STUDIES ON LIVER-TUMOR PROMOTION IN THE RAT BY OROTIC-ACID - DOSE ANDMINIMUM EXPOSURE TIME REQUIRED FOR DIETARY OROTIC-ACID TO PROMOTE HEPATOCARCINOGENESIS

Our earlier studies indicated that orotic acid, a precursor for pyrimi dine nucleotide biosynthesis, exerts a promoting effect on rat hepatoc arcinogenesis. The present study was designed to determine the optimum conditions of exposure to orotic acid required for promotion of hepat ocarcinogenesis in the initiated rats. The first series of experiments was designed to determine the optimum dose of orotic acid needed to e xert its liver tumor promoting effect. Accordingly male Fischer rats w ere given diethylnitrosamine (200 mg/kg, i.p.) or 0.9% NaCl. One week later carcinogen-injected rats were divided into six groups and fed ei ther basal diet or the same diet containing 0.1, 0.5, 1, 2 or 4% oroti c acid. Rats given 0.9% NaCl were fed 4% orotic acid. Two-thirds parti al hepatectomy was performed on all animals 10 weeks after starting on their respective diets, and all groups were killed 3 weeks thereafter . Analysis of gamma-glutamyltransferase-positive foci and nodules reve aled that 0.5-1% orotic acid in the diet is sufficient to exert a sign ificant promoting effect on the selective growth of initiated hepatocy tes, while higher concentrations of orotic acid were only marginally m ore effective. No gamma-glutamyl-transferase-positive foci were observ ed in animals given 4% orotic acid diet following saline injection. Us ing 1% orotic acid as the promoting regimen, in the next series, the m inimum exposure time required for dietary orotic acid to promote liver carcinogenesis was determined. Male Fischer 344 rats were given i.p. either 1,2-dimethylhydrazine dihydrochloride (100 mg/kg) or 0.9% NaCl 18 h after 2/3 partial hepatectomy. After 1 week of recovery one group of rats was continued on a semisynthetic basal diet, while others wer e transferred to the same basal diet containing 1% orotic acid. Rats t hat were on the 1% orotic acid diet were progressively transferred to the basal diet after 5, 10, 20, 29 and 40 weeks of exposure. All rats were sacrificed 54 weeks after the beginning of the experiment. The re sults indicate that 100% of the initiated rats developed hepatic nodul es whether or not they were exposed to an orotic acid-containing diet. However, the incidence of hepatocellular carcinoma was greatly increa sed in animals exposed to the orotic acid diet, with 42% incidence in initiated rats given orotic acid diet for 10 weeks and up to 75% in th ose exposed to this diet for 40 weeks. Further, promotion by orotic ac id exhibited a high metastatic potential with 33-60% metastasis to the lungs. On the other hand, initiated rats exposed to the basal diet de veloped only a 21% incidence of hepatocellular carcinoma, with no meta stasis to the lungs. These results indicate that a minimum of 10-20 we eks of exposure to orotic acid diet was found to be sufficient to indu ce maximum liver tumor promoting effect. Further, initiated rats, whet her or not exposed to orotic acid, exhibited a higher incidence of ext ra-hepatic tumors. However, their significance cannot be assessed beca use of low frequency of incidence at any given site.