The evolving role of investigative toxicology in the pharmaceutical industry.

For decades, preclinical toxicology was essentially a descriptive discipline in which treatment-related effects were carefully reported and used as a basis to calculate safety margins for drug candidates. In recent years, however, technological advances have increasingly enabled researchers to gain insights into toxicity mechanisms, supporting greater understanding of species relevance and translatability to humans, prediction of safety events, mitigation of side effects and development of safety biomarkers. Consequently, investigative (or mechanistic) toxicology has been gaining momentum and is now a key capability in the pharmaceutical industry. Here, we provide an overview of the current status of the field using case studies and discuss the potential impact of ongoing technological developments, based on a survey of investigative toxicologists from 14 European-based medium-sized to large pharmaceutical companies.

Introducing the concept of virtual control groups into preclinical toxicology testing.

Sharing legacy data from in vivo toxicity studies offers the opportunity to analyze the variability of control groups stratified for strain, age, duration of study, vehicle and other experimental conditions. Historical animal control group data may lead to a repository, which could be used to construct virtual control groups (VCGs) for toxicity studies. VCGs are an established concept in clinical trials, but the idea of replacing living beings with virtual data sets has so far not been introduced into the design of regulatory animal studies. The use of VCGs has the potential of a 25% reduction in animal use by replacing the control group animals with existing randomized data sets. Prerequisites for such an approach are the availability of large and well-structured control data sets as well as thorough statistical evaluations. the foundation of data sharing has been laid within the Innovative Medicines Initiatives projects eTOX and eTRANSAFE. For a proof of principle participating companies have started to collect control group data for subacute (4-week) GLP studies with Wistar rats (the strain preferentially used in Europe) and are characterizing these data for its variability. In a second step, the control group data will be shared among the companies and cross-company variability will be investigated. In a third step, a set of studies will be analyzed to assess whether the use of VCG data would have influenced the outcome of the study compared to the real control group.

Biology-inspired microphysiological systems to advance patient benefit and animal welfare in drug development

The first microfluidic microphysiological systems (MPS) entered the academic scene more than 15 years ago and were considered an enabling technology to human (patho)biology in vitro and, therefore, provide alternative approaches to laboratory animals in pharmaceutical drug development and academic research. Nowadays, the field generates more than a thousand scientific publications per year. Despite the MPS hype in academia and by platform providers, which says this technology is about to reshape the entire in vitro culture landscape in basic and applied research, MPS approaches have neither been widely adopted by the pharmaceutical industry yet nor reached regulated drug authorization processes at all. Here, 46 leading experts from all stakeholders - academia, MPS supplier industry, pharmaceutical and consumer products industries, and leading regulatory agencies - worldwide have analyzed existing challenges and hurdles along the MPS-based assay life cycle in a second workshop of this kind in June 2019. They identified that the level of qualification of MPS-based assays for a given context of use and a communication gap between stakeholders are the major challenges for industrial adoption by end-users. Finally, a regulatory acceptance dilemma exists against that background. This t4 report elaborates on these findings in detail and summarizes solutions how to overcome the roadblocks. It provides recommendations and a roadmap towards regulatory accepted MPS-based models and assays for patients' benefit and further laboratory animal reduction in drug development. Finally, experts highlighted the potential of MPS-based human disease models to feedback into laboratory animal replacement in basic life science research.

Optimizing drug discovery by Investigative Toxicology: Current and future trends.

Investigative Toxicology describes the de-risking and mechanistic elucidation of toxicities, supporting early safety decisions in the pharmaceutical industry. Recently, Investigative Toxicology has contributed to a shift in pharmaceutical toxicology, from a descriptive to an evidence-based, mechanistic discipline. This was triggered by high costs and low throughput of Good Laboratory Practice in vivo studies, and increasing demands for adhering to the 3R (Replacement, Reduction and Refinement) principles of animal welfare. Outside the boundaries of regulatory toxicology, Investigative Toxicology has the flexibility to embrace new technologies, enhancing translational steps from in silico, in vitro to in vivo mechanistic understanding to eventually predict human response. One major goal of Investigative Toxicology is improving preclinical decisions, which coincides with the concept of animal-free safety testing. Currently, compounds under preclinical development are being discarded due to the use of inappropriate animal models. Progress in Investigative Toxicology could lead to humanized in vitro test systems and the development of medicines less reliant on animal tests. To advance this field a group of 14 European-based leaders from the pharmaceutical industry founded the Investigative Toxicology Leaders Forum (ITLF), an open, non-exclusive and pre-competitive group that shares knowledge and experience. The ITLF collaborated with the Centre for Alternatives to Animal Testing Europe (CAAT-Europe) to organize an "Investigative Toxicology Think-Tank", which aimed to enhance the interaction with experts from academia and regulatory bodies in the field. Summarizing the topics and discussion of the workshop, this article highlights Investigative Toxicology's position by identifying key challenges and perspectives.

