As studies on the reactions of carcinogens with cellular macromolecules progressed, it became
apparent that most of these interactions resulted from covalent bond formation between an electrophilic
form of the carcinogen and the nucleophilic sites in proteins (e.g., sulfur, oxygen, and
nitrogen atoms in cysteine, tyrosine, and histidine, respectively) and nucleic acids (e.g., purine
or pyrimidine ring nitrogens and oxygens). Frequently, the parent compound itself did not
interact in vitro with macromolecules until it had been incubated with liver homogenates or
liver microsomal fractions. These studies led to the realization that metabolic activation of certain
carcinogenic agents is necessary to produce the ‘‘ultimate carcinogen’’ that actually reacts
with crucial molecules in target cells. With the exception of the very chemically reactive alkylating
agents, which are activated in aqueous solution at physiologic pH (e.g., N-methyl-Nnitrosourea),
and the agents that intercalate into the DNA double helix by forming tight noncovalent
bonds (e.g., daunorubicin), most of the known chemical carcinogens undergo some
metabolic conversions that appear to be required for their carcinogenic action. Some examples
of these metabolic conversions are given next.
Donors of Simple Alkyl Groups
Included in this group are the dialkylnitrosamines,dialkylhydrazines, aryldialkyltriasenes,
alkylnitrosamides, and alkylnitrosimides. The alkylnitrosamides and alkylnitrosimides do not
require enzymatic activation because they can react directly with water or cellular nucleophilic
groups. The alkylnitrosamines, alkylhydrazines, and alkyltriazenes,however, undergo anenzymemediated
activation step to form the reactive electrophile (Fig. 2–6). These agents are metabolically
dealkylated by the mixed-function oxidase system in the microsomal fraction (endoplasmic
reticulum) of cells, primarily liver cells. The monoalkyl derivatives then undergo a nonenzymatic,
spontaneous conversion to monoalkyldiazonium ions that donate an alkyl to cellular
nucleophilic groups in DNA, RNA, and protein.8
Cytochrome P-450–Mediated
Activation
A number of carcinogenic chemicals are chemically inert nucleophilic agents until they are converted
to active nucleophiles by the cytochrome p-450–dependent mixed function oxidases, or
CYPs So far, 57 genes encoding these enzymes have been identified in the human genome. The
CYPs most involved in carcinogen activation are CYP1A1, 1A2, 1B1, 2A6, and 3A4.A wide variety
of chemical carcinogens such as aromatic and heterocyclic amines, aminoazo dyes, polycyclic
aromatic hydrocarbons, N-nitrosamines, and halogenated olefins are activated by one or more of these CYPs (Fig. 2–7). Some of these compounds are further activated by subsequent steps; for
example, 2-acetylaminofluorene (AAF) is further modified by a sulfotransferase to form the ultimate
DNA-binding moiety. Somewhat surprisingly, glutathione-S-trans ferase (GST), which had been thought to be involved only in detoxifying carcinogens, has been shown to activate some industrial chemicals,
7 so GST appears to have a dual role, depending on the chemical.
2-Acetylaminofluorene
Themetabolic interconversions of this compound were studied in detail by the Millers and colleagues.
9,10 In 1960, itwas shown that AAF is converted to a more potent carcinogen, N-hydroxy-
AAF, after the parent compound was fed to rats. Although both AAF and N-hydroxyl-AAF are
carcinogenic in vivo, neither compound reacted in vitro with nucleic acids or proteins, suggesting
that the ultimate carcinogen was another, as-yet unidentified metabolite. Subsequent studies
showed that N-hydroxy-AAF is converted in rat liver to a sulfate, N-sulfonoxy-AAF, by means
of a cytosol sulfotransferase activity .This compound reacts with nucleic acids and
proteins and appears to be the ultimate carcinogen in vivo. It is also highly mutagenic, as determined
by assays of DNA-transforming activity .
Other enzymatic conversions of AAF occur in rat liver, for example,N-hydroxy-AAFisconverted
to N-acetoxy-AAF, N-acetoxy-2-aminofluorene and theO-glucuronide (conjugate with glucoronic
acid). These enzymatic reactions may also be involved in the conversion of AAF to carcinogenic
metabolites,especiallyinnonhepatictissues,which often have low sulfotransferase activity for Nhydroxy-
AAF. The acetyltransferase-mediated activity converts N-hydroxy-AAF to N-acetoxy-2-
aminofluorene, which is also a strong electrophile andmay be the ultimate carcinogen in nonhepatic
tissues.
