FDA draft guidance operationalizes animal testing phaseout

Background

As discussed in our prior alert on FDA’s April 2025 Roadmap on reducing animal testing, amendments to 21 U.S.C. § 355(z) under the FDA Modernization Act 2.0 expanded the definition of “nonclinical tests” to include cell based assays, organoids, microphysiological systems, computational modeling, and other non animal, human relevant methodologies. Consistent with that statutory change, FDA’s roadmap encouraged the use of scientifically validated NAMs in preclinical safety studies, including in silico tools such as AI/ML based predictive modeling, PBPK and QSP models; organoids and organ on chip platforms to assess target specific and off target effects; ex vivo human tissues; high throughput human cell based screening; microdosing and imaging studies in human volunteers; and refined in vivo approaches such as humanized transgenic mice as interim measures. Subsequently, the September 2025 MAHA Commission’s Strategy Report further reinforced FDA’s efforts to reduce animal testing by identifying it as a key priority for the Department of Health and Human Services.

Building on these efforts, a team of senior pharmacologists, toxicologists, and reviewers at the Center of Drug Evaluation and Research (CDER) published a December 2025 International Journal of Toxicology article describing the Center’s experience evaluating NAMs in regulatory submissions. The authors state that FDA has already accepted validated NAMs as replacements for specific animal tests (e.g., in vitro assays which are the current standard for ocular irritation, and skin sensitization) and has relied on fit for purpose, weight of evidence NAM approaches—including in vitro, and organ based models—to support investigational new drug applications, expand indications, and, in limited cases, inform approval where traditional animal models were not informative or feasible.

Notably, the authors state that NAM submissions have been challenged or rejected by the agency because the sponsor’s use of NAMs was insufficiently supported because the sponsor asserted an unclear context of use (COU), an insufficient demonstration of human biological relevance, an incomplete technical characterization, and/or data gaps that prevented FDA from relying on NAMs as a standalone basis for regulatory decision making. Taken together, these developments reflect a broader policy shift from encouraging NAM development to operationalizing their use in regulatory decision-making.

Most recently, FDA announced that it had met its first year milestones under the April 2025 Roadmap. Among other steps it has taken, the agency reported that it had qualified its first artificial intelligence based drug development tool, underscoring its growing willingness to rely on advanced in silico models to inform regulatory decision making when appropriately supported. It also launched a publicly searchable database identifying contexts in which alternative methods may be used in place of animal studies and increased coordination with international regulators to promote global alignment on NAM acceptance and validation strategies. FDA stated that, going forward, it intends to expand acceptable NAM use across additional endpoints and therapeutic areas.

Validation considerations for NAMs

FDA highlights four key features that sponsors should incorporate into their validation approach to ensure NAM studies can adequately support regulatory decision making. Collectively, these elements reflect FDA’s emphasis on a “fit-for-purpose” validation paradigm grounded in scientific context rather than a one-size-fits-all evidentiary standard:

1. Context of Use (COU): Similar to FDA’s expectation for other drug development tools, FDA emphasizes that each NAM should have a clearly defined regulatory purpose, a COU, which is tied to a specific scientific and/or drug development question. Adequate COUs should address data gaps and advance specific drug‑development objectives, such as informing patient monitoring in clinical trials, supporting dose selection, elucidating mechanisms underlying adverse events in animals or humans, justifying the exclusion of an animal species from nonclinical testing, or supporting a weight‑of‑evidence approach to predict risks not typically measured in humans.

2. Human biological relevance: FDA expects sponsors to demonstrate that the information generated from a NAM meaningfully reflects human biology and can inform assessment of drug safety in humans. FDA recommends that sponsors:

1. Describe the physiological features that will be assessed by the NAM, including the relevance of the cell types used, species of origin, and how the anatomical and physiological characteristics involved in toxicology findings may be mimicked in the NAM platform;
2. Demonstrate how relevant toxicological findings can be reliably evaluated in the NAM; and
3. Describe how the biological mechanisms investigated in the NAM are applicable to outcomes measured in human clinical trials.

