DNA Alkylation Damage by Nitrosamines and Relevant DNA Repair Pathways

Fahrer, Jörg; Christmann, Markus

doi:10.3390/ijms24054684

Cited by 25 publications

(14 citation statements)

References 257 publications

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“…A strictly linear relationship is not expected at very low exposures where saturation of detoxification pathways such as DNA repair and metabolism is highly unlikely. Fahrer and Christmann pointed out that many N -nitrosamines, particularly those found in drugs, are insufficiently studied so far with respect to DNA damage induction, and suggest that more studies are required to understand the structure–adduct repair relationship of more complex nitrosamines, such as NDSRIs. This knowledge would further improve the risk assessment.…”

Section: Resultsmentioning

confidence: 99%

Mechanisms of Nitrosamine Mutagenicity and Their Relationship to Rodent Carcinogenic Potency

Snodin,

Trejo-Martin,

Ponting

et al. 2024

Chem. Res. Toxicol.

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A thorough literature review was undertaken to understand how the pathways of N-nitrosamine transformation relate to mutagenic potential and carcinogenic potency in rodents. Empirical and computational evidence indicates that a common radical intermediate is created by CYP-mediated hydrogen abstraction at the α-carbon; it is responsible for both activation, leading to the formation of DNA-reactive diazonium species, and deactivation by denitrosation. There are competing sites of CYP metabolism (e.g., β-carbon), and other reactive species can form following initial bioactivation, although these alternative pathways tend to decrease rather than enhance carcinogenic potency. The activation pathway, oxidative dealkylation, is a common reaction in drug metabolism and evidence indicates that the carbonyl byproduct, e.g., formaldehyde, does not contribute to the toxic properties of N-nitrosamines. Nitric oxide (NO), a side product of denitrosation, can similarly be discounted as an enhancer of N-nitrosamine toxicity based on carcinogenicity data for substances that act as NO-donors. However, not all N-nitrosamines are potent rodent carcinogens. In a significant number of cases, there is a potency overlap with non-N-nitrosamine carcinogens that are not in the Cohort of Concern (CoC; high-potency rodent carcinogens comprising aflatoxin-like-, N-nitroso-, and alkyl-azoxy compounds), while other N-nitrosamines are devoid of carcinogenic potential. In this context, mutagenicity is a useful surrogate for carcinogenicity, as proposed in the ICH M7 (R2) (2023) guidance. Thus, in the safety assessment and control of N-nitrosamines in medicines, it is important to understand those complementary attributes of mechanisms of mutagenicity and structure−activity relationships that translate to elevated potency versus those which are associated with a reduction in, or absence of, carcinogenic potency.

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Section: Resultsmentioning

confidence: 99%

Mechanisms of Nitrosamine Mutagenicity and Their Relationship to Rodent Carcinogenic Potency

Snodin,

Trejo-Martin,

Ponting

et al. 2024

Chem. Res. Toxicol.

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“…Second, human methyltransferases have a much lower rate for removing O 4 -MeT adducts compared to the rat methyltransferases (Zak et al 1994 ). Third, N 3 -MeA adducts are repaired by alkyladenine glycosylase (AAG)-initiated base excision repair (BER) and AAG levels have remarkable effects on NDMA-induced point mutations and tumors in rodent liver (Fahrer and Christmann 2023 ; Kay et al 2021 ). It has been shown that AAG levels in peripheral blood mononuclear cells vary up to tenfold between individual humans (Fahrer and Christmann 2023 ).…”

Section: Discussionmentioning

confidence: 99%

“…Third, N 3 -MeA adducts are repaired by alkyladenine glycosylase (AAG)-initiated base excision repair (BER) and AAG levels have remarkable effects on NDMA-induced point mutations and tumors in rodent liver (Fahrer and Christmann 2023 ; Kay et al 2021 ). It has been shown that AAG levels in peripheral blood mononuclear cells vary up to tenfold between individual humans (Fahrer and Christmann 2023 ). Since the HepaRG cell line is derived from a single donor with unknown AAG levels, this mutation may not be common to all human hepatic cells.…”

Section: Discussionmentioning

confidence: 99%

Evaluating the mutagenicity of N-nitrosodimethylamine in 2D and 3D HepaRG cell cultures using error-corrected next generation sequencing

