Free radicals, metals and antioxidants in oxidative stress-induced cancer

Elevated rates of reactive oxygen species (ROS) have been detected in almost all cancers, where they promote many aspects of tumour development and progression. However, tumour cells also express increased levels of antioxidant proteins to detoxify from ROS, suggesting that a delicate balance of intracellular ROS levels is required for cancer cell function. Further, the radical generated, the location of its generation, as well as the local concentration is important for the cellular functions of ROS in cancer. A challenge for novel therapeutic strategies will be the fine tuning of intracellular ROS signalling to effectively deprive cells from ROS-induced tumour promoting events, towards tipping the balance to ROS-induced apoptotic signalling. Alternatively, therapeutic antioxidants may prevent early events in tumour development, where ROS are important. However, to effectively target cancer cells specific ROS-sensing signalling pathways that mediate the diverse stress-regulated cellular functions need to be identified. This review discusses the generation of ROS within tumour cells, their detoxification, their cellular effects, as well as the major signalling cascades they utilize, but also provides an outlook on their modulation in therapeutics.

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Reactive oxygen species in cancer

It is increasingly proposed that reactive oxygen species (ROS) and reactive nitrogen species (RNS) play a key role in human cancer development [1–6], especially as evidence is growing that antioxidants may prevent or delay the onset of some types of cancer (reviewed in [7,8]). ROS is a collective term often used by biologists to include oxygen radicals [superoxide (O # J−), hydroxyl (OHJ), peroxyl (RO # J) and alkoxyl (ROJ)] and certain nonradicals that are either oxidizing agents and}or are easily converted into radicals, such as HOCl, ozone (O $ ), peroxynitrite (ONOO−), singlet oxygen ("O # ) and H # O # . RNS is a similar collective term that includes nitric oxide radical (NOJ), ONOO−, nitrogen dioxide radical (NO # J), other oxides of nitrogen and products arising when NOJ reacts with O # J−, ROJ and RO # J. ‘Reactive ’ is not always an appropriate term; H # O # , NOJ and O # J− react quickly with very few molecules, whereas OHJ reacts quickly with almost anything. RO # J, ROJ, HOCl, NO # J, ONOO− and O $ have intermediate reactivities. ROS and RNS have been shown to possess many characteristics of carcinogens [4] (Figure 1). Mutagenesis by ROS}RNS could contribute to the initiation of cancer, in addition to being important in the promotion and progression phases. For example, ROS}RNS can have the following effects. (1) Cause structural alterations in DNA, e.g. base pair mutations, rearrangements, deletions, insertions and sequence amplification. OHJ is especially damaging, but "O # , RO # J, ROJ, HNO # , O $ , ONOO− and the decomposition products of ONOO− are also effective [9–13]. ROS can produce gross chromosomal alterations in addition to point mutations and thus could be involved in the inactivation or loss of the second wild-type allele of a mutated proto-oncogene or tumour-suppressor gene that can occur during tumour promotion and progression, allowing expression of the mutated phenotype [4]. (2) Affect cytoplasmic and nuclear signal transduction pathways [14,15]. For example, H # O # (which crosses cell and organelle membranes easily) can lead to displacement of the inhibitory subunit from the cytoplasmic transcription factor nuclear factor κB, allowing the activated factor to migrate to the nucleus [14]. Nitration of tyrosine residues by ONOO− may block phosphorylation. (3) Modulate the activity of the proteins and genes that respond to stress and which act to regulate the genes that are related to cell proliferation, differentiation and apoptosis [4,14–17]. For example, H # O # can stimulate transcription of c-jun

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Damage to DNA by reactive oxygen and nitrogen species: role in inflammatory disease and progression to cancer.

Oxidative Nucleobase Modifications Leading to Strand Scission

Organisms are constantly exposed to various forms of reactive oxygen species (ROS) that lead to oxidation of proteins, nucleic acids, and lipids. Protein oxidation can involve cleavage of the polypeptide chain, modification of amino acid side chains, and conversion of the protein to derivatives that are highly sensitive to proteolytic degradation. Unlike other types of modification (except cysteine oxidation), oxidation of methionine residues to methionine sulfoxide is reversible; thus, cyclic oxidation and reduction of methionine residues leads to consumption of ROS and thereby increases the resistance of proteins to oxidation. The importance of protein oxidation in aging is supported by the observation that levels of oxidized proteins increase with animal age. The age-related accumulation of oxidized proteins may reflect age-related increases in rates of ROS generation, decreases in antioxidant activities, or losses in the capacity to degrade oxidized proteins.

