New approach methodologies (NAMs) are nonclinical tools that offer potential to improve biological relevance and predictive accuracy of safety and efficacy assessments, while reducing animal use and shortening drug development timelines. Methods and approaches, such as NAMs, that support the principles of the 3Rs (replacement, reduction, refinement) in research and drug development have long had widespread support from the scientific community, ICH guidance, and animal welfare groups, but recent announcements from regulatory authorities have spurred new momentum in using NAMs to support regulatory decisions. In April 2025, FDA published a roadmap to reduce animal testing in preclinical safety studies1. More recently, EMA published a reflection paper aiming to reduce the use of non-human primates (NHPs)2, and the UK government announced a new initiative to reduce the use of dogs and NHPs by at least 35% by 2030 that will be backed by £75 million in funding to support development of new testing methods3.
Though regulatory expectations are evolving, as evidenced by new guidance from FDA indicating that toxicology studies longer than 3 months in duration are no longer warranted for monospecific antibodies4, animal studies will continue to play a role in establishing exposure-response relationships, identifying potential adverse events, and determining safe doses. Sponsors who conduct animal studies routinely contend with challenges, including animal models that do not always accurately predict human toxicities, due to species differences in metabolism, antigen expression, or other physiological differences; moreover, these studies are costly and time-consuming. Additional constraints, such as limited availability of NHPs, can further complicate the ability to conduct these studies.
NAMs includes a wide array of in vitro, in silico, and in chemico methods and are commonly employed during drug discovery to advance the best candidate molecule. However, new efforts from many stakeholders seek to explore additional opportunities where NAMs can be applied to modernize nonclinical pharmacology and toxicology studies and address some of these long-standing challenges. Importantly, NAMs could be used in combination with animal studies or other NAMs to form integrated frameworks that improve human relevance. A few examples that may be considered are listed below.
- Microphysiologic systems (MPS) are in vitro systems that use human cells and tissues to model complex interactions and include 3D tissue culture methods (such as organoids and tissue spheroids) that may better reflect in vivo physiologic and mechanical properties compared to traditional 2D cell culture methods. For example, tumor spheroids derived from patient biopsies are used to model the tumor microenvironment and assess the anti-tumor activity of novel therapeutics.
- Microfluidic systems include organ-on-a-chip assays and have been developed for most human organs. Multiple different organ-on-a-chip assays can be linked to replicate interactions between organ systems.
- Omics-based methods, such as genomic, transcriptomic, proteomic, and metabolomic approaches, can be used for high throughput screening. These methods often generate large data sets that allow for multiple analyses.
- Computational methods, like quantitative-structure activity relationship (QSAR), are also available to predict and model biological properties and activity of new molecules.
- Large language models (LLMs) and artificial intelligence (AI) can aid in evaluating large volumes of existing data generated for successful and failed drug candidates to determine what types of data and assays are informative. While historical data can provide a reference that can be queried for comparative data, data will likely need to be transformed to a standardized format to allow for analysis.
NAMs may not replace pivotal animal studies entirely, but they can be used to de-risk or refine study design to increase the likelihood of successful animal studies when they are needed. Assays that improve dose selection or identify key biological parameters may justify streamlined toxicology study designs that use fewer dose levels or phylogenetically lower species, thereby reducing unnecessary animal use. NAMs could also be used to provide additional supplemental scientific evidence to support the translatability (or lack thereof) of findings from animal studies. Similar approaches are currently used in drug development as evidenced by the adoption of in vitro assays to support safety assessments, such as in vitro hERG assays to evaluate potential for cardiac toxicity. Tiered approaches, where in vitro assays are conducted to determine whether additional in vivo studies are warranted, such as those applied to phototoxicity or dermal toxicity assessments, also support the principles of the 3Rs and reduce animal use.
Sponsors seeking to leverage NAMs in their nonclinical development strategy should discuss their intended use with the appropriate regulatory authorities. A recent publication provides an FDA/CDER pharmacology/toxicology perspective on NAM-based approaches that have potential for use in regulatory decision making5. Examples of NAMs submitted by Sponsors are presented along with regulatory outcomes. While the findings in this paper are encouraging, challenges with using NAMs to replace in vivo testing were highlighted, such as assessing off-target effects in systems that do not fully replicate the complex physiology of a whole organism. Further, this paper emphasizes the essential need to define the intended context-of-use (COU) in order to determine whether an assay is suitable or “fit-for-purpose”. Another hurdle for regulatory acceptance is the need for qualification, standardization, and validation to ensure reliability in regulatory decision making. When engaging with regulators, a clear narrative describing the NAM and its scope and limitations, as well as any efforts to validate the assay, is key for advancing its use. Asking specific and focused questions in meetings and other interactions with regulators can also help guide conversations to identify and address central issues. As more case studies and robust data sets are shared, regulators will be able to better define expectations and formulate clear guidance for Sponsors on the use of NAMs.
Antibody-based therapeutics are especially well-suited for NAM approaches due to their highly specific targets and the extensive clinical experience with large biologic molecules. Data collection and conversations are ongoing to re-evaluate the relevance of chronic toxicology studies using NHPs for some products, like CD3-bispecific antibodies and antibody-drug conjugates (ADCs) that use a cytotoxic payload, where long-term toxicology studies in NHPs may not be informative. For example, peripheral neuropathy and ocular toxicity are common adverse effects for some ADCs, but these findings are not always detected in NHPs. In vitro assays using iPSC-derived neurons or corneal cells, respectively, are currently being evaluated to better understand the mechanisms for these toxicities and predict when they may occur clinically. These types of studies could supplement animal studies to provide a more comprehensive assessment of potential toxicity. Platform-based approaches are increasingly common for many types of products, including ADCs, and could be used to inform on safety and reduce the need for additional studies by using data generated across development programs. In some cases, a weight-of-evidence (WoE)-based approach that includes all available nonclinical and clinical data and relevant published literature may be used in lieu of some toxicology studies; as described in ICH S1B(R1)6, a WoE approach can be used instead of a 2-year rat carcinogenicity study for certain products.
As mechanism-driven, human-based tools, NAMs offer the potential for translational insights that traditional in vivo testing alone may miss, while also reducing development time and cost, and minimizing the need for animal testing. Advancing regulatory acceptance will require greater collaboration and data sharing among all stakeholders. Continued research, case studies, performance benchmarks, and side-by-side comparisons with current in vivo studies are critical to define the COU, validate methods, and establish confidence in NAMs. Such efforts will support consistent decision-making and facilitate global harmonization that is needed to transition NAMs from exploratory tools to regulatory standards without compromising human risk evaluation.
Salamandra is actively following these developments to support our clients in optimizing their development strategy and aligning with current regulatory expectations. If you have questions about your toxicology program or development strategy, reach out at hello@salamandra.net.