Improving the Safety Assessment of Chemicals and Drug Candidates by the Integration of Bioinformatics and Chemoinformatics Data.

The application of read-across and in silico tools for regulatory decision-making has been limited for pharmaceutical compounds to the assessment of genotoxic impurity. In contrast, the broad availability of toxicity data for industrial chemicals has triggered regulatory frameworks for read-across (e.g. ECHA Read-Across Assessment Framework), software tools and public databases for an automated process of gap filling in the context of safety assessment. This MiniReview provides an overview of the currently existing in silico and read-across approaches for chemicals together with recent developments for pharmaceutical compounds in these areas. It also highlights the differences and commonalities in the in silico safety assessment of industrial chemicals and drug candidates. Whereas toxicity data collection and sharing is now common practice for chemicals falling under the European REACH regulation (Registration, Evaluation, Authorisation and Restriction of Chemicals), the biggest hurdle for establishing preclinical safety databases for pharmaceutical compounds is the unwillingness to share proprietary data and lack of published data sets. In a recent consortium approach, thirteen pharmaceutical companies, eleven academic partners and six small to medium size enterprises (SMEs) of the bioinformatics sector joined forces over the last 7 years within the European Innovative Medicines Initiative project eTOX ('electronic toxicity') to design and implement a strategy for leveraging these preclinical data for small molecules and sharing them across project partners. The eTOX database has evolved as the largest preclinical toxicity database for drugs and drug candidates and currently contains more than 1900 different chemical structures and more than 8000 in vivo toxicity study data sets. It can be foreseen that the development and application of such databases for drugs or drug candidates will in the future also cross-fertilize the read-across and the in silico assessment of industrial or consumer chemicals particularly as soon as human safety data from clinical trials are integrated too.

Legacy data sharing to improve drug safety assessment: the eTOX project.

The sharing of legacy preclinical safety data among pharmaceutical companies and its integration with other information sources offers unprecedented opportunities to improve the early assessment of drug safety. Here, we discuss the experience of the eTOX project, which was established through the Innovative Medicines Initiative to explore this possibility.

Nonclinical Safety and Toxicology.

Nonclinical safety pharmacology and toxicology testing of drug candidates assess the potential adverse effects caused by the drug in relation to its intended use in humans. Hazards related to a drug have to be identified and the potential risks at the intended exposure have to be evaluated in comparison to the potential benefit of the drug. Preclinical safety is thus an integral part of drug discovery and drug development. It still causes significant attrition during drug development.Therefore, there is a need for smart selection of drug candidates in drug discovery including screening of important safety endpoints. In the recent years,there was significant progress in computational and in vitro technology allowing in silico assessment as well as high-throughput screening of some endpoints at very early stages of discovery. Despite all this progress, in vivo evaluation of drug candidates is still an important part to safety testing. The chapter provides an overview on the most important areas of nonclinical safety screening during drug discovery of small molecules.

Inroads to predict in vivo toxicology-an introduction to the eTOX Project.

There is a widespread awareness that the wealth of preclinical toxicity data that the pharmaceutical industry has generated in recent decades is not exploited as efficiently as it could be. Enhanced data availability for compound comparison ("read-across"), or for data mining to build predictive tools, should lead to a more efficient drug development process and contribute to the reduction of animal use (3Rs principle). In order to achieve these goals, a consortium approach, grouping numbers of relevant partners, is required. The eTOX ("electronic toxicity") consortium represents such a project and is a public-private partnership within the framework of the European Innovative Medicines Initiative (IMI). The project aims at the development of in silico prediction systems for organ and in vivo toxicity. The backbone of the project will be a database consisting of preclinical toxicity data for drug compounds or candidates extracted from previously unpublished, legacy reports from thirteen European and European operation-based pharmaceutical companies. The database will be enhanced by incorporation of publically available, high quality toxicology data. Seven academic institutes and five small-to-medium size enterprises (SMEs) contribute with their expertise in data gathering, database curation, data mining, chemoinformatics and predictive systems development. The outcome of the project will be a predictive system contributing to early potential hazard identification and risk assessment during the drug development process. The concept and strategy of the eTOX project is described here, together with current achievements and future deliverables.

