Is read-across for chemicals comparable to medical device equivalence and where to use it for conformity assessment?

Novel medical devices must conform to medical device regulation (MDR) for European market entry. Likewise, chemicals must comply with the Registration, Evaluation, Authorization and Restriction of Chemicals (REACh) regulation. Both pose regulatory challenges for manufacturers, but concordantly provide an approach for transferring data from an already registered device or compound to the one undergoing accreditation. This is called equivalence for medical devices and read-across for chemicals. Although read-across is not explicitly prohibited in the process of medical device accreditation, it is usually not performed due to a lack of guidance and acceptance criteria from the authorities. Nonetheless, a scientifically justified read-across of material-based endpoints, as well as toxicological assessment of chemical aspects, such as extractables and leachables, can prevent failure of MDR device equivalence if data is lacking. Further, read-across, if applied correctly can facilitate the standard MDR conformity assessment. The need for read-across within medical device registration should let authorities to reconsider device accreditation and the formulation of respective guidance documents. Acceptance criteria like in the European Chemicals Agency (ECHA) read-across assessment framework (RAAF) are needed. This can reduce the impact of the MDR and help with keeping high European innovation device rate, beneficial for medical device patients.

New supporting data to guide the use of evident toxicity in acute oral toxicity studies (OECD TG 420).

Currently there are three test guidelines (TG) for acute oral toxicity studies of substances or mixtures from the Organisation for Economic Co-operation and Development (OECD). TG 423 and TG 425 use lethality as an endpoint, while TG 420 replaces death with 'evident toxicity', defined as clear signs that exposure to a higher dose would result in death. However, the perceived subjectivity of 'evident toxicity' may be preventing wider use of TG 420. To address this, the UK National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs) and the European Partnership for Alternative Approaches to Animal Testing (EPAA) collaborated to provide recommendations on the recognition of 'evident toxicity'. Historical data from acute oral toxicity studies were analysed for clinical signs at the lower dose that could have predicted death at the higher dose. Several signs including ataxia, laboured respiration, and eyes partially closed, alone or in combination, are highly predictive. Others such as lethargy, decreased respiration, and loose faeces have lower but still appreciable positive predictive value (PPV). The data has been used to develop recommendations to promote use of TG 420 and thus reduce the suffering and numbers of animals used in acute oral toxicity studies.

International regulatory uses of acute systemic toxicity data and integration of new approach methodologies.

Chemical regulatory authorities around the world require systemic toxicity data from acute exposures via the oral, dermal, and inhalation routes for human health risk assessment. To identify opportunities for regulatory uses of non-animal replacements for these tests, we reviewed acute systemic toxicity testing requirements for jurisdictions that participate in the International Cooperation on Alternative Test Methods (ICATM): Brazil, Canada, China, the European Union, Japan, South Korea, Taiwan, and the USA. The chemical sectors included in our review of each jurisdiction were cosmetics, consumer products, industrial chemicals, pharmaceuticals, medical devices, and pesticides. We found acute systemic toxicity data were most often required for hazard assessment, classification, and labeling, and to a lesser extent quantitative risk assessment. Where animal methods were required, animal reduction methods were typically recommended. For many jurisdictions and chemical sectors, non-animal alternatives are not accepted, but several jurisdictions provide guidance to support the use of test waivers to reduce animal use for specific applications. An understanding of international regulatory requirements for acute systemic toxicity testing will inform ICATM's strategy for the development, acceptance, and implementation of non-animal alternatives to assess the health hazards and risks associated with acute toxicity.

Exploring the potential of ToxCast™ data for mechanism-based prioritization of chemicals in regulatory context: Case study with priority existing chemicals (PECs) under K-REACH.

