## 5 Iron, ROS and Iron Chelators That Can Cause DNA Damage in Cancer  
ROS in normal cells can be beneficial for signalling in cell defence or development; ROS mainly arises in the mitochondria as a by-product of oxygen metabolism by oxygensincomplete reduction [158]. Low levels of ROS in normal cells contribute to the cell's proliferation, whereas ROS can contribute to apoptosis at intermediate levels. High levels of ROS contribute to necrosis; however, high levels can also be kept in check by antioxidant enzymes such as glutathione peroxidase [159]. In cancer, sustained growth signalling and proliferation factors are constantly turned on; consequently, there is an increased metabolic rate and hypoxia, resulting in heightened levels of ROS in cancer cells [160]. Cancer cells can also adapt to higher concentration of ROS by producing more antioxidants. The more elevated ROS occurring during oncogenic signalling by c-Myc are typically just under levels that would cause toxic levels of damage [161]. Cancer is known to rapidly grow in response to iron levels which enable DNA replication. Cancer cells therefore have an increased labile iron pool, and this resulting iron participates in the Fenton reaction to produce toxic radicals, contributing to a higher basal level of ROS. These high levels of iron can result in genomic instability due to the production of hydroxyl radicals that can damage DNA [162; 163]. Hydroxyl radicals produced by the Fenton reaction can react with membrane phospholipids causing lipid peroxidation. The hydroxyl radical can react with the fatty acid chain, generating a hydroperoxidised lipid and an alkyl radical, which can cause further damage [10]. Lipid peroxidation can result in cytotoxic aldehydes such as malonaldehyde and hydroxynonenal, which can react with the amino group in DNA bases to form adducts [164]. The hydroxyl radical from the Fenton reaction leads to hydrogen atom transfer (HAT) in deoxyribose, occurring at different carbon sites on the deoxyribose. ROS can react with the nitrogen bases in DNA, resulting in DNA fragmentation and causing issues involving the coiling of DNA in chromatin [165]. On pyrimidine, this often occurs at the 5, 6 pyrimidine bond and results in the HAT from the methyl group. In thymine, the hydroxyl radical can also react with the methyl group leading to 5-(uracily)methyl radical [166]. This can react with other bases such as adenine and guanine to produce interstrand cross-links. The most studied and common example of DNA damage by ROS is the addition of the hydroxyl radical with carbon 8 on guanine generating 8-hydroxy-7,8-dihydroquan-8-yl radicals. This either undergoes oxidation to form 8-oxo-guanine or reduction resulting in the imidazole ring opening and formation of N5-substituted formamidopyrimidine (N5-R-FAPy) lesions [167]. DNA damage can produce replication errors, aberrant transcription or signalling pathways, and, in theory, results in greater mutations and clonal evolution. This is also aided by the fact that high volumes of ROS can contribute to mitogenic signalling giving tumour cells a further growth advantage. An example is that the oncogenic KRAS requires ROS for proliferation [168]. ROS can cause changes to DNA and thus the genome leading to changes downstream in metabolic pathways or cell signalling, potentially favouring sustained proliferation. Mitochondrial DNA is even more susceptible to ROS as it has no histones and cannot participate in nuclear excisions repair [159]. This, in part, has given rise to the 'horse and cart' model. The model links mitochondrial metabolism (the 'horse') to (the 'cart') gene expression, where interconnected reactions in the mitochondria can lead to changes in gene expression [169]. When the balance shifts to favour ROS, it creates an environment that lends itself to genetic instability and self-deterioration. The mitochondrial labile iron pool is increased in cancer to fill the need for the incorporation of iron into crucial clusters.