**3.3 Mutations**

The failure of repair mechanisms and constant exposure to PAHs induce mutagenesis in cells. These mutations are present in multiple genes including those that participate in cell survival. In particular, p53 mutations are associated with risk of carcinogenesis in PAHexposed individuals. Since the p53 protein is a transcription factor that regulates cell proliferation, differentiation, apoptosis, and DNA repair, mutations induced in this important protein could lead to severe damage in cells and genes. Some studies have associated p53 mutations to PAH exposure (Mordukhovich, et al., 2010, Yoon, et al., 2003). Another common target of mutagenesis is the ras gene (Ross & Nesnow, 1999). A study by Gray et al. (2001) revealed that exposure to BaP in mice increases mutation of the K-ras gene.

#### **3.4 Carcinogenesis**

Forming adducts in DNA repair-related genes is not the only mechanism by which PAHs induce carcinogenesis. An additional danger amounts from their resemblance to steroid hormones allowing PAHs the ability to activate estrogen receptors and metabolism. The ability of several PAHs to displace natural estrogens and occupy ER binding sites, at least to some extent, implies a potential mechanism of action in endocrine tissues that is ER-

DNA Damage Caused by Polycyclic Aromatic Hydrocarbons: Mechanisms and Markers 133

the initial detection of damaged DNA. On the other hand, TC-NER does not require XPC, however the stalled RNA Polymerase complex is displaced in order to allow the NER proteins to access the damaged DNA. After this process, TC-NER and GG- NER proceed in identical ways. XPA and RPA (Replication Protein-A) then bind at the location of injury and further aid in damage detection. Subsequently, the XPB and XPD helicases unwind the DNA duplex in the surrounding area of the lesion. The endonucleases XPG and ERCC1 (Excision Repair Cross-Complementing group-1)/XPF then cleave one strand of the DNA at positions 3' and 5' to the damage, respectively, generating a 30 base oligonucleotide containing the lesion. This oligonucleotide is displaced, permitting gap repair synthesis (performed by DNA Pol Delta/Epsilon, and other accessory proteins). Finally, DNA ligase seals the nick in the repaired strand (Fig. 6). Several studies demonstrate that certain polymorphisms in NER genes alter the efficiency of DNA repair (Shen, et al., 2006, Vodicka, et al., 2004). Four polymorphisms, XPA - 4G/A (rs1800975), ERCC1 C8092A (rs3212986), XPD Lys751Gln (rs1052559), and XPF Ser835Ser (rs1799801), are associated with a reduced capacity for DNA repair and an increased susceptibility to various cancers (Hu, et al., 2004, Monzo, et al., 2007). However, a recent study reports that significant opportunities exist for an interaction between the XPA-4 G/A polymorphism and PAH exposure on sperm DNA damage (Gu, et al., 2010). Although some PAHs lack "bay" and/or "fjord" regions, such as anthracene for example, this molecule also induces DNA damage, activating repair mechanisms, and has in fact has been shown to induce NER and MMR pathways (Desler, et al., 2009).These findings provide support for the

BER involves the combined activity of some specific proteins that recognize and excise DNA damage, replacing the damaged moiety with normal nucleotides. PAH adducts are repaired by this mechanism. BER consists of three important steps: first, removal of the incorrect base by an appropriate DNA N-glycosylase to create an AP site (apurinic/apyrimidinic site); second, cutting off the damaged DNA strand by AP endonuclease upstream of the AP site, therefore creating a 3'-OH terminus adjacent to the AP site and finally, extension of the 3'- OH terminus by DNA polymerase, accompanied by excision of the AP site. Several enzymes are required to complete these three steps. In humans, there are at least six different glycosylases that bind specifically to a target base and hydrolyze the N-glycosylic bond generating the AP or abasic site. Next, the AP site is processed by the APE1 system (AP Endonuclease-1, or HAP1/REF1/APEX), which cuts the phosphodiester backbone adjacent to the 5' end of the AP site, resulting in a 3' hydroxyl group and a transient 5' dRP (abasic deoxyribose phosphate). The removal of the dRP is accomplished by DNA Pol Beta (polymerase beta) activity, which adds one nucleotide to the 3' end of the nick and removes the dRP moiety through the action of an AP lyase (Bennett, et al., 1997). DNA Pol Beta also interacts with XRCC1. DNA Pol Beta is therefore crucial for the inclusion of different components of BER at sites of DNA damage and promoting repair efficiency (Fig. 6). The BER pathway deals with smaller damage to individual bases, such as oxidation, methylation, depurination, and deamination. If the adducts are left unrepaired, they may cause permanent mutations (Boysen & Hecht, 2003). If these mutations are situated at critical sites, including tumor suppressor genes, DNA repair-related genes or oncogenes, they may lead to cellular transformation and the development of tumors. A recent study demonstrates that BER plays an important role in DNA repair in cells exposed to PAHs. Chinese hamster ovary cells (CHO) deficient in the BER pathway were found to be more

importance of NER pathway in DNA damage induced by PAHs.

**4.2 Base excision repair (BER)** 

mediated (Santodonato, 1997). Nevertheless, the most important mechanism of carcinogenesis is a deficient DNA repair system in key genes involved in cell cycle control. Since chronic exposure to PAHs is related to a high rate of mutagenesis it is probable that damage to DNA is cumulative. Several studies have associated chronic occupational PAH exposure to multiple types of cancer including cancer of the bladder, lung, kidney, liver, and breast (Boffetta, et al., 1997, Dickey, et al., 1997, Karami, et al., 2011, Shen, et al., 2003).

Fig. 5. Structures and interaction of two common adducts: (A) BPDE-dG adduct and (B) BPDE-dA adduct.