Recent studies have highlighted the potential of the ToxCast™ database for mechanism-based prioritization of chemicals. To explore the applicability of ToxCast data in the context of regulatory inventory chemicals, we screened 510 priority existing chemicals (PECs) regulated under the Act on the Registration and Evaluation, etc. of Chemical Substances (K-REACH) using ToxCast bioassays. In our analysis, a hit-call data matrix containing 298984 chemical-gene interactions was computed for 949 bioassays with the intended target genes, which enabled the identification of the putative toxicity mechanisms. Based on the reactivity to the chemicals, we analyzed 412 bioassays whose intended target gene families were cytochrome P450, oxidoreductase, transporter, nuclear receptor, steroid hormone, and DNA-binding. We also identified 141 chemicals based on their reactivity in the bioassays. These chemicals are mainly in consumer products including colorants, preservatives, air fresheners, and detergents. Our analysis revealed that in vitro bioactivities were involved in the relevant mechanisms inducing in vivo toxicity; however, this was not sufficient to predict more hazardous chemicals. Overall, the current results point to a potential and limitation in using ToxCast data for chemical prioritization in regulatory context in the absence of suitable in vivo data.

Unexpected neurotoxicity in chronic toxicity studies with a HBV viral expression inhibitor.

The non-clinical safety profile of the small molecule hepatitis B virus viral expression inhibitor RG7834 was studied in a package consisting of safety pharmacology, genotoxicity, repeat dose toxicity and reproductive toxicity studies. The chronic monkey toxicity study identified dose- and time-dependent symptoms of polyneuropathy, with correlating nerve conduction velocity reductions and axonal degeneration in peripheral nerves and spinal cord, in all compound treatment groups with no evidence of reversibility after approximately 3 months of treatment cessation. Similar histopathological findings were observed in the chronic rat toxicity study. Subsequent in vitro neurotoxicity investigations and ion channel electrophysiology did not elucidate a potential mechanism for the late toxicity. However, based on similar findings observed with a structurally different molecule, an inhibition of their common pharmacological targets, PAPD5 & PAPD 7, was considered as a possible mechanism of toxicity. In conclusion, the marked neuropathies, only observed after chronic dosing, did not support further clinical development of RG7834 because of its foreseen clinical treatment duration of up to 48 weeks in chronic HBV patients.

Gaps and challenges in nonclinical assessments of pharmaceuticals: An FDA/CDER perspective on considerations for development of new approach methodologies.

Previously, we provided an FDA/CDER perspective on nonclinical testing strategies and briefly discussed the opportunities and challenges of using new approach methodologies (NAMs) in drug development, especially for regulatory purposes. To facilitate the integration of NAMs into nonclinical regulatory testing, we surveyed the CDER Pharmacology/Toxicology community to identify the nonclinical challenges faced by CDER review staff, including gaps and areas of concern underserved by current nonclinical testing approaches, and to understand how development of NAMs with specific contexts of use (COUs) could potentially alleviate them. Survey outcomes were coalesced into CDER-identified needs for which NAMs with specific COUs could potentially be developed to address gaps and challenges in nonclinical safety assessments. We also discussed the current FDA procedure for validation and qualification of NAMs intended to inform regulatory decisions. This manuscript is intended to facilitate productive discussions and collaborations with regulatory, government, and academic stakeholders within the drug development community regarding the development and regulatory use of NAMs and their role in safety and efficacy assessment of pharmaceuticals.

Towards a future regulatory framework for chemicals in the European Union - Chemicals 2.0.

The body of EU chemicals legislation has evolved since the 1960s, producing the largest knowledge base on chemicals worldwide. Like any evolving system, however, it has become increasingly diverse and complex, resulting in inefficiencies and potential inconsistencies. In the light of the EU Chemicals Strategy for Sustainability, it is therefore timely and reasonable to consider how aspects of the system could be simplified and streamlined, without losing the hard-earned benefits to human health and the environment. In this commentary, we propose a conceptual framework that could be the basis of Chemicals 2.0 - a future safety assessment and management approach that is based on the application of New Approach Methodologies (NAMs), mechanistic reasoning and cost-benefit considerations. Chemicals 2.0 is designed to be a more efficient and more effective approach for assessing chemicals, and to comply with the EU goal to completely replace animal testing, in line with Directive 2010/63/EU. We propose five design criteria for Chemicals 2.0 to define what the future system should achieve. The approach is centered on a classification matrix in which NAMs for toxicodynamics and toxicokinetics are used to classify chemicals according to their level of concern. An important principle is the need to ensure an equivalent, or higher, protection level.

