This project is composed of two areas of research: (I) Cellular control of nucleoside diphosphate hexose synthesis and glycosaminoglycan production and (II) Regulation of pyrimidine nucleotide synthesis in vivo. Studies focused on the cellular regulation of hyaluronic acid synthesis, a glycosaminoglycan that is associated with tumor invasion and metastasis. Assays were developed for three enzymes involved in the synthesis of UDP-hexoses that are precursors for hyaluronic acid (UDP-glucose dehydrogenase, glucosamine-6-phosphate synthetase, and UDP-glucose synthetase). Current studies are evaluating these three enzymes, along with hyaluronic acid synthetase, as control points for regulating hyaluronic acid synthesis and as possible targets for drug design. Disruption of tumor-stromal interactions was achieved with short-chained oligosaccharides derived from hyaluronic acid by controlled hyaluronidase digestion. Oligosaccharides of different compositions were obtained by varying the hyaluronidase source. Oligosaccharides were separated by size using gel exclusion chromatography; purity was determined by DIONEX anion-exchange chromatography; and molecular weights by GC/MS. Current studies will determine the effect of size and composition of hyaluronic-acid derived oligosaccharides on the invasion/metastasis of B16F10 melanoma cells in mice and on tumor cell migration in vitro. An ongoing study that compares rates of pyrimidine synthesis in tumors and host tissues of intact mice and the differential effects of inhibitors of pyrimidine synthesis using stable isotopes and GC/MS analysis was continued to include doses of PALA that are used clinically as a bio-modulator of FUra activity.
