A pancreatic cancer cell line, PC-1, was established from a pancreatic ductal carcinoma induced in a hamster by N-nitrosobis(2-oxopropyl)amine (BOP). The cells grew in a monolayer with a doubling time of 38 h, and floated or piled up to form a duct-like structure. Chromosome counts ranged from 42 to 89. Light and electron microscopic studies of PC-1 cells revealed production of conspicuous amounts of amorphous substance. Injection of PC-1 cells into the homologous hamster pancreas resulted in tumor formation, histopathologically indistinguishable from the original primary pancreatic ductal carcinoma. Immunohistochemical expression of blood group-related antigens (BGRAs), A, B, H, Leb, Lex and Ley was observed both in the cells in the culture, and in tumor transplanted into the pancreas. In the culture supernatant, a high titer of blood group A antigen was detected. This cell line may provide a unique tool for studying the mechanism of BGRA synthesis and release in malignant cells.

Cytochrome P-450IA1 (Cyto-P450IA1) is the isozyme most closely associated with aryl hydrocarbon hydroxylase (AHH). At least two distinct high-affinity binding proteins may regulate its expression, the 4S protein that primarily binds polycyclic aromatic hydrocarbons (PAHs), and the 8S Ah receptor that binds 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and like congeners. The present study was conducted to investigate ligand binding characteristics of the 4S and 8S binding proteins before and after separation from liver cytosol in the presence and absence of sodium molybdate. Liver cytosol and 4S and 8S receptor-enriched fractions from livers of male Sprague-Dawley rats (AHH-responsive), and from C57BL/6N (AHH-responsive) and DBA/2N and AKR/N mice (AHH-nonresponsive) served as sources of these proteins. Competitive binding studies were performed using 10 nM [3H]benzo[a]pyrene (BaP) or [3H]-TCDD in the presence and absence of a 200-fold excess of BaP, 3-methylcholanthrene (3-MC), and tetrachlorodibenzofuran (TCDBF). Specific PAH-binding activity was assayed by using either sucrose density gradient analysis or a hydroxylapatite assay. Our results indicate that before and after the separation of liver cytosol into 4S and 8S fractions, ligand binding characteristics were relatively unaltered for the 4S protein in comparison to that for the Ah receptor, particularly in the presence of molybdate. The 4S protein had high affinity for BaP and 3-MC but very low affinity for TCDBF; the 8S protein had high affinity for TCDBF, lesser affinity for 3-MC, and low affinity for BaP. In the presence of sodium molybdate, the Ah receptor fractions were significantly stabilized, whereas the 4S protein was relatively unaffected. After the separation of Ah receptor fraction from liver cytosol in the presence of molybdate, 3-MC consistently bound to a greater extent. These results affirm the existence of two distinct PAH-binding proteins.

A protein which specifically binds [3H]benzo[a]pyrene and other polycyclic aromatic hydrocarbons has been purified over 6000-fold from rat hepatic cytosol by using ion-exchange, gel permeation, and hydrophobic interaction chromatography. The binding protein differs from the 9S binding protein characterized in other laboratories. A Stokes radius of 2.75 nm was determined by gel filtration on Sephadex G-100. A sedimentation coefficient of 3.3 S was determined by using sucrose gradient analysis. The ability of this protein to bind total rat liver DNA as well as subclones containing portions of the rat cytochrome P-450c gene was investigated. Under high stringency conditions, this binding protein was found to interact in a specific and saturable manner with several subclones of the rat cytochrome P-450c gene containing 5'-upstream sequences, as well as portions of intron 1. Binding was not observed to the coding portions of the gene. These data implicate the "4S" binding protein in the transregulation of rat cytochrome P-450c expression.

In the rat, expression of the CYP1A1 gene is closely associated with arylhydrocarbon hydroxylase (AHH) enzyme activity. AHH is an inducile enzyme activity known to play an important role in the bioactivation of polycyclic aromatic hydrocarbons (PAHs) to mutagenic and carcinogenic metabolites. PAH-induced expression of the CYP1A1 gene appears to be regulated by several trans-acting factors, including the Ah receptor and the 4S PAH-binding protein. In this study, we used the PAH isomers benzo[a]pyrene (BaP) and benzo[e]pyrene (BeP) to further evaluate the role of the 4S PAH-binding protein in induction of the CYP1A1 gene in H4-II-E rat hepatoma cells. Although BaP is believed to bind to both the Ah receptor and the 4S protein, BeP has been reported to bind exclusively to the 4S protein. The results of the study presented here indicate that BaP and BeP induce the expression of the CYP1A1 gene, as measured by ethoxyresorufin O-deethylase (EROD) activity, in a concentration-dependent manner. However, BaP is about 25 times as potent as BeP in inducing EROD activity in these cells. Slot-blot analysis of total RNA isolated from these cells indicated that BeP, BaP, and 3-methylcholanthrene increased the level of CYP1A1 mRNA expression. Sucrose-gradient analysis of BeP binding activity indicated that BeP bound with high affinity to the 4S PAH-binding protein, but not to the Ah receptor. These results suggest that the 4S protein may play a role in the PAH-induced expression of the CYP1A1 gene in rat H4-II-E cells.