This project is composed of two major areas of research: (1) studies of pyrimidine nucleotide synthesis in vivo and (2) cellular control of nucleoside diphosphate hexose synthesis and glycosaminoglycan production. The relative contribution of de novo and salvage synthesis to tissue pyrimidine nucleotide pools is an important parameter for the rational design of antipyrimidine therapies. Both pathways were measured in the intact mouse using stable isotopes to quantitate de novo activity and radiolabeled uridine to quantitate salvage. Low doses of PALA cause a complete inhibition of hepatic de novo synthesis for at least 48 hours; whereas, inhibition of intestinal de novo synthesis is short lived with high doses of PALA and an actual stimulation of de novo pathway activity takes place at low PALA doses. These data support the concept of hepatic export of salvageable uridine as one determinant of the selective toxicity of antipyrimidines; a concept important to the design of clinical protocols. Data from our Laboratory establish a link between mitogenic activation of quiescent fibroblasts and the synthesis of UDP-hexoses used in extracellular matrix formation. An important factor in the synthesis of hyaluronan, a glycosaminoglycan associated with invasion and metastasis, is UDP-glucose dehydrogenase (UDPGDH). Using an HPLC method developed to quantitate UDPGDH activity in crude homogenates, we found activity to be directly related to cell proliferation and the cellular concentration of UDP-glucuronic acid. The early activation of UDPGDH is independent of protein synthesis; UDPGDH is a likely link between signal transduction pathways and hyaluronan synthesis. Metabolic compartmentation studies indicate that the glucuronic acid moiety of UDP-glucuronic acid is derived from exogenous glucose and not from glycogen stores. Hyaluronan synthesis can be inhibited by Suramin and by agents that are glucuronidated.