3. Technical characterization: Consistent with “OECD Guidance Document on Good In Vitro Method Practices (GIVIMP), OECD Series on Testing and Assessment,” published in December 2018, FDA underscores the importance of robust technical characterization to ensure robustness, reliability and reproducibility of NAM-generated data. Sponsors should:

1. Describe test method details, including dosing, frequency of dosing, test substance preparation and storage, assay material compatibility, detection methods and instrumentation and any steps or processes that could introduce assay variability;
2. Explain statistical methods and criteria used for data analysis and interpretation, including criteria for positive, negative, or inconclusive results;
3. Demonstrate predictive performance of the NAM for the specified COU (e.g., sensitivity and specificity);
4. Provide information on assay stability and working duration, including consistency across batches of materials;
5. Document cell type used (e.g., primary, immortalized, stem cell-derived), cell/tissue isolation and/or differentiation methods, cell source (e.g., commercial, patient-derived, catalog and batch number), and species;
6. Address biological variability (e.g., genetics, donor variability) and donor cell phenotype (e.g., strain, sex, age) as they relate to the COU;
7. Justify the selection of reference compounds and controls; and
8. For complex platforms such as organ on a-chip systems, sponsors should address additional technical factors (e.g., flow, matrix, shear stress, and media), critical scaffold and matrix characteristics (e.g., biocompatibility, fabrication method, mechanical and biochemical properties, architecture, and substrate compatibility) and their impact on experimental variability and biological relevance, potential interactions with the test article and the platform, device, cell culture wells or material interface (e.g., material leaching or absorption that may alter toxicity or reduce target exposure, or biocompatibility issues of the biomaterial with the material interface).

4. Fit-for-purpose: Like other drug development tools, FDA expects that NAMs will be fit-for-purpose. FDA will find that a NAM is fit to support regulatory decision making if it replaces or offers an alternative approach to animal studies, fills a data gap or confirms and/or complements findings from traditional nonclinical testing. FDA recommends that sponsors demonstrate a NAM meaningfully supports regulatory decision making by ensuring the relevant regulatory submission:

1. Demonstrates that the NAM characterizes risk (e.g., sensitivity, specificity, positive and negative predictivity, false positive, and negative rates) at least as well as an established method where comparators exist;
2. Transparently describes the NAM’s strengths and limitations especially as it relates to the NAM’s reliability, reproducibility, and/or ability to inform a risk assessment for its intended COU; and
3. Explains how the NAM’s findings inform overall human health safety assessment.

Practical considerations for sponsors

Sponsors considering the use of NAMs should expect FDA to scrutinize their approach to validation. Early engagement with FDA review divisions remains critical, particularly where a sponsor intends to leverage a NAM to replace animal studies traditionally relied upon for IND enabling toxicology. The guidance signals that FDA expects NAM use to evolve iteratively. With this in mind, sponsors pursuing broader NAM use within their research and development efforts may consider submitting animal studies alongside corresponding NAM data to help FDA assess the reliability of the NAM and build agency familiarity and confidence in the tool’s performance. Sponsors may also consider using unvalidated and/or unqualified NAM data to support existing nonclinical data where there is a strong body of other evidence. Furthermore, sponsors should ensure that NAM development plans are prospectively aligned with the intended regulatory context of use, rather than retrofitting NAM data to support regulatory arguments after generation.

FDA has indicated that this draft guidance is part of a broader series of policy actions related to NAMs. Additional guidance addressing specific NAM technologies or therapeutic areas may be obtained from the agency through early consultation. FDA encourages drug developers to consult with the appropriate FDA review division when considering NAMs, particularly for indication-, disease-, organ-, and endpoint-specific applications. FDA has also indicated that it will continue to engage stakeholders through public workshops and interagency coordination on validation and acceptance of alternative methods.

Recently, the biotechnology industry collectively approached the International Society for Stem Cell Research (ISSCR) about generating consensus standards for NAMs, and in particular for organoids and assembloids. They expressed concern that variability in existing models is a significant challenge for stakeholders that need to be able to rely on these models to reduce reliance on animal testing—and to convince regulators to do the same. Biologics companies may wish to weigh in on such voluntary consensus standards as they are being developed.

Feel free to reach out to any of the authors of this alert or the Hogan Lovells attorney with whom you regularly work with any questions.

Authored by Heidi Gertner, Mike Druckman, Ashley Grey, and Mike Mortillo.