How to Deal with the Nitrosamine Impurities in Human Drugs

The discovery in 2018 of nitrosamines in humans showed the need for better monitoring of these potent carcinogens

We are all exposed to some level of nitrosamines, also known as N-nitrosamines (NNAs). They commonly occur in water and foods, including cured and grilled meats, dairy products, and vegetables. Nitrosamines form a category of potent carcinogens known to induce tumours in animals. Nitrosamines can also be formed endogenously in the human body. The concern that these compounds might be ending up in human drugs has been raised since mid-2018, when, thanks to more sensitive analytical methods, the presence of several nitrosamine impurities has been detected in a type of blood pressure lowering drugs known as sartans. This discovery showed the need for better monitoring of NNA impurities in drug products, and led to several drug recalls including ranitidine, metformin and rifampicin.

What happened next?

Following these discoveries, global health authorities such as the US Food and Drug Administration and the European Medicines Agency have published guidance for drug manufacturers and authorization holders on how to control the levels of nitrosamine impurities in human drugs or how to avoid them entirely.

According to the guidance, the risk of possible formation of nitrosamine impurities in active substances or final drug products should be proactively assessed and the manufacturing processes should be designed in such a way that the formation of nitrosamines can be prevented or mitigated. If a risk of nitrosamine formation has been identified based on theoretical assessment involving the manufacturing process, starting materials, intermediates and excipients, confirmatory testing should be performed in order to detect the actual presence of nitrosamines in drug substances and products. It goes without saying that testing outcomes should be reported to the authorities.

At what levels should NNAs be controlled?

Since the Acceptable Intake (AI) for (potential) genotoxic impurities as defined by ICH-M7 is not sufficiently conservative for NNAs, several international authorities have developed the establishment of specific AI limit values for nitrosamines detected in human medicines, to ensure that the additional risk of cancer development in a patient from a lifelong intake of a nitrosamine impurity in a drug product does not exceed the acceptable rate of 1:100000. If a nitrosamine or a mixture of nitrosamines for which the AI has already been established, is detected in a drug substance or drug product, registrants should provide evidence that the AI is not exceeded for their drug products, taking into consideration the maximum daily dose of the drug product and the levels of the detected nitrosamine impurity.

For novel nitrosamines, for which no AIs have yet been set, the so-called Carcinogenic Potency Categorisation Approach (CPCA) should be followed to classify the nitrosamine in one of the 5 defined potency categories based on its chemical structure. Each potency category has been assigned a specific AI. Alternatively, if a structurally similar nitrosamine is available for which robust carcinogenicity data are available, a more specific AI could be derived, based on read-across – one approach frequently used in the risk assessment of substances that lack information on their toxicological properties—from the structural analogue.

Testing strategy of NNAs

Although many nitrosamines are potent genotoxic carcinogens, some nitrosamines, in particularly those derived directly from the drug substances (so-called Nitrosamine Drug Substance-Related Impurities, or NDSRIs) appear to exhibit lower potency or even to be not mutagenic. Their lower potency may be caused by the presence of bulky side chains in their chemical structure making them less accessible for activation by hepatic enzymes, or due to the presence of deactivating substituents.

For this type of NNAs, the generic approach may be too conservative. To investigate their real potency, nitrosamines can be tested in the Enhanced Ames Test (EAT), which is an in vitro reverse bacterial mutagenicity test designed specifically for nitrosamines. In comparison to the standard Ames test, the EAT has some additional elements, involving the use of a hamster metabolic activation system besides the standard rat system, and the additional use of two concurrent nitrosamine positive controls.

CRL contributed to the validation of this test as a partner in the HESI-GTTC Nitrosamine Research Program. A negative result in a GLP-compliant well-performed EAT is considered sufficient to control the nitrosamine at the AI level of 1.5 µg/day, which is the standard AI defined in the ICH-M7 guideline for mutagenic substances. Finally, to further de-risk nitrosamines, an in vivo mutagenicity study may be conducted to demonstrate that a nitrosamine is not mutagenic in vivo. A negative outcome for this endpoint would allow to control the impurity as any non-mutagenic impurity in drug substance or drug product in accordance with ICH-Q3A and/or ICH-Q3B guidelines.

Charles River can support you!

Due to our unique combination of experimental facilities and regulatory consulting activities, Charles River can support you each step of the way to make sure you fulfil your obligations, if a risk of nitrosamine formation is identified for your drug substance or drug product. You can learn more about the services we offer here. Feel free to reach out if you have questions about possible nitrosamine presence in your drug substances or drug products!