Aflatoxin epoxidation

Figure 24.12. Examples of DNA adducts and oxidized guanine nucleotide. (A) Benzo[a]pyrene-7,8-diol-9,10-epoxide-M-2 guanine adduct. (B) Aflatoxin epoxide N7 guanine adduct. (C) 8-Oxoguanine is commonly produced by reactive oxygen species, dR = deoxyribose.

A specific mutation in the p53 tumour suppressor gene has been detected in 10-70% of HCCs from areas of high AFBi exposure and is absent from HCCs from areas with negligible AFBi exposure [28-30]. Support for the implication that the mutation, a G- T transversion at the third base of codon 249, is caused by exposure to aflatoxin has come from in vitro studies. Aflatoxin exposure in bacteria almost exclusively causes G—transversions [31] and the aflatoxin-epoxide has been shown to bind to codon 249 of p53 in vitro [32]. Moreover, human... 

DNA alkylation has the potential to yield a time-dependent spectrum of adducts, in which initially formed kinetically favored lesions give way to the thermodynamically favored adducts over time. Reversible alkylation has been observed at several of the nucleophilic sites in DNA, including N3A (CC-1065,7, Scheme 8.10, duocarmycin, 8)," " NIA (qui-none methide, 9)," N7G (leinamycin, Schane 8.11, aflatoxin Bj epoxide, 10 and quinone methide, 9),57.ii4.ii8 (quinone methide, 9)," and bPG (ecteinascidin 743,11)." The bidentate Nl/ISPG adduct of malondialdehyde also forms reversibly. ... 

The normal pathway toward excretion of foreign molecules like aflatoxin Bi and dibenzo [a,/]pyrene, however, also involves nucleophilic substitution reactions of their epoxides. 

The mode of action of this carcinogen is believed to involve epoxidation of a double bond (49), as indicated in Figure 3 in this Figure the similarities of the shapes, and particularly of the sites of activation of aflatoxin, BP and DMBA are demonstrated. Aflatoxin has functional groups at each end of the molecule, unlike an activated PAH which has functional groups only at one end of the molecule. 

Figure 3. (a) Similarities in the shapes of BP, DMBA, a base pair of guanine and cytosine, and the steroid, estradiol. In (b) the shapes of BD and DMBA (filled bonds) are compared with that of aflatoxin B] (open bonds) and the sites of activation of each to an epoxide is indicated by the arrows. 

The potent carcinogen aflatoxin B1 is activated by generation of an epoxide on a dihydrofuran moiety [43]. The potent MBIs contained in grapefruit juice and in the herbal medication St J ohn s Wort are furanocoumarin compounds [44—46]. A similar moiety is also incorporated in the HIV protease inhibitor L-754,394, a potent CYP3A4 MBI [47]. 

Fig. 10.30. Structure of aflatoxin B1 exo-8,9-epoxide (10.132), the dihydrodiol resulting from hydrolysis (10.133), and the reactive a-hydroxy dialdehyde (10.134) that exists in equilibrium with the diol under alkaline conditions [204]...

In the pH range of 5 - 10, H20-catalyzed hydrolysis is the predominant mechanism (see Fig. 10.11, Pathway b), resulting in the formation of the (8R,9R)-dihydrodiol (10.133, Fig. 10.30). Thus, aflatoxin B1 exo-8,9-epoxide is possibly the most reactive oxirane of biological relevance. Such an extreme reactivity is mostly due to the electronic influence of 0(7), as also influenced by stereolectronic factors, i.e., the difference between the exo- and endo-epoxides. The structural and mechanistic analogies with the dihydro-diol epoxides of polycyclic aromatic hydrocarbons (Sect. 10.4.4) are worth noting. 