Alternatives to Monkey Reproductive Toxicology Testing for Biotherapeutics.

Embryofetal toxicity studies are conducted to support inclusion of women of childbearing potential in clinical trials and to support labeling for the marketed pharmaceutical product. For biopharmaceuticals, which frequently lack activity in the rodent or rabbit, the nonhuman primate is the standard model to evaluate embryofetal toxicity. These studies have become increasingly challenging to conduct due to the small number of facilities capable of performing them and a shortage of sexually mature monkeys. The low number of animals per group and the high rate of spontaneous abortion in cynomolgus monkeys further complicate interpretation of the data. Recent FDA guidance has proposed a weight of evidence (WoE) approach to support product labeling for reproductive toxicity of products intended to be used for the treatment of cancer (Oncology Pharmaceuticals: Reproductive Toxicity Testing and Labeling Recommendations), an approach that has also supported the approval of biotherapeutics for non-cancer indications. Considerations to determine the appropriateness and content of a WoE approach to support product labeling for embryofetal risk include known class effects in humans; findings from genetically modified animals with or without drug administration; information from surrogate compounds; literature-based assessments about the developmental role of the pharmaceutical target; and the anticipated exposure during embryofetal development. This paper summarizes the content of a session presented at the 42nd annual meeting at the American College of Toxicology, which explored the conditions under which alternative approaches may be appropriate to support product labeling for reproductive risk, and how sponsors can best justify the use of this approach.

Scientific and Regulatory Policy Committee Best Practices: Recommended ("Best") Practices for Informed (Non-blinded) Versus Masked (Blinded) Microscopic Evaluation in Animal Toxicity Studies.

This article describes the Society of Toxicologic Pathology's (STP) five recommended ("best") practices for appropriate use of informed (non-blinded) versus masked (blinded) microscopic evaluation in animal toxicity studies intended for regulatory review. (1) Informed microscopic evaluation is the default approach for animal toxicity studies. (2) Masked microscopic evaluation has merit for confirming preliminary diagnoses for target organs and/or defining thresholds ("no observed adverse effect level" and similar values) identified during an initial informed evaluation, addressing focused hypotheses, or satisfying guidance or requests from regulatory agencies. (3) If used as the approach for an animal toxicity study to investigate a specific research question, masking of the initial microscopic evaluation should be limited to withholding only information about the group (control or test article-treated) and dose equivalents. (4) The decision regarding whether or not to perform a masked microscopic evaluation is best made by a toxicologic pathologist with relevant experience. (5) Pathology peer review, performed to verify the microscopic diagnoses and interpretations by the study pathologist, should use an informed evaluation approach. The STP maintains that implementing these five best practices has and will continue to consistently deliver robust microscopic data with high sensitivity for animal toxicity studies intended for regulatory review. Consequently, when conducting animal toxicity studies, the advantages of informed microscopic evaluation for maximizing sensitivity outweigh the perceived advantages of minimizing bias through masked microscopic examination.

The use of Bayesian methodology in the development and validation of a tiered assessment approach towards prediction of rat acute oral toxicity.

There exists consensus that the traditional means by which safety of chemicals is assessed-namely through reliance upon apical outcomes obtained following in vivo testing-is increasingly unfit for purpose. Whilst efforts in development of suitable alternatives continue, few have achieved levels of robustness required for regulatory acceptance. An array of "new approach methodologies" (NAM) for determining toxic effect, spanning in vitro and in silico spheres, have by now emerged. It has been suggested, intuitively, that combining data obtained from across these sources might serve to enhance overall confidence in derived judgment. This concept may be formalised in the "tiered assessment" approach, whereby evidence gathered through a sequential NAM testing strategy is exploited so to infer the properties of a compound of interest. Our intention has been to provide an illustration of how such a scheme might be developed and applied within a practical setting-adopting for this purpose the endpoint of rat acute oral lethality. Bayesian statistical inference is drawn upon to enable quantification of degree of confidence that a substance might ultimately belong to one of five LD50-associated toxicity categories. Informing this is evidence acquired both from existing in silico and in vitro resources, alongside a purposely-constructed random forest model and structural alert set. Results indicate that the combination of in silico methodologies provides moderately conservative estimations of hazard, conducive for application in safety assessment, and for which levels of certainty are defined. Accordingly, scope for potential extension of approach to further toxicological endpoints is demonstrated.

