The objective of this proposal is to understand the mechanisms by which environmental chemicals influence liver tumor promotion. To accomplish this task, we propose to use the prototype carcinogen, 2,3,7,8-tetrachlorodibenzo- ? p-dioxin (dioxin), as a promoter of hepatocellular carcinoma in the two-stage mouse model. The choice of dioxin is related to its biological potency and the large volume of genetic and pharmacological evidence that suggests its effects are mediated through a single ligand-activated transcription factor known as the Ah receptor (AHR). Given the central nature of this signaling protein, efforts will be made to elucidate the molecular details of the dioxin-AHR pathway as it relates to liver tumor promotion. To this end, we will first optimize the murine model of liver tumor promotion, with particular emphasis on genetics and statistical power. We will then use gene-targeting approaches to manipulate various functional domains and expression patterns of the AHR, as well as its heterodimeric partner ARNT. In particular, focus will be on determining if dioxin's tumor promoting activity is a cell autonomous process and if it is the direct result of the AHR's or ARNT's activity as transcription factors. This understanding will guide the later experiments that will be directed at identifying those particular transcriptional outputs that lie in the pathway to tumor promotion. In this last respect, emphasis will be placed on direct test for the involvement of candidate genes such as Cyp1a1, Cyp1a2 and Cyp1b1, as well as the IL1-like inflammatory cytokines. As a backup approach, we will begin to develop novel screening approaches to identify candidate genes using high throughput transcriptional profiling of preneoplastic lesions and SiRNA technologies to elucidate functional roles in promotion. ? ?
