Nucleic acid oxidation by the peroxidase systems of activated phagocytes: Products, mechanisms, and biological activity.

摘要

Oxidative modification of DNA has been implicated in the development of inflammation-associated cancers. However, the mechanisms by which this process may occur remain poorly understood. Genetic epidemiologic studies have suggested that myeloperoxidase (MPO), a phagocytic heme enzyme that produces halogenating oxidants, may play an important role in the development of some human cancers.; Our studies began with the observation that, among the four major nucleosides, deoxycytidine was particularly susceptible to modification by MPO. We demonstrated that the deoxycytidine oxidation product is 5-chlorodeoxycytidine and is readily produced upon stimulation of human neutrophils in culture. We also observed that eosinophil peroxidase (EPO), an enzyme related to MPO, generates brominating oxidants to produce 5-bromodeoxycytidine in an analogous reaction. Both phagocytic peroxidases were also found to halogenate the RNA base uracil to the mutagenic thymine analogs 5-chloro- and 5-bromouracil.; To determine which oxidants are responsible for the observed nucleobase halogenation, we replaced the enzyme and cell systems with purified hypochlorous and hypobromous acid, the primary products of MPO and EPO, respectively. The experimental results support a role for hypochlorous acid, chlorine gas (Cl 2), hypobromous acid, and bromamines in nucleobase halogenation by phagocytes and their peroxidases. Bromination of deoxycytidine and uracil by neutrophils and MPO was found to be consistent with formation of the interhalogen gas bromine chloride (BrCl).; Because experiments with bacteria indicated that DNA is protected from halogenation relative to RNA, we examined the ability of halogenated DNA precursors to become incorporated into DNA. Consistent with this scenario, we found that EPO-generated 5-bromodeoxycytidine was deaminated and incorporated into the DNA of cultured mammalian cells. We also observed that 5-bromouracil generated by EPO was converted to the mutagenic DNA precursor 5-bromodeoxyuridine by the thymine salvage enzyme thymidine phosphorylase.; Next, to determine if nucleobase halogenation occurs in vivo , we analyzed human tissue for free halogenated uracil. Mass spectrometric analyses revealed high levels of 5-chlorouracil and 5-bromouracil in inflammatory exudate and undetectable levels in normal human plasma and urine. Collectively, our studies demonstrate that phagocytic peroxidases generate mutagens which may play a role in mutagenesis and the development of cancer at sites of inflammation.