Integrating toxicokinetics into toxicology studies and the human health risk assessment process for chemicals: Reduced uncertainty, better health protection.

The database of practical examples where toxicokinetic (TK) data has benefitted all stages of the human health risk assessment process are increasingly being published and accepted. This review aimed to highlight and summarise notable examples and to describe the "state of the art" in this field. The overall recommendation is that for any in vivo animal study conducted, measurements of TK should be very carefully considered for inclusion as the numerous benefits this brings continues to grow, particularly during the current march towards animal free toxicology testing and ambitions to eventually conduct human health risk assessments entirely based upon non-animal methods.

Utilizing Whole Slide Images for the Primary Evaluation and Peer Review of a GLP-Compliant Rodent Toxicology Study.

The approach undertaken to deliver a Good Laboratory Practice (GLP) validation of whole slide images (WSIs) and the associated workflow for the digital primary evaluation and peer review of a GLP-compliant rodent inhalation toxicity study is described. The contract research organization (CRO) undertook validation of the slide scanner, scanner software, and associated database software. This provided a GLP validated environment within the database software for the primary histopathologic evaluation using WSI and viewed with the database software web viewer. The CRO also validated a cloud-based digital pathology platform that supported the upload and transfer of WSI and metadata to a cache within the sponsor's local area network. The sponsor undertook a separate GLP validation of the same cloud-based digital pathology platform to cover the download and review of the WSI. The establishment of a fit-for-purpose GLP-compliant workflow for WSI and successful deployment for the digital primary evaluation and peer review of a large GLP toxicology study enabled flexibility in accelerated global working and potential future reuse of digitized data for advanced artificial intelligence and machine learning image analysis.

Serial blood sampling effects in rat embryo-fetal development studies for toxicokinetics.

Serial blood sampling for toxicokinetics is generally conducted in regulatory embryo-fetal development (EFD) studies in rats. EFD studies are designed to detect the potential adverse effects of pharmaceuticals on pregnant females and their fetuses; this information is useful for understanding the relationships between systemic exposure levels and toxicity profiles. However, additional satellite pregnant females are needed for toxicokinetics because comprehensive information regarding the potential impact of serial blood sampling on pregnant females is scarce. Here, in this study, we investigated the potential impact of serial blood sampling in pregnant female rats using a typical EFD study design. Additionally, we investigated the additional endpoints (clinical pathology, organ weights, and histopathology) that were deemed likely to be sensitive to blood sampling. Results indicated that serial blood sampling in pregnant females induced physiological adaptive changes and did not affect the general endpoints in EFD studies. Nevertheless, inclusion of satellite groups in EFD studies may be a more prudent approach considering the physiological changes in pregnant females and potential off-target effects of candidate pharmaceuticals. These results provide background information on the impact of serial blood sampling in pregnant females and will be useful to design the regulatory EFD studies.

Comprehensive analysis of chronic rodent inhalation toxicity studies for methyl acrylate with attention to test conditions exceeding a maximum tolerated concentration.

MA is a chemical intermediate, manufactured and processed within closed systems. While so far available subacute to chronic inhalation toxicity studies performed in compliance with OECD TG principles gave no indication of any carcinogenic potential for MA, a recent 2-year inhalation study with F344/DuCrlCrlj rats published in 2017 by the JBRC showed a statistically significant increase of squamous cell carcinoma in the nasal cavity of male rats at the highest tested concentration of 160 ppm. However, the results of the different studies in total indicate that this high concentration exceeded the MTC. As MA has a low potential for genotoxic and mutagenic activity, the increased tumour incidence can be attributed to a non-genotoxic mechanism, namely to a strong inflammatory response observed in this study. Together with mechanistic and epidemiologic data for other compounds related to nasal carcinogenesis via this mode of action, it can be concluded that the relevance of this increased tumour incidence in male rats for humans is questionable. Also, a long-term exposure to higher concentrations of MA is highly unlikely to be reached in the environment or at workplaces. Therefore, a risk for humans including cancer hazard is considered implausible.

