Tumor Initiation

The early concept of tumor initiation indicated that the initial changes in chemical carcinogenesis are irreversible genetic damage. However, recent data from molecular studies of preneoplastic human lung and colon tissues implicate epigenetic changes as an early event in carcinogenesis. DNA methylation of promoter regions of genes can transcriptionally silence tumor-suppressor genes.²¹ For mutations to accumulate, they must arise in cells that proliferate and survive the lifetime of the organism. A chemical carcinogen causes a genetic error by modifying the molecular structure of DNA that can lead to a mutation during DNA synthesis. Most often, this is brought about by forming an adduct between the chemical carcinogen or one of its functional groups and a nucleotide in DNA.¹⁶ (The process by which this occurs for the major classes of chemical carcinogens is discussed in detail under “Carcinogen Metabolism”). In general, a positive correlation is found between the amount of carcinogen-DNA adducts that can be detected in animal models and the number of tumors that develop.^25–27 Thus, tumors rarely develop in tissues that do not form carcinogen-DNA adducts. Carcinogen-DNA adduct formation is central to theories of chemical carcinogenesis, and it may be a necessary, but not a sufficient, prerequisite for tumor initiation. DNA adduct formation that causes either the activation of a protooncogene or the inactivation of a tumor-suppressor gene can be categorized as a tumor-initiating event (see “Tumor Progression,” “Oncogenes,” and “Tumor-Suppressor Genes”).

Tumor Promotion

Tumor promotion comprises the selective clonal expansion of initiated cells. Because the accumulation rate of mutations is proportional to the rate of cell division, or at least the rate at which stem cells are replaced, clonal expansion of initiated cells thus, produces a larger population of cells that are at risk of further genetic changes and malignant conversion.^19,28 Tumor promoters are generally nonmutagenic, are not carcinogenic alone, and often (but not always) are able to mediate their biologic effects without metabolic activation. These agents are characterized by their ability to reduce the latency period for tumor formation after exposure of a tissue to a tumor initiator, or to increase the number of tumors formed in that tissue. In addition, they induce tumor formation in conjunction with a dose of an initiator that is too low to be carcinogenic alone. Croton oil (isolated from Croton tiglium seeds) is used widely as a tumor promoter in murine skin carcinogenesis, and the mechanism of action for its most potent constituent, 12-otetradecanoylphorbol-13-acetate, via protein kinase C activation, is arguably the best understood among tumor promoters.²⁷ Chemicals or agents capable of both tumor initiation and promotion are known as complete carcinogens, eg, benzo[a]pyrene and 4-aminobiphenyl.

Identification of new tumor promoters in animal models has accelerated with the sophisticated development of model systems designed to assay for tumor promotion. Furthermore, ligand-binding properties can be determined in recombinant protein kinase C isozymes that are expressed in cell cultures.²⁷ Chemicals, complex mixtures of chemicals, or other agents that have been shown to have tumor-promoting properties include dioxin, benzoyl peroxide, macrocyclic lactones, bromomethylbenzanthracene, anthralin, phenol, saccharin, tryptophan, dichlo- rodiphenyltrichloroethane (DDT), phenobarbital, cigarette-smoke condensate, polychlorinated biphenyls (PCBs), teleocidins, cyclamates, estrogens and other hormones, bile acids, ultraviolet light, wounding, abrasion, and other chronic irritation (ie, saline lavage).¹⁶ In addition, protein kinase C is activated and cellular diacylglycerol elevated in laboratory animals maintained on high-fat diets.^29,30

Malignant Conversion

Malignant conversion is the transformation of a preneoplastic cell into one that expresses the malignant phenotype. This process requires further genetic changes. The total dose of a tumor promoter is less significant than frequently repeated administrations, and if the tumor promoter is discontinued before malignant conversion has occurred, premalignant or benign lesions may regress. Tumor promotion contributes to the process of carcinogenesis by the expansion of a population of initiated cells that will then be at risk for malignant conversion. Conversion of a fraction of these cells to malignancy will be accelerated in proportion to the rate of cell division and the quantity of dividing cells in the benign tumor or preneoplastic lesion. In part, these further genetic changes may result from infidelity of DNA synthesis.³¹ The relatively low probability of malignant conversion can be increased substantially by the exposure of preneoplastic cells to DNA-damaging agents,¹⁶ and this process may be mediated through the activation of protooncogenes and inactivation of tumor suppressor genes.

Tumor Progression

Tumor progression comprises the expression of the malignant phenotype and the tendency of malignant cells to acquire more aggressive characteristics over time. Also, metastasis may involve the ability of tumor cells to secrete proteases that allow invasion beyond the immediate primary tumor location. A prominent characteristic of the malignant phenotype is the propensity for genomic instability and uncontrolled growth.³² During this process, further genetic and epigenetic changes can occur, again including the activation of protooncogenes and the functional loss of tumor suppressor genes. Frequently, protooncogenes are activated by two major mechanisms: in the case of the ras gene family, point mutations are found in highly specific regions of the gene (ie, the twelfth, thirteenth, fifty-ninth, or sixty-first codons), and members of the myc, raf, HER2, and jun multigene families can be overexpressed, sometimes involving amplification of chromosomal segments containing these genes. Some genes are overexpressed if they are translocated and become juxtaposed to a powerful promoter (eg, the relationship of bcl-2 and immunoglobulin heavy chain gene promoter regions in B-cell malignancies; see also Philadelphia chromosome on page 8, under Clonal Evolution). Loss of function of tumor-suppressor genes usually occurs in a bimodal fashion, and most frequently involves point mutations in one allele and loss of the second allele by a deletion, recombinational event, or chromosomal nondisjunction. These phenomena confer to the cells a growth advantage as well as the capacity for regional invasion, and ultimately, distant metas-tatic spread. Despite evidence for an apparent scheduling of certain mutational events, it is the accumulation of these mutations, and not the order or the stage of tumorigenesis in which they occur, that appears to be the determining factor.^23,24