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Protein oxidation and aging

Endogenous DNA adducts: potential and paradox

The peroxidation of polyunsaturated lipids generates a range of substances that possess DNA damaging potential. This includes lipid hydroperoxides and various species that contain unpaired electrons, such as the alkoxyl and peroxyl radicals. In addition, a range of genotoxic carbonyl-containing compounds are formed, such as malondialdehyde, various 4-hydroxy-2-alkenals such as 4-hydroxynonenal and a number of 2-alkenals. It has previously been assumed that the antioxidants and electrophile scavenging enzymes existing in mammalian cells effectively protect the genetic material against these substances. However, thanks to recent analytical advances in the detection of low levels of DNA adducts, it is now evident that DNA adducts formed from a range of lipid peroxidation products are abundant in both rodent and human genomes. This suggests that the cellular defence system is not 100% efficient and that a proportion of endogenously produced lipid peroxidation products escape detoxification and cause DNA damage. This review surveys the genotoxic properties of the major classes of lipid peroxidation products, focusing on their chemistry of DNA adduction, the mutagenic properties of such damage and the evidence that it occurs in intact biological systems. Furthermore, avenues of future research that will clarify the significance of such damage to spontaneous mutagenesis and carcinogenesis are proposed and discussed.

Genotoxic lipid peroxidation products : their DNA damaging properties and role in formation of endogenous DNA adducts

The toxicology of the cyanobacterial alkaloid cylindrospermopsin (CYN), a potent inhibitor of protein synthesis, appears complex and is not well understood. In exposed mice the liver is the main target for the toxic effects of CYN. In this study primary mouse hepatocyte cultures were used to investigate the mechanisms involved in CYN toxicity. The results show that 1–5 μM CYN caused significant concentration-dependent cytotoxicity (52%–82% cell death) at 18 h. Protein synthesis inhibition was a sensitive, early indicator of cellular responses to CYN. Following removal of the toxin, the inhibition of protein synthesis could not be reversed, showing behavior similar to that of the irreversible inhibitor emetine. In contrast to the LDH leakage, protein synthesis was maximally inhibited by 0.5 μM CYN. No protein synthesis occurred over 4–18 h at or above this concentration. Inhibition of cytochrome P450 (CYP450) activity with 50 μM proadifen or 50 μM ketoconazole diminished the toxicity of CYN but not the effects on protein synthesis. These findings imply a dissociation of the two events and implicate the involvement of CYP450-derived metabolites in the toxicity process, but not in the impairment of protein synthesis. Thus, the total abolition of protein synthesis may exaggerate the metabolite effects but cannot be considered a primary cause of cell death in hepatocytes over an acute time frame. In cell types deficient in CYP450 enzymes, protein synthesis inhibition may play a more crucial role in the development of cytotoxicity. © 2003 Wiley Periodicals, Inc. Environ Toxicol 18: 243–251, 2003.

Cylindrospermopsin-induced protein synthesis inhibition and its dissociation from acute toxicity in mouse hepatocytes

Acetaminophen Toxicity Results in Site-specific Mitochondrial Damage in Isolated Mouse Hepatocytes*

Cylindrospermopsin (CYN) is a cyanobacterial toxin found in drinking-water sources world wide. It was the likely cause of human poisonings in Australia and possibly Brazil. Although CYN itself is a potent protein synthesis inhibitor, its acute toxicity appears to be mediated by cytochrome p-450 (CYP450)-generated metabolites. CYN also induces genotoxic effects both in vitro and in vivo, and preliminary evidence suggests that tumors are generated by oral exposure to CYN. To understand the role of CYP450-activated CYN metabolites on in vitro genotoxicity, this study quantified the process in primary mouse hepatocytes using the COMET assay in both the presence and absence of CYP450 inhibitors known to block acute CYN cytotoxicity. CYN was cytotoxic at concentrations above 0.1 μM(EC50 = 0.5 μM) but produced significant increases in Comet tail length, area, and tail moment at 0.05 μM and above; hence genotoxicity is unlikely to be secondary to metabolic disruption due to toxicity. The CYP450 inhibitors omepraz...

Philip C. Burcham

Papers

Endogenous DNA adducts: potential and paradox

Genotoxic lipid peroxidation products : their DNA damaging properties and role in formation of endogenous DNA adducts

Cylindrospermopsin-induced protein synthesis inhibition and its dissociation from acute toxicity in mouse hepatocytes

Acetaminophen Toxicity Results in Site-specific Mitochondrial Damage in Isolated Mouse Hepatocytes*

Cylindrospermopsin genotoxicity and cytotoxicity: role of cytochrome P-450 and oxidative stress.