Sequential combined tumorigenic effect of HPV-16 and chemical carcinogens

Abstract

We immortalized oral keratinocytes by transfection with recombinant human papillomavirus type 16 (HPV-16) DNA and established two cell lines, human oral keratinocytes-16A (HOK-16A) and -16B (HOK-16B). These cell lines were morphologically different from the normal counterpart, contained HPV-16 DNA as integrated form and expressed numerous viral genes. However, these cells proliferated only in culture medium containing low calcium (0.15 mM) and are not tumorigenic in nude mice. To test the hypothesis that tumors can be developed by sequential combined effect of human papillomavirus and chemical carcinogens in the oral cavity, these immortalized cell lines were chemically transformed by exposure to either benzo[a]pyrene or methanesulfonic acid ethyl ester. Such transformants proliferated in medium containing physiological calcium levels (1.5 mM) and demonstrated enhanced growth potential in nude mice, whereas primary human oral keratinocytes treated with these chemical carcinogens failed to show any evidence of transformation. Chemically transformed cells contained integrated, intact HPV-16 sequences and transcribed significantly higher amount of HPV-16 E6/E7 messages and transforming growth factor-alpha (TGF-alpha) compared with the immortalized oral keratinocytes. Like the HPV-immortalized cell lines, the chemically transformed oral keratinocytes contained lower levels of newly synthesized, wild-type p53 proteins compared to normal cells, and expressed wild-type c-Ha-ras. These results indicate that this in vitro system is useful for investigating the mechanisms of multistep oral carcinogenesis.

Identification of biomarkers that distinguish human papillomavirus (HPV)-positive versus HPV-negative head and neck cancers in a mouse model

“…Transformation studies using oral keratinocytes argue for a synergy between the viral oncogenes and tobacco carcinogens. Even though HPV16 E6 and E7 are sufficient to immortalize human oral keratinocytes, exposure to tobacco carcinogens is required for these cells to become tumorigenic in nude mice (37,38). Synergy between a carcinogen and a papillomavirus to induce tumors in the esophagus of animals has also been shown with bovine papillomavirus 4-infected cattle that were fed a diet of bracken fern, which contains the flavonoid carcinogen quercetin (39).…”

Downregulation of Cdc2/CDK1 Kinase Activity Induces the Synthesis of Noninfectious Human Papillomavirus Type 31b Virions in Organotypic Tissues Exposed to Benzo[ a ]pyrene

“…Epidemiological studies have established a causal relationship between cigarette smoking and the development of human papillomavirus (HPV)-associated cervical cancer (10,43,55), with the risk of developing cancer doubling among HPV-infected women who are smokers compared with those who never smoked (79). Polycyclic aromatic hydrocarbons (PAHs), such as benzo-[a]pyrene (BaP), and tobacco-specific nitrosamines, such as 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and their respective metabolites, which are major carcinogenic constituents in cigarette smoke, have been shown to be present in the cervical mucus of smokers (47,63).…”

The Cigarette Smoke Carcinogen Benzo[ a ]pyrene Enhances Human Papillomavirus Synthesis

“…Epidemiological studies suggest that environmental and host-related cofactors act in conjunction with HPV to promote malignant progression of cervical lesions (7). It has been proposed that cigarette smoking among HPV-positive women is one of the cofactors which likely influences the risk for cervical cancer progression (15,21,27,38,41,50,52,54). Tobacco-specific polycyclic aromatic hydrocarbons such as benzo[a]pyrene (BaP) (31) and nitrosamines such as 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) (43), which are potent inducers of carcinogenesis (12,16,28), have been detected in the cervical mucus of women who smoke and exhibit cervical dysplasias (19,29,47).…”