Advances in science are transforming how scientists study health and disease. Biomedical and behavioral research can involve working with animal models, cells in test tubes, computer modeling, and clinical studies with people. Each research approach is critical to advancing our knowledge of health and disease.

Over the last decade, improved technological capabilities enable scientists to reduce their reliance on animal models for specific types of studies. These advances are important because testing in animals can pose ethical issues, takes significant time and resources, and the relevance to human health is not always certain.

Recent advances in alternative methods, such as computational, biochemical, or cell-based model systems that can replicate human biology, have been shown in some cases to perform the same as or even better than standard animal models. These “New Approach Methodologies” (NAMs, sometimes called Novel Alternative Methods, Non-Animal Methods, or New Alternative Methods) can provide a complementary approach to traditional models. They can offer a way to improve understanding of the human system and its susceptibility to toxic effects, and to discover effective treatments for human conditions.

NIEHS has been a leader in the development of NAMs to reduce the use of animals and increase the human relevance of models in health effects testing. NIEHS is working with the scientific community and other stakeholders to increasingly apply these methods to policy and regulatory decision-making processes or to replace traditional testing requirements.

The 3Rs

NIEHS is also committed to efforts that replace, reduce, or refine (the “3Rs”) the use of animal models in studies. This concept of replacing, reducing, or refining animal use in research and testing was first described more than 65 years ago. The 3Rs became a tenet for the scientific community to address animal welfare in a meaningful way.

• Replacing: The method substitutes traditional animal models with non-animal systems such as computer models or biochemical or cell-based systems, or one animal species is replaced with a less developed one (for example, replacing a mouse with a worm).
• Reducing: The method decreases the number of animals required for testing to a minimum while still achieving testing objectives.
• Refining: The method eliminates pain or distress in animals, or enhances animal well-being, such as by providing better housing or enrichment.

Types of Alternative Methods

The scientific toolbox used for toxicological assessment continually expands. In a 2023 report, NIH categorized NAMs approaches into three general categories. Definitions and examples of each method are below.

In chemico: Experiments performed on biological molecules, such as proteins and DNA, outside of cells, which may be used to study how these molecules interact with each other and with drugs.

• Non-mammalian models, such as zebrafish, nematodes, and planaria, can enable the rapid and cost-effective assessment of the effects of chemicals on biology and behavior.

In silico: Experiments performed by computing platforms or custom hardware, encompassing mathematical modeling and simulation, machine learning, and other computational techniques.

• Computational model is a general term describing the use of computers to simulate complex systems, such as those that can be used as a NAM.
• Artificial intelligence and machine learning (AI/ML) approaches can enhance and support human intellect in assessing the safety of chemical exposures. For example, advanced computer simulations can be used to model biological processes and predict the effects of previously untested chemicals and drugs.

In vitro: Experiments performed on cells outside of the body, including various types of cell, organoid, and tissue culture techniques.

• These methods may involve culturing cells in the laboratory and can take advantage of high-throughput screening of large numbers of chemical compounds.
• Microphysiological systems or organs-on-chips are complex, cell-based devices that mimic key physiological aspects of tissues or organs by incorporating microenvironments that align with those in the human body. A particular system might incorporate flow and shear stress, appropriate pH and oxygen levels, biochemical and electrical stimuli, and other factors that replicate various features of animal-based models.
• Mini-organs or organoids are 3D tissue-like structures, derived from stem cells, that closely replicate the complexity and function of human organs. These tiny, artificially grown tissues contain various specialized cell types similar to those found in full-sized organs.

Different human populations experience different levels of susceptibility to toxic effects from chemical exposure, presenting a complex problem for chemical risk assessment. Alternative methods, such as those above, could provide more rapid and human-relevant techniques to assess chemical safety for communities affected by environmental exposures. Alternatives to animal testing can be used for toxicological research, such as chemical safety and drug effectiveness testing, or other types of biomedical research that examine biological mechanisms of disease.

NIH Leadership for NAMs

The NIH Common Fund’s Complement Animal Research In Experimentation program (Complement-ARIE) aims to speed the development, standardization, validation, and use of human-based NAMs. It will significantly advance the understanding of human health and disease and approaches to basic, translational, toxicological, and clinical research. NIEHS plays a lead role in planning activities for this program.

Complement-ARIE brings together many areas of expertise through a consortium of researchers in the following efforts:

• Technology centers will develop NAMs projects to fill in areas of greatest need. Projects will emphasize biological complexity, high throughput techniques, combining approaches, and data sharing.
• A resource coordinating center will create integrated data structures and a searchable NAMs repository.
• A validation and qualification network will accelerate deployment and regulatory approval of NAMs for biomedical research.
• Community engagement and training will promote the development of an inclusive and diverse biomedical research workforce with the skills to build and use new NAMs.
• Strategic engagement with key partners will advance opportunities in the development and use of NAMs in basic, translational, and clinical research.