Mechanism-Based Insights into Removing the Mutagenicity of Aromatic Amines by Small Structural Alterations

Aromatic and heteroaromatic amines (ArNH₂) are activated by cytochrome P450 monooxygenases, primarily CYP1A2, into reactive N-arylhydroxylamines that can lead to covalent adducts with DNA nucleobases. Hereby, we give hands-on mechanism-based guidelines to design mutagenicity-free ArNH₂. The mechanism of N-hydroxylation of ArNH₂ by CYP1A2 is investigated by density functional theory (DFT) calculations. Two putative pathways are considered, the radicaloid route that goes via the classical ferryl-oxo oxidant and an alternative anionic pathway through Fenton-like oxidation by ferriheme-bound H₂O₂. Results suggest that bioactivation of ArNH₂ follows the anionic pathway. We demonstrate that H-bonding and/or geometric fit of ArNH₂ to CYP1A2 as well as feasibility of both proton abstraction by the ferriheme-peroxo base and heterolytic cleavage of arylhydroxylamines render molecules mutagenic. Mutagenicity of ArNH₂ can be removed by structural alterations that disrupt geometric and/or electrostatic fit to CYP1A2, decrease the acidity of the NH₂ group, destabilize arylnitrenium ions, or disrupt their pre-covalent transition states with guanine.