Scoping review

Scoping Review: New Approach Methodologies (NAMs) for Mutagenicity Assessment of Nitrosamine Drug Substance-Related Impurities (NDSRIs) under ICH M7(R2)

Rationale

Since 2018, an emerging problem has been identified globally in pharmaceutical manufacturing: the formation of nitrosamine impurities through the reaction of active pharmaceutical ingredients (APIs) with a nitrosating agent (EMA, 2025; FDA, 2023; Ruep et al., 2021). Although nitrosamine formation is a natural process, when it occurs at unacceptable levels during medicine production it may result in carcinogenic contamination. Several pharmaceuticals have a documented history of nitrosamine contamination, including sartans, ranitidine, nizatidine, rifampicin, and metformin, in which the contaminating impurities, referred to as nitrosamine drug substance related impurities (NDSRIs), led to recalls of medicines in the US market (reviewed in Aishwarya et al., 2025; Vikram et al., 2024). This emerging issue represents a significant economic and public health problem, as nitrosamines have been associated with conditions like lung, liver, oesophageal, and breast cancer (Vikram et al., 2024).
To address this issue, the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) updated its M7 guideline (ICH M7(R2)). This guideline addresses the identification, categorization, qualification, and control of mutagenic impurities to limit potential carcinogenic risk from nitrosamine exposure. It favors the use of alternatives to animal testing, such as the Ames assay, literature based assessment, and computational models. However, when resulting impurity levels cannot be controlled to an acceptable limit, ICH M7(R2) recommends confirming the relevance of the bacterial mutagenicity result with an in vivo gene mutation assay.
Some authors argue that animal based methods still offer a superior approach for the control of mutagenic contamination (Aishwarya et al., 2025; Dande et al., 2025). This raises the question that motivated the present review: what efforts have been made to increase the use of NAMs for this confirmatory purpose.

Objectives
The objective of this project is to conduct a scoping review to map the use of NAMs to assess mutagenicity in compliance with ICH M7(R2), and to map the biological and methodological factors underlying the advantages and disadvantages of these approaches compared to current in vivo methods.

Methodology
This scoping review will follow the PRISMA-ScR (Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews) checklist (Tricco et al., 2018). A review protocol has been developed a priori, in line with Peters et al. (2020), and will be made publicly available on a dedicated platform ahead of the review’s execution. Any amendments to the protocol will be clearly documented, justified, and reported in the resulting publication.
A preliminary literature search was conducted to assess the feasibility of the review, and the results supported moving forward with a full scoping review on this topic. The formal search strategy will draw on multiple biomedical databases, with records screened independently by two investigators against predefined eligibility criteria.

Status and outcomes
This project is ongoing. If successful, it will produce a structured evidence framework categorizing current options for mutagenicity assessment of NDSRIs using NAMs, summarizing the biological, methodological, and regulatory considerations associated with these approaches and highlighting where further research is needed. Findings will be submitted to a peer reviewed journal in toxicology, regulatory science, or alternative methods, and shared through this platform and our LinkedIn page.

Open to collaborations: Yes. If you are interested in contributing to this project, get in touch.

References

Aishwarya, D., Ramakant Dhampalwar, V., Pallaprolu, N., & Peraman, R. (2025). Nitrosamine drug substance-related impurities (NDSRIs) in pharmaceuticals: Formation, mitigation strategies, and emphasis on mutagenicity risks. Pharmaceutical Research, 42(4), 547–578. https://doi.org/10.1007/s11095-025-03857-9

Dande, A., Chandra Mouli, H. M., Nandy, J., Shreya, S., Pallaprolu, N., Samanthula, G., & Peraman, R. (2025). Risk assessment, detection and control of mutagenic impurities in pharmaceuticals: Emphasis on nitrosamines. Critical Reviews in Analytical Chemistry. Advance online publication. https://doi.org/10.1080/10408347.2025.2517357

European Medicines Agency. (2025, February 28). Nitrosamine impurities in specific medicines. https://www.ema.europa.eu/en/human-regulatory-overview/post-authorisation/pharmacovigilance-post-authorisation/referral-procedures-human-medicines/nitrosamine-impurities/nitrosamine-impurities-specific-medicines

Peters MDJ, Godfrey C, McInerney P, Munn Z, Tricco AC, Khalil, H. Scoping Reviews (2020). Aromataris E, Lockwood C, Porritt K, Pilla B, Jordan Z, editors. JBI Manual for Evidence Synthesis. JBI; 2024. Available from: https://synthesismanual.jbi.global.  https://doi.org/10.46658/JBIMES-24-09

Ruepp, R., Frötschl, R., Bream, R., Filancia, M., Girard, T., Spinei, A., Weise, M., & Whomsley, R. (2021). The EU response to the presence of nitrosamine impurities in medicines. Frontiers in Medicine, 8, Article 782536. https://doi.org/10.3389/fmed.2021.782536

Tricco, A. C., Lillie, E., Zarin, W., O’Brien, K. K., Colquhoun, H., Levac, D., Moher, D., Peters, M. D. J., Horsley, T., Weeks, L., Hempel, S., Akl, E. A., Chang, C., McGowan, J., Stewart, L., Hartling, L., Aldcroft, A., Wilson, M. G., Garritty, C., Lewin, S., Godfrey, C. M., Macdonald, M. T., Langlois, E. V., Soares-Weiser, K., Moriarty, J., Clifford, T., Tunçalp, Ö., & Straus, S. E. (2018). PRISMA extension for scoping reviews (PRISMA-ScR): Checklist and explanation. Annals of Internal Medicine, 169(7), 467–473. https://doi.org/10.7326/M18-0850

U.S. Food and Drug Administration. (2023). Recommended acceptable intake limits for nitrosamine drug substance-related impurities (NDSRIs): Guidance for industry. Center for Drug Evaluation and Research.

Vikram, H. P. R., Kumar, T. P., Kumar, G., Beeraka, N. M., Deka, R., Suhail, S. M., Jat, S., Bannimath, N., Padmanabhan, G., Chandan, R. S., Kumar, P., & Gurupadayya, B. (2024). Nitrosamines crisis in pharmaceuticals—Insights on toxicological implications, root causes and risk assessment: A systematic review. Journal of Pharmaceutical Analysis, 14(5), Article 100919. https://doi.org/10.1016/j.jpha.2023.12.009