Seo,

Le,

Revollo

et al. 2024

Arch Toxicol

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Human liver-derived metabolically competent HepaRG cells have been successfully employed in both two-dimensional (2D) and 3D spheroid formats for performing the comet assay and micronucleus (MN) assay. In the present study, we have investigated expanding the genotoxicity endpoints evaluated in HepaRG cells by detecting mutagenesis using two error-corrected next generation sequencing (ecNGS) technologies, Duplex Sequencing (DS) and High-Fidelity (HiFi) Sequencing. Both HepaRG 2D cells and 3D spheroids were exposed for 72 h to N-nitrosodimethylamine (NDMA), followed by an additional incubation for the fixation of induced mutations. NDMA-induced DNA damage, chromosomal damage, and mutagenesis were determined using the comet assay, MN assay, and ecNGS, respectively. The 72-h treatment with NDMA resulted in concentration-dependent increases in cytotoxicity, DNA damage, MN formation, and mutation frequency in both 2D and 3D cultures, with greater responses observed in the 3D spheroids compared to 2D cells. The mutational spectrum analysis showed that NDMA induced predominantly A:T → G:C transitions, along with a lower frequency of G:C → A:T transitions, and exhibited a different trinucleotide signature relative to the negative control. These results demonstrate that the HepaRG 2D cells and 3D spheroid models can be used for mutagenesis assessment using both DS and HiFi Sequencing, with the caveat that severe cytotoxic concentrations should be avoided when conducting DS. With further validation, the HepaRG 2D/3D system may become a powerful human-based metabolically competent platform for genotoxicity testing.

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“…Temozolomide is manufactured through a highly verticalized synthetic route, where the process for the preparation is based on the patent WO2023102627 . Temozolomide is an antineoplastic alkylating agent and a pro-drug used in the oncological treatment of newly diagnosed patients with glioblastoma multiform, malignant glioma, and advanced metastatic malignant melanoma . After being absorbed in the body, it is converted into its active form, where it is capable of binding to DNA cell molecules, preventing their division and delaying the growth of the tumor mass.…”

Section: Introductionmentioning

confidence: 99%

Risk Assessment Study of Nitrosamines for the API Temozolomide Industrial Process

Secatto,

Massoni,

De Sio

2024

Org. Process Res. Dev.

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Since the International Agency for Research on Cancer (IARC) classified most of N-nitrosamines as potentially carcinogenic or mutagenic harmful agents, in addition to the recent worldwide episode where the presence of these substances was identified on medicines derived from sartans (e.g., Valsartan) and other active pharmaceutical ingredients (APIs) (e.g., Ranitidine, Nizatidine, and Metformin), regulatory agencies such as FDA, EMA, and ANVISA became concerned about the needs of pharmaceutical companies for evaluating their products and their respective manufacturing steps to ensure safety and quality. As the technical and regulatory approach to the subject advanced, it was established that the presence of these substances could be originated from different steps, both from API and finished product processes. Thus, several multidisciplinary committees were formed to discuss effective strategies to assess the qualitative and quantitative risk of contamination and formation of nitrosamines on products. In perspective on the production chain of active pharmaceutical ingredients, the aim of this study is to show the relevant results obtained from the risk assessment of nitrosamines for the API Temozolomide industrial process, produced by the pharmaceutical company Cristaĺia. The study focused on realizing a complete and robust risk assessment from the stages of synthesis and equipment evaluation to packaging and stability condition, applying in silico analyses verified by confirmatory analytical test through gas chromatography (GC) methodology.

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DNA Alkylation Damage by Nitrosamines and Relevant DNA Repair Pathways

Cited by 25 publications

References 257 publications

Mechanisms of Nitrosamine Mutagenicity and Their Relationship to Rodent Carcinogenic Potency

Mechanisms of Nitrosamine Mutagenicity and Their Relationship to Rodent Carcinogenic Potency

Evaluating the mutagenicity of N-nitrosodimethylamine in 2D and 3D HepaRG cell cultures using error-corrected next generation sequencing

Risk Assessment Study of Nitrosamines for the API Temozolomide Industrial Process

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