if the test were positive in animals having only viable brain cysts.Oxfendazole is highly effective for the treatment of cysticerci in muscle tissues, although efficacy is limited in the treatment of brain cysts . It is uA consensus was reached based on feedback from respondents that a qualitative test would be fit for purpose, hence this was changed from the proposal for a quantitative test that was included in the draft TPP which had been circulated. Analytical sensitivity was defined at 1 cyst for the Optimal attribute. A variety of views were expressed in this respect for the Minimal attribute, and a decision was taken to define the limit of detection for this test to also be 1 cyst.Sensitivity and specificity were also reviewed based on the feedback provided. Based on the use case, priority was determined to be specificity and hence this was maintained at a high level as per the original draft. Sensitivity was reviewed down, because of the challenges in achieving good sensitivity in an antigen test. A test with a relatively low sensitivity would nevertheless be useful in monitoring progress of control efforts, so long as the specificity was high.T. solium transmission and a consequent reduction in the incidence of human NCC.The TPPs were finalized based on the contributions of many subject matter experts serving different roles in academia, government, industry, NGOs and international organizations. Often a variety of views were expressed about a particular test attribute and, through additional discussions, in most cases a consensus was determined and this then formed the attribute included in the final TPPs. Each TPP is likely to need to be reviewed and updated in the future, as knowledge and technology progresses. It is expected that the TPPs presented here will assist researchers and diagnostic developers in guiding their efforts and ultimately development of the tools needed to assist in the control of