EMA Issues Paper On Qualifying Non-Mutagenic Impurities For Drug Toxicological Safety

By Tim Sandle, Ph.D.

Impurities are unwanted in pharmaceutical products, either as something that renders the product undesirable to the consumer or a substance that either affects drug efficacy or can cause patient harm.¹ They can be defined as any component of the drug product that is not the drug substance or an excipient.

Diverse sources of impurity in pharmaceutical products include residues of reagents, heavy metals, ligands, catalysts, and extracted materials like filter aids. Impurities also include degraded end products post-manufacturing as the result of hydrolysis, oxidative degradation, and other chemical reactions.² Impurities are conventionally categorized as organic impurities, inorganic impurities, or residual solvents.³ Identifying impurities is not always straightforward, and this may require the identification of the toxicophore (the portion of a structure that is related to the toxic properties of a chemical). To add to the complexity, some toxicophores require metabolic activation.⁴

Current regulatory guidance offers a framework for qualifying non-mutagenic impurities in various drug substances and products. However, there remains only limited guidance for assessing new or elevated impurity levels. To address this, the European Medicines Agency (EMA) has issued a “reflection paper” to consider what is needed for an adequate safety evaluation and how assessments can be made. ⁵ For assessments, the paper provides alternatives to animal studies in keeping the European Commission’s 3Rs initiative (replace, reduce, and refine the use of animal models) as per Directive 2010/63/EU.⁶ Impurities in advanced therapy medicinal products (ATMPs), in herbal medicinal products, and in biological and biotechnologically derived pharmaceuticals are out of scope, although companies working on products wherein both chemically synthesized and biotechnologically derived moieties are present (e.g., antibody–drug conjugates) may wish to consider the document.

Risk Evaluation

The reflection paper discusses the “Threshold of Toxicological Concern” (TTC) as the most effective tool for assessing low-level exposures by a user of a product to the pharmaceutical product itself. This is a concept referring to the establishment of a level of exposure for all chemicals, whether or not there are chemical-specific toxicity data below which there would be no appreciable risk to human health.⁷

The premise is, where the chemical structure of an impurity is known, then the likely health risk can be evaluated using generic human thresholds of exposure or TTC values. These values exist for established substances of similar chemical structure. The process also accounts for the likelihood of toxicity, drawing on published toxicological data. Substances can be considered as being either low, moderate, or high toxicity.

Risk Framework And Multifactorial Matters

Non-mutagenic impurities need to be considered using a suitable risk framework and such a risk framework needs to consider what occurs when more than one impurity is present. The EMA paper calls for a multifactorial approach, considering dependent factors, including genetic and environmental factors. Of concern for pharmaceuticals are the exposure level, route of administration, physicochemical properties, bioavailability, degradability, clinical conditions, and target population (such as noting any applicable differences between children and adults).

Given the complexity of the risk assessment, the use of an in silico tool is recommended. Advances in machine learning, artificial intelligence, and quantitative structure–activity relationship models can strengthen computational toxicological approaches.⁸

Data Generation

In the absence of adequate historical data, the reflection paper describes alternative assessments described as New Approach Methodologies (NAMs). These involve the characterization of the chemical properties of the impurity and the application of computational toxicology tools. Examples of NAM include biological observations from in vitro assays, toxicogenomics, metabolomics, and receptor binding screens. Often NAM approaches use physiologically based kinetic modeling to inform scientists about systemic toxicological exposure.⁹ Recommended alternatives to animal models include 2D and 3D cell systems and microphysiological systems.

Where alternative methods fail to generate the required data, then the paper recommends that animal models can be considered to assess the toxicological nature of the substance. As well as animal welfare, a limitation of the use of animal models is a general inability to detect toxic impurities in comparison with alternative assays.¹⁰ Where animal models are deployed, the paper recommends that a suitable duration is built into the experiment (such as assessing the animal reaction to the product over the course of 28 days).

Risk Outcomes

After either a single risk evaluation or a multifactorial evaluation has been completed (depending on the weight of factors and number of identified impurities), the reflection paper requires an acceptable level (AL) to be calculated. This is an estimation of the product-specific safe level of exposure to impurities. To complete this assessment, an understanding of the permitted daily exposure (PDE), bioavailability, read-across (RAX) data, and product-specific considerations need to be known. A weight-of-evidence approach can be used to assess the different areas of concern. This allows a combination of information from several independent sources to be used and assessed to strengthen the predictive model.