Dying for change: A roadmap to refine the fish acute toxicity test after 40 years of applying a lethal endpoint.

The fish acute toxicity test (TG203; OECD, 2019) is frequently used and highly embedded in hazard and risk assessment globally. The test estimates the concentration of a chemical that kills 50% of the fish (LC50) over a 96 h exposure and is considered one of the most severe scientific procedures undertaken. Over the years, discussions at the Organisation for Economic Co-operation and Development (OECD) have resulted in changes to the test which reduce the number of fish used, as well as the development of a (potential) replacement test (TG236, OECD, 2013). However, refinement of the mortality endpoint with an earlier (moribundity) endpoint was not considered feasible during the Test Guideline's (TG) last update in 2019. Several stakeholders met at a UK-based workshop to discuss how TG203 can be refined, and identified two key opportunities to reduce fish suffering: (1) application of clinical signs that predict mortality and (2) shortening the test duration. However, several aspects need to be addressed before these refinements can be adopted. TG203 has required recording of major categories of sublethal clinical signs since its conception, with the option to record more detailed signs introduced in the 2019 update. However, in the absence of guidance, differences in identification, recording and reporting of clinical signs between technicians and laboratories is likely to have generated piecemeal data of varying quality. Harmonisation of reporting templates, and training in clinical sign recognition and recording are needed to standardise clinical sign data. This is critical to enable robust data-driven detection of clinical signs that predict mortality. Discussions suggested that the 96 h duration of TG203 cannot stand up to scientific scrutiny. Feedback and data from UK contract research organisations (CROs) conducting the test were that a substantial proportion of mortalities occur in the first 24 h. Refinement of TG203 by shortening the test duration would reduce suffering (and test failure rate) but requires a mechanism to correct new results to previous 96 h LC50 data. The actions needed to implement both refinement opportunities are summarised here within a roadmap. A shift in regulatory assessment, where the 96 h LC50 is a familiar base for decisions, will also be critical.

Scientific and Regulatory Policy Committee Points to Consider*: Approaches to the Conduct and Interpretation of Vaccine Safety Studies for Clinical and Anatomic Pathologists.

The design and execution of toxicology studies supporting vaccine development have some unique considerations relative to those supporting traditional small molecules and biologics. A working group of the Society of Toxicologic Pathology Scientific and Regulatory Policy Committee conducted a review of the scientific, technical, and regulatory considerations for veterinary pathologists and toxicologists related to the design and evaluation of regulatory toxicology studies supporting vaccine clinical trials. Much of the information in this document focuses on the development of prophylactic vaccines for infectious agents. Many of these considerations also apply to therapeutic vaccine development (such as vaccines directed against cancer epitopes); important differences will be identified in various sections as appropriate. The topics addressed in this Points to Consider article include regulatory guidelines for nonclinical vaccine studies, study design (including species selection), technical considerations in dosing and injection site collection, study end point evaluation, and data interpretation. The intent of this publication is to share learnings related to nonclinical studies to support vaccine development to help others as they move into this therapeutic area. [Box: see text].

Alternatives to statistical decision trees in regulatory (eco-)toxicological bioassays.

The goal of (eco-) toxicological testing is to experimentally establish a dose or concentration-response and to identify a threshold with a biologically relevant and probably non-random deviation from "normal". Statistical tests aid this process. Most statistical tests have distributional assumptions that need to be satisfied for reliable performance. Therefore, most statistical analyses used in (eco-)toxicological bioassays use subsequent pre- or assumption-tests to identify the most appropriate main test, so-called statistical decision trees. There are however several deficiencies with the approach, based on study design, type of tests used and subsequent statistical testing in general. When multiple comparisons are used to identify a non-random change against negative control, we propose to use robust testing, which can be generically applied without the need of decision trees. Visualization techniques and reference ranges also offer advantages over the current pre-testing approaches. We aim to promulgate the concepts in the (eco-) toxicological community and initiate a discussion for regulatory acceptance.

