Predicting DNA-Reactivity of N-Nitrosamines: A Quantum Chemical Approach

Abstract

N-Nitrosamines (NAs) are a class of reactive organic chemicals that humans may be exposed to from environmental sources, food but also impurities in pharmaceutical preparations. Some NAs were identified as DNA-reactive mutagens and many of those have been classified as probable human carcinogens. Beyond high-potency mutagenic carcinogens that need to be strictly controlled, NAs of low potency need to be considered for risk assessment as well. NA impurities and nitrosylated products of active pharmaceutical ingredients (APIs) often arise from production processes or degradation. Most NAs require metabolic activation to ultimately become carcinogens, and their activation can be appropriately described by first-principles computational chemistry approaches. To this end, we treat NAinduced DNA alkylation as a series of subsequent association and dissociation reaction steps that can be calculated stringently by density functional theory (DFT), including α-hydroxylation, proton transfer, hydroxyl elimination, direct S N 2/S N Ar DNA alkylation, competing hydrolysis and S N 1 reactions. Both toxification and detoxification reactions are considered. The activation reactions are modeled by DFT at a high level of theory with an appropriate solvent model to compute Gibbs free energies of the reactions (thermodynamical effects) and activation barriers (kinetic effects). We study congeneric series of aliphatic and cyclic NAs to identify trends. Overall, this work reveals detailed insight into mechanisms of activation for NAs, suggesting that individual steric and electronic factors have directing and rate-determining influence on the formation of carbenium ions as the ultimate pro-mutagens and thus carcinogens. Therefore, an individual risk assessment of NAs is suggested, as exemplified for the complex API-like 4-(N-nitroso-N-methyl)aminoantipyrine which is considered as low-potency NA by in silico prediction.

Quantum-Mechanical Approach to Predicting the Carcinogenic Potency of N-Nitroso Impurities in Pharmaceuticals

“…16−18 For NOCs, quantum-mechanics (QM) modeling of molecular interactions in key events (KEs) is particularly appealing given the limited data and the welldescribed mechanisms of reactivity and in turn the carcinogenicity. 18 This line of reasoning is consistent with the recent report by Cross and Ponting, which emphasized the need for SAR improvements based on N-nitroso reaction mechanisms. 5 Furthermore, because mutagenicity has shown high sensitivity in the Ames test in predicting rodent carcinogenicity, 6,19 the KEs can be related to activation mechanisms that control DNA alkylation rates.…”

“…Fortunately, today’s in silico toolkit is not limited to statistics-focused approaches; established SARs can fuel the development of expert systems, such as those reported by Thomas et al, or “physics-led” models based on computational chemistry. − For NOCs, quantum-mechanics (QM) modeling of molecular interactions in key events (KEs) is particularly appealing given the limited data and the well-described mechanisms of reactivity and in turn the carcinogenicity . This line of reasoning is consistent with the recent report by Cross and Ponting, which emphasized the need for SAR improvements based on N -nitroso reaction mechanisms .…”

“…Furthermore, because mutagenicity has shown high sensitivity in the Ames test in predicting rodent carcinogenicity, , the KEs can be related to activation mechanisms that control DNA alkylation rates . While QM approaches to study N -nitroso reactivity are not new, ,− to the best of our knowledge, no study has incorporated modern density functional theory (DFT) methods into a robust tool to predict carcinogenic potency in support of substance-specific AI calculations as per ICH M7­(R1) guidelines . This effort requires careful deconstruction of QM-based reaction-pathway modeling into computationally less-demanding steric and electronic factors that drive the formation of mutagens (and thus carcinogens) and can afford hazard assessments in reasonable timeframes competitive with other nonanimal approaches.…”

A Quantum-Mechanical Approach to Predicting Carcinogenic Potency of N-nitrosamine Impurities in Pharmaceuticals

“…14,15 For N-nitrosamines, quantum-mechanics (QM) modeling of molecular interactions in key initiating events (KIEs) is the logical next step in predicting their reactivity, and in turn carcinogenicity. 16 This line of reasoning is consistent with the recent report by Cross and Ponting, which emphasized the need for SAR improvements based on N-nitrosamine reaction mechanisms. 4 Furthermore, because mutagenicity has shown high sensitivity in Ames tests in predicting rodent carcinogenicity, 5,17 the KIEs can be related to the metabolic activation mechanisms by Cytochrome P450 (CYP), which control DNA alkylation rates of N-nitroso compounds.…”

“…4 Furthermore, because mutagenicity has shown high sensitivity in Ames tests in predicting rodent carcinogenicity, 5,17 the KIEs can be related to the metabolic activation mechanisms by Cytochrome P450 (CYP), which control DNA alkylation rates of N-nitroso compounds. 18 While QM approaches to study N-nitrosamine reactivity and the P450 metabolism are not new, 16,[19][20][21] to the best of our knowledge, no study has incorporated modern density functional theory (DFT) methods into a robust tool to predict carcinogenicity potency in support of substance-specific AI calculations as per ICH M7(R1) guidelines. 22 This effort requires careful deconstruction of QMbased reaction-pathway modeling 16 into computationally less-demanding steric and electronic factors that drive the formation of mutagens (and thus carcinogens), and can afford hazard assessments in reasonable timeframes competitive with other nonanimal approaches.…”