The current paradigm used to determine the safety of an ingredient involves several in vitro, animal, and human studies. However, recent advances in in vitro methodologies have spurred the development of new approach methods, or NAMs. A NAM is defined by the US Environmental Protection Agency (EPA) as, “a technology, methodology, approach, or combination that can provide information on chemical hazard and risk assessment to avoid the use of animal testing”. NAMs are also commonly defined to include in silico, in chemico, and in vitro assays, as well as information about the exposure of chemicals in the context of hazard assessment. In silico methods are computational in nature, in chemico refers to purely chemical methods, such as testing a chemical by seeing if it reacts with another chemical or isolated protein in the absence of cells or tissues, and in vitro refers to experiments conducted in cell or tissue cultures. The ultimate goal of NAMs is to replace, reduce, or refine the use of animal testing in chemical safety assessment.
PROMOTING THE ADVANCEMENT OF NON-ANIMAL METHODS IN CHEMICAL SAFETY ASSESSMENT
To encourage the development, validation, and implementation of NAMs, the US has several dedicated groups working across multiple agencies. For example, the FDA has a working group dedicated to the advancement of alternative methods, and the US National Toxicology Program (NTP) has the interagency center for the evaluation of alternative toxicological methods, known as NICEATM. In other regions such as Canada and the EU, NAMs are being developed in response to bans on animal testing for new cosmetic ingredients. This ban has been in place in the EU since 2013, and in Canada since December, 2023. Additionally, the Organization for Economic Co-operation and Development (OECD) regularly publishes test guidelines, including in vitro methods for safety and toxicity testing.
ADVANCES IN IN VITRO METHODS FOR ASSESSING ENDOCRINE DISRUPTION
One area with several established in vitro methods is endocrine disruption. Briefly, an endocrine disruptor is a chemical that impacts the normal hormonal function of an organism. Classic examples include BPA found in plastics, phthalates found in products such as dryer sheets, and perfluoroalkyl and polyfluoroalkyl substances (PFAS) or “forever chemicals” found in things like non-stick cookware and many water repellant materials. Testing for endocrine disruption is also very important in the context of functional food, natural health product, and dietary supplement safety. For example, with the expansion of plant-based proteins and other soy-based ingredients, assessment of their potential for endocrine disruption is important because soy contains various phytoestrogens which may impact normal hormonal function, depending on the dose and the individual consuming them. Thus, several in vitro methods are currently in use to determine if a substance causes endocrine disruption. There are several such methods currently available from the OECD, and in general, these methods are used to determine if a chemical affects the production of sex hormones (testosterone and estrogen) or acts on the sex hormone receptors.
THE LEGACY OF HeLa CELLS
Recently, the cells used in endocrine disruption testing made the news. HeLa cells are a type of cervical cancer cell that have been used in biomedical research since 1951. Most notably, these cells were used early in the development of the polio vaccine, are used for in vitro fertilization (IVF), and to determine if chemicals interact with the estrogen receptor. The original donor of these cells, Henrietta Lacks, never gave consent for her cells to be used for research purposes after receiving surgery. The family of Henrietta Lacks recently reached an undisclosed settlement with ThermoFisher, with the settlement being reached on Henrietta’s 103rd birthday.
ADVANCES IN REPRODUCTIVE AND DEVELOPMENTAL TOXICITY ASSESSMENT
Another area of chemical safety testing with new in vitro methods is reproductive and developmental toxicity. In general, this refers to the ability of a chemical to adversely affect; (1) the ability of male or female organisms to reproduce, (2) the ability of a female to deliver a successful pregnancy to full term, or (3) the growth and development of the resultant offspring. These tests are typically performed with rodents or rabbits. However, a recently developed model is looking to provide a reliable in vitro alternative. This new test system uses human embryonic stem cells and measures biomarkers of normal cell metabolism to predict changes that might indicate toxicity in a developing embryo. This method, along with methods in zebrafish and C. elegans (a type of worm) are currently being evaluated by the National Institute of Environmental Health Sciences (NIEHS) to determine if these methods together can accurately predict the potential for food ingredients to induce reproductive or developmental toxicity.
CONCLUSION
NAMs are typically in vitro methods that aim to replace, reduce, or refine the use of animals in determining the safety of a chemical. There are many such methods currently being developed, with most being used to reduce or refine animal use. The development of NAMs is rapidly expanding, with new methods being introduced across several areas of chemical safety. Regulators around the world are actively working to encourage the development, validation, and implementation of these methods. With so many methods at various stages of development and with varying applications, it is important to understand which methods are accepted by regulators and which are relevant for a specific chemical or ingredient. The experts at SGS-Nutrasource can help you determine which test methods are right for your specific needs to ensure that the safety of your ingredient is properly characterized.
References:
-
EPA, 2023. EPA NAMs Training & Outreach Strategy | Science Inventory | US EPA
-
FDA (2022). Advancing Alternative Methods at FDA | FDA
-
National Toxicology Program (2023). https://ntp.niehs.nih.gov/go/niceatm
-
Health Canada (2023). Health Canada Announces the End of Cosmetic Animal Testing in Canada - Canada.ca
-
devTOX quickPredict Predicts Developmental Toxicity i | stemina.com
