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Purine and Pyrimidine Enzymic Programs and Nucleotide Pattern in Sarcoma¹

Laboratory for Experimental Oncology, Indiana University School of Medicine, Indianapolis, Indiana 46223 [G. W., R. C. J., N. P., M. S. L., E. T.], and the Surgery Branch, National Cancer Institute, NIH, Bethesda, Maryland 20205 [M. E. B.]

The purpose of this study was to elucidate the enzymic program and nucleotide pattern of a chemically induced, transplantable sarcoma and to compare the biochemical makeup with that of normal and differentiating skeletal muscle in the rat.

The activities of 28 key enzymes of pyrimidine, purine, and carbohydrate metabolism were determined in the 100,000 x g supernatant fluid or in purified extracts. The concentrations of the adenosine, guanosine, uridine, and cytidine mono-, di-, and triphosphates were measured by high-pressure liquid chromatography of samples prepared by the freeze-clamp method.

The results of enzymic and metabolite assays were given in nmol/hr/mg protein and nmol/g, respectively, and for comparability were also expressed as percentages of the muscle of adult rat as the reference normal tissue in this study. These percentages denote that the activities in sarcoma or in differentiating muscle were higher or lower than those in the muscle of adult rats.

In pyrimidine metabolism, the specific activities of cytidine diphosphate reductase, cytidine triphosphate synthetase, and thymidine kinase increased in the sarcoma 60-, 78-, and 80-fold over those of the muscle. The activities of the key glycolytic enzymes, hexokinase and phosphofructokinase, increased 7- and 3-fold, whereas that of pyruvate kinase decreased to 35%. The activities of glucose-6-phosphatase and fructose-1,6-diphosphatase declined to 42 and 48%, respectively. The activities of enzymes involved in pentose phosphate production and utilization increased, with that of the glucose-6-phosphate dehydrogenase being elevated 288-fold. The activity of galactokinase was unchanged, whereas that of uridine diphosphoglucose pyrophosphorylase decreased to 22%. In purine metabolism, the activities of the first three enzymes of guanosine triphosphate biosynthesis, inosine monophosphate dehydrogenase, guanosine monophosphate synthetase, and guanosine monophosphate kinase, increased 22-, 2-, and 5-fold, respectively. In contrast, the activities of adenylosuccinate lyase and adenosine monophosphate deaminase decreased to 28 and 42%. The activities of adenosine deaminase and kinase increased 1.8- and 3.5-fold. The activity of the rate-limiting enzyme of de novo inosine monophosphate biosynthesis, amidotransferase, increased 13-fold, whereas that of the rate-limiting purine-catabolic enzyme, xanthine oxidase, decreased to 54%.

In the sarcoma, the concentrations of adenosine tri-, di-, and monophosphate decreased to 24, 49, and 76%. In contrast, the pools of guanosine tri-, di-, and monophosphate increased 9- to 11-fold, and those of the uridylates were also elevated 4.5- to 12-fold. The cytidine triphosphate concentration increased 17-fold. In the muscle, the concentrations of cytidine diphosphate, inosine monophosphate, and xanthosine monophosphate were too low for the sensitivity of our method to detect, but these pools were well measurable in the sarcoma.

The enzymic program of sarcoma was distinguished from that of the muscle of 6-day-old rats by the quantitatively more pronounced alterations in the sarcoma. The activities of uridine phosphorylase, hexokinase, 6-phosphogluconate dehydrogenase, and adenosine deaminase were higher in sarcoma but were lower or not significantly altered in 6-day-old muscle as compared to activities of adult muscle. The activities of adenosine monophosphate deaminase and xanthine oxidase decreased in sarcoma, whereas they were unchanged or increased, respectively, in 6-day-old muscle.

In the sarcoma, important segments of alterations in enzymology of carbohydrate, purine, and pyrimidine metabolism were identified that proved to be a program shared with that observed in chemically induced and virus-derived transplantable rat and avian hepatocellular carcinomas and in primary human liver, kidney, and colon neoplasms. There were also sarcoma-specific markers in the enzymic programs and nucleotide patterns that readily distinguish this tumor from other examined neoplasms.

The present investigation revealed a formidable biochemical capacity for replication in the sarcoma cells. These biochemical studies also point out possible targets in the strategy of anticancer drug treatment of sarcoma.

¹ This investigation was supported by USPHS Grants CA-13526, CA-05034, and CA-10792.

² To whom requests for reprints should be addressed.

Received 7/22/82. Accepted 12/ 3/82.