Role of ribonucleotide reductase in expression of the neoplastic program

Evidence is presented for the tight linkage of ribonucleotide reductase activity with normal and neoplastic proliferation. A sensitive and reproducible assay was worked out to measure CDP reductase activity in rat in normal liver and various tissues, hepatomas of different growth rates, kidney tumors and sarcoma and tissue culture cells of hepatoma 3924A. In the standard assay, linear kinetics were obtained and the reductase activity of the rat liver was 23 ± 3 pmol CDP metabolized per hr/mg protein. When hepatoma 3924A tissue culture cells that had accumulated in plateau phase were replated, allowed to go through lag and log phases and again into the plateau phase during a 96-hr period, ribonucleotide reductase activity rose at 6 hr after cells were plated, the activity was maintained at high levels during the first 48-hr period, and returned to the resting level at 72 and 96 hr. This rise was earlier than that of 6 other enzymes of pyrimidine $\text{de novo}$ and salvage pathways (thymidine kinase, CTP synthetase, orotidine-5′-phosphate decarboxylase, orotate phosphoribosyltransferase, uridine phosphoribosyltransferase, and uridine-cytidine kinase). The rise in reductase activity was synchronous with the increase in incorporation of cytidine and deoxycytidine in the hepatoma cells. The reductase activity was markedly elevated in kidney tumors (31-fold) and in sarcoma (60-fold) as compared to the kidney cortex and muscle, respectively. In 14 lines of transplantable solid hepatomas, reductase activity was increased from 6.2- to 326-fold of that of normal rat liver. The rise in reductase activity positively correlated with the growth rate of the hepatomas; the behavior of CDP reductase was both transformation- and progression-linked. Reductase activity was also high in differentiating and regenerating liver; thus, it also was linked with normal proliferation. However, the elevation in activity was more marked in the rapidly-growing solid hepatoma 3924A (97-fold) than in normal tissues with the same replicative rate, such as regenerating (56-fold) or differentiating (46-fold) liver. Reductase activity was also high in organs of active cell renewal (thymus, bone marrow, spleen and intestine). Since in the solid hepatomas the levels of the substrate for the reductase, the ribonucleoside diphosphates, were generally unaltered, the marked elevation observed in the concentration of deoxynucleoside triphosphates may be attributed primarily to the early and marked rise in CDP reductase activity.