A further remarkable finding in the hydrolysis of aflatoxin B1 exo-8,9-epoxide is the relative instability of the dihydrodiol, which under basic conditions exists in equilibrium with an aflatoxin dialdehyde, more precisely a furofuran-ring-opened oxy anionic a-hydroxy dialdehyde (10.134, Fig. 10.30). The dihydrodiol is the predominant or exclusive species at pH < 7, whereas this is true for the dialdehyde at pH >9, the pK value of the equilibrium being 8.2 [204], The dialdehyde is known to form Schiff bases with primary amino groups leading to protein adducts. However, the slow rate of dialdehyde formation at physiological pH and its reduction by rat and human aldo-keto reductases cast doubts on the toxicological relevance of this pathway [206]. 

The exceptional reactivity of aflatoxin B1 exo-8,9-epoxide raises the question of its potential detoxification by EHs. Despite the short half-life, the epoxide does react with DNA (toxification) and glutathione 5-transferases (detoxification), but a role for EH appeared dubious [207], Rat liver or recombinant rat EH has since been shown to provide a modest enhancement of up to 22% in the hydrolysis rate of aflatoxin B1 exo-S,9-epoxide, and to decrease somewhat the genotoxicity of aflatoxin B1 when the ratio of EH to cytochrome P450 is high (ca. 50-fold). Purified human EH provided no such enhancement in hydrolysis, nor did it have a clear effect on genotoxicity. Thus, little evidence exists to support a role for EH in the detoxification of aflatoxin B1 [208],... 

F. P. Guengerich, W. W. Johnson, Y. F. Ueng, H. Yamazaki, T. Shimada, Involvement of Cytochrome P450, Glutathione 5-Transferase, and Epoxide Hydrolase in the Metabolism of Aflatoxin B1 and Relevance to Risk of Human Liver Cancer , Environ. Health Perspect. 1996, 104 (Suppl. 3), 557 - 562. 

Denissenko, M.F. et al., Quantitation and mapping of aflatoxin DNA damage in genomic DNA using aflatoxin Bi-8,9-epoxide and microsomal activation systems, Mutat. Res., 425, 205, 1999. 

Fortunately, nature provides an alternative nucleophile whose role is to mop up dangerous electrophiles such as aflatoxin Bi epoxide before they can do damage, and to remove them from the body. This compound is glutathione (see Box 6.6), a tripeptide composed of glutamic acid, cysteine, and glycine. 

Electron-rich alkenes are the more reactive jr-bond snbstrates towards epoxidation by the electrophilic dioxiranes Some typical examples of these oxidations are snm-marized in Scheme 2. Since the resnlting epoxides are nsnally hydrolytically and ther-molytically qnite labile, snch oxidations are best carried ont with isolated dioxiranes. For example, the 8,9 epoxide of the well-known aflatoxin B, postnlated as potent carcinogen in the oxidative metabolism of this natural product, escaped numerous efforts to prepare it by conventional epoxidations because of its sensitivity towards hydrolysis . The synthesis of this labile epoxide was readily accomplished by employing a solution of the isolated DMD at room temperature (equation 2), and its mutagenicity unequivocally... 

Figure 5.14 Routes of oxidative metabolism of aflatoxin showing activation to an epoxide catalyzed by CYP1A2 and CYP3A4 and conjugation with glutathione catalyzed by a-GST. The 8,9-exoepoxide is the enantiomer, which binds extensively to DNA and is carcinogenic. The endoepoxide binds less readily. Abbreviation. a-GST, a-glutathione transferase.

Epoxidation/hydroxylation Aldrin, benzo(a)pyrene, aflatoxin, bromobenzene... 

Aliphatic Epoxidation. Many aliphatic and alicylcic compounds containing unsaturated carbon atoms are thought to be metabolized to epoxide intermediates (Figure 7.4). In the case of aldrin the product, dieldrin, is an extremely stable epoxide and represents the principle residue found in animals exposed to aldrin. Epoxide formation in the case of aflatoxin is believed to be the final step in formation of the ultimate carcinogenic species and is, therefore, an activation reaction. 

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