The value of avian gross pathology in identifying endocrine disrupting properties.

The European Chemical Agency and European Food Safety Authority recommend that gross pathology findings, from avian reproduction studies, be used to support assessment of potential endocrine disrupting properties of active pesticidal and biocidal substances. In open literature, little information is available on the utility of gross pathology data for informing endocrine evaluations. Here the gross pathology data from historical control groups of 51 northern bobwhite and 51 mallard reproduction tests is analyzed to evaluate the utility of such information. Incidence of gross morphology findings in untreated birds may aid the interpretation of some gross abnormalities, potentially indicative of an endocrine interaction (e.g. reproductive condition). Statistical analysis of the historical control data indicates that gross pathology is not likely to be useful for detecting endocrine effects as abnormalities with relatively high increases in prevalence (more than 20-30%, depending on prevalence in controls) are reliably interpreted as a treatment response. Gross pathology changes are only indicative and not diagnostic of endocrine interactions making it difficult to distinguish gross pathology abnormalities, due to endocrine-mediated effects, from systemic toxicity. This work demonstrates the utility of using historical control analyses to establish the value and properties of selected endpoints for regulatory applications.

Advances in selecting appropriate non-rodent species for regulatory toxicology research: Policy, ethical, and experimental considerations.

In vivo animal studies are required by regulatory agencies to investigate drug safety before clinical trials. In this review, we summarize the process of selecting a relevant non-rodent species for preclinical studies. The dog is the primary, default non-rodent used in toxicology studies with multiple scientific advantages, including adequate background data and availability. Rabbit has many regulatory advantages as the first non-rodent for the evaluation of reproductive and developmental as well as local toxicity. Recently, minipigs have increasingly replaced dogs and rabbits in toxicology studies due to ethical and scientific advantages including similarity to humans and breeding habits. When these species are not relevant, nonhuman primates (NHPs) can be used as the available animal models, especially in toxicology studies investigating biotherapeutics. Particularly, based on the phylogenetic relationships, the use of New-World marmosets can be considered before Old-World monkeys, especially cynomolgus with robust historical data. Importantly, the use of NHPs should be justified in terms of scientific benefits considering target affinity, expression pattern, and pharmacological cross-reactivity. Strict standards are required for the use of animals. Therefore, this review is helpful for the selection of appropriate non-rodent in regulatory toxicology studies by providing sufficient regulatory, ethical, and scientific data for each species.

The Implementation of the Three Rs in Regulatory Toxicity and Biosafety Assessment: The Indian Perspective.

Animal models have long served as a basis for scientific experimentation, biomedical research, drug development and testing, disease modelling and toxicity studies, as they are widely thought to provide meaningful, human-relevant predictions. However, many of these systems are resource intensive and time-consuming, have low predictive value and are associated with great social and ethical dilemmas. Often drugs appear to be effective and safe in these classical animal models, but later prove to be ineffective and/or unsafe in clinical trials. These issues have paved the way for a paradigm shift from the use of in vivo approaches, toward the 'science of alternatives'. This has fuelled several research and regulatory initiatives, including the ban on the testing of cosmetics on animals. The new paradigm has been shifted toward increasing the relevance of the models for human predictivity and translational efficacy, and this has resulted in the recent development of many new methodologies, from 3-D bio-organoids to bioengineered 'human-on-a-chip' models. These improvements have the potential to significantly advance medical research globally. This paper offers a stance on the existing strategies and practices that utilise alternatives to animals, and outlines progress on the incorporation of these models into basic and applied research and education, specifically in India. It also seeks to provide a strategic roadmap to streamline the future directions for the country's policy changes and investments. This strategic roadmap could be a useful resource to guide research institutions, industries, regulatory agencies, contract research organisations and other stakeholders in transitioning toward modern approaches to safety and risk assessment that could replace or reduce the use of animals without compromising the safety of humans or the environment.