With each of the estimates:

• The PDE represents a substance-specific daily dose that is unlikely to cause an adverse effect if an individual is exposed each day for a lifetime. This requires knowledge of “critical effects” and understanding the no-observed-adverse-effect level, with the use of several adjustment factors to account for various uncertainties.¹¹
• Bioavailability refers to the extent a substance or drug becomes completely available to its intended biological destination(s). By the same implication, the impurity could have the same level of availability as the active drug substance.¹²
  • Importantly, bioavailability via different administration routes may vary greatly.
• RAX is a type of chemical assessment where a “read-across” method is used to fill data gaps. In the context of TTC, the approach involves looking for toxicological similarities of known chemical substances to the impurity.¹³
• Product-specific considerations are concerned with the safety of medicine. This is determined by direct or indirect pharmacological evidence, including immunogenic properties of the active substance, of the excipients, and of process-related impurities.¹⁴

In summary, the reflection paper encourages pharmaceutical developers to adopt an integrated risk assessment approach to determine whether a non-mutagenic impurity can be considered safe or otherwise. The EMA is accepting public comments through April 30, 2025.

References

1. ICH. Specifications: Test Procedures And Acceptance Criteria For New Drug Substances And New Drug Products: Chemical Substances Q6A, ICH, 1999: https://database.ich.org/sites/default/files/Q6A%20Guideline.pdf
2. Pilaniya K, Chandrawanshi HK, Pilaniya U et al. Recent trends in the impurity profile of pharmaceuticals. J Adv Pharm Technol Res. 2010;1(3):302-10. doi: 10.4103/0110-5558
3. ICH Q3D Elemental impurities - Scientific guideline, ICH, September 2022: https://www.ema.europa.eu/en/documents/scientific-guideline/international-conference-harmonisation-technical-requirements-registration-pharmaceuticals-human-use-ich-q3d-elemental-impurities-step-5-revision-2_en.pdf
4. Williams, D.P.; Naisbitt, D.J. (2002) Toxicophores: Groups and Metabolic Routes Associated with Increased Safety Risk. Current Opinion in Drug Discovery & Development. 5 (1): 104–115
5. EMA. Reflection paper on the qualification of non-mutagenic impurities, European Medicines Agency, EMA/CHMP/543397/2024, 2nd December 2024: https://www.ema.europa.eu/en/documents/scientific-guideline/draft-reflection-paper-qualification-non-mutagenic-impurities_en.pdf
6. Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes, EUR-lex, 26th June 2019: https://eur-lex.europa.eu/legal-content/EN/TXT/?uri=celex:32010L0063
7. Kroes R, Kleiner J, Renwick A. The threshold of toxicological concern concept in risk assessment. Toxicol Sci. 2005;86(2):226-30
8. OECD (Q)SAR Assessment Framework: Guidance for the regulatory assessment of (Quantitative) Structure Activity Relationship models and predictions, https://one.oecd.org/document/ENV/CBC/MONO(2023)32/en/pdf
9. Alexander-White C, Bury D, Cronin M. et al. A 10-step framework for use of read-across (RAX) in next generation risk assessment (NGRA) for cosmetics safety assessment. Regul Toxicol Pharmacol. 2022;129:105094
10. Slikkerveer A, Doehr O, Claude N. et al. New limits proposed for the management of non-mutagenic impurities. Regul. Toxicol. Pharmacol. 2024:150:105647
11. Ball DJ, Beierschmitt WP. Permitted Daily Exposure Values: Application Considerations in Toxicological Risk Assessments. Int J Toxicol. 2020;39(6):577-585
12. Welling, P. G.; Tse, F. L.; Dighe, S V. (1991) Pharmaceutical Bioequivalence. Drugs and the Pharmaceutical Sciences. Vol. 48. New York, NY: Marcel Dekker.
13. Ouedraogo G, Alexander-White C, Bury D, et al. Cosmetics Europe. Read-across and new approach methodologies applied in a 10-step framework for cosmetics safety assessment - A case study with parabens. Regul Toxicol Pharmacol. 2022 Jul;132:105161.
14. Engelhardt JA, Dorato MA The no-observed-adverse-effect-level in drug safety evaluations: Use, issues, and definition(s). Regul Toxicol Pharmacol. 2005; 42 (3): 265–274

About The Author:

Tim Sandle, Ph.D., is a pharmaceutical professional with wide experience in microbiology and quality assurance. He is the author of more than 30 books relating to pharmaceuticals, healthcare, and life sciences, as well as over 170 peer-reviewed papers and some 500 technical articles. Sandle has presented at over 200 events and he currently works at Bio Products Laboratory Ltd. (BPL), and he is a visiting professor at the University of Manchester and University College London, as well as a consultant to the pharmaceutical industry. Visit his microbiology website at https://www.pharmamicroresources.com.