Incorporation of in silico biodegradability screening in early drug development--a feasible approach?

INTRODUCTION: The concentration of a pharmaceutical found in the environment is determined by the amount used by the patient, the excretion and metabolism pattern, and eventually by its persistence. Biological degradation or persistence of a pharmaceutical is experimentally tested rather late in the development of a pharmaceutical, often shortly before submission of the dossier to regulatory authorities. MATERIALS AND METHODS: To investigate whether the aspect of persistence of a compound could be assessed early during drug development, we investigated whether biodegradation of pharmaceuticals could be predicted with the help of in silico tools. To assess the value of in silico prediction, we collected results for the OECD 301 degradation test ("ready biodegradability") of 42 drugs or drug synthesis intermediates and compared them to the prediction of the in silico tool BIOWIN. RESULTS AND DISCUSSION: Of these compounds, 38 were predictable with BIOWIN, which is a module of the Estimation Programs Interface (EPI) Suite™ provided by the US EPA. The program failed to predict the two drugs which proved to be readily biodegradable in the degradation tests. On the other hand, BIOWIN predicted two compounds to be readily biodegradable which, however, proved to be persistent in the test setting. CONCLUSION: The comparison of experimental data with the predicted one resulted in a specificity of 94% and a sensitivity of 0%. The results of this study do not indicate that application of the biodegradation prediction tool BIOWIN is a feasible approach to assess the ready biodegradability during early drug development.

Evaluating the role of zinc in the occurrence of fibrosis of the skin: a preclinical study.

PURPOSE: To investigate the possible role of Zn as a trigger for NSF we were using a previously established preclinical model. The depletion of endogenous Zinc ions (Zn) caused by the administration of gadolinium-based contrast agents (GBCAs) has been suggested as a possible pathomechanism for nephrogenic systemic fibrosis (NSF). MATERIALS AND METHODS: In the Zn supplementation study, rats were injected with Gadodiamide, Omniscan, and Magnevist with or without Zn supplementation. In the Zn depletion study, animals were kept on a Zn-deficient diet or a special control diet and received injections of Omniscan, OptiMARK, Magnevist, Gadovist, and Gd-EDTA. Gd, Zn, and Cu concentrations in tissue were measured and histology of the skin was performed. RESULTS: As seen in earlier studies, a difference in Gd concentration in the skin was observed following treatment with the different GBCAs. High Gd concentration in the skin correlated with the occurrence of NSF-like skin lesions. We observed no differences in the occurrence of skin lesions between the Zn supplementation and the Zn-deficient groups compared to their respective control groups. CONCLUSION: We found no significant effect of Zn on the initiation of NSF-like skin lesions. The results further support data from previous studies highlighting the importance of complex stability of the investigated GBCAs.

Preclinical safety assessment of Vasovist (Gadofosveset trisodium), a new magnetic resonance imaging contrast agent for angiography.

OBJECTIVES: Vasovist (EPIX Pharmaceuticals and Schering AG) is a newly developed blood pool contrast agent for magnetic resonance imaging with a high affinity for human albumin, making it an ideal tool for the detection of structural abnormalities such as stenosis and aneurysm. For the risk assessment of the single diagnostic use in patients, the toxicity of this compound was investigated. MATERIALS AND METHODS: Studies of acute, repeated-dose, reproductive, and developmental toxicity as well as local tolerance, immunotoxicity, and mutagenic potential were performed. RESULTS: Lethality was observed in rodents after single intravenous administration at doses of at least 2 orders of magnitude higher than the anticipated human dose of 0.03 mmol/kg. The no observed adverse effect level after repeated daily administration over the course of 4 weeks to monkeys exceeded the single diagnostic dose by a factor of 3.3. The main effect of repeated dosing in both rats and monkeys was vacuolation in kidney proximal tubules without concomitant effect on kidney function. Studies into reproduction toxicity have shown no evidence of effects on fertility or perinatal and postnatal development. Signs of embryo-fetal toxicity were observed in rabbits after repeated administration of high doses. No indications of immunotoxic and mutagenic effects were observed. In local tolerance testing, Vasovist was well tolerated after intravenous administration. CONCLUSIONS: Vasovist was well tolerated with reasonable safety margins between the single diagnostic dose of 0.03 mmol/kg in humans and the doses resulting in adverse effects in animal studies.