This proposal is aimed at providing information basic to the development of improved drugs for cancer chemotherapy. Two approaches will be taken. In the first, the molecular basis of specificity of pteridines and related compounds will be determined so that anti-folates may be designed which are specific for dihydrofolate reductase and which do not interfere with other pteridine utilizing enzymes. Series of substituted pteridines, pyrimidines and pyrimidodiazepines will be studied for their ability to bind to the aromatic amino acid hydroxylases. These enzymes have a specific requirement for a pteridine cofactor, tetrahydrobiopterin (BH4), and are involved in the biosynthesis of serotonin and the catecholamine neurotransmitters. The effects of the pteridines and pyrimidines will also be assessed on dihydropteridine reductase, the enzyme which in vivo regenerates BH4. Any pteridine like drug aimed at dihydrofolate reductase, including pyrimidines and pyrimidodiazepines, has the potential to restrict BH4 dependent neurotransmitter synthesis. Binding studies to be conducted both in vitro and in vivo, may also yield information leading to an understanding of the elevated urinary neopterin levels observed in patients with malignant disease, and found to correlate with the clinical status of the patient. The only known source of neopterin in mammalian metabolism is from the degradation of neopterin triphosphate, and intermediate in the biosynthesis of BH4 from GTP. The second approach is to develop mechanism based active-site directed irreversible inhibitors of thymidylate synthase, another target enzyme in cancer chemotherapy. Analogs of the cofactor 5-10-methylene-tetrahydrofolate will be synthesized which have a second substituent at the 6-position of the pterin ring. Since thymidylate synthase specifically requires the 6-hydrogen, blocking of this position, with e.g. methyl, will prevent the reaction from proceeding. If 6,6-disubstituted compounds bind such that a reaction is initiated, they should act as suicide substrates, and become irreversibly bound at the active site. These compounds will also provide a means of further probing the enzyme mechanism. Although it is known that the hydrogen from the 6-position of 5-10-methylene-tetrahydrofolate is incorporated into the methyl group of TMP, the route by which it arrives there is still unknown.
| Bailey, S W; Dillard, S B; Thomas, K B et al. (1989) Changes in the cofactor binding domain of bovine striatal tyrosine hydroxylase at physiological pH upon cAMP-dependent phosphorylation mapped with tetrahydrobiopterin analogues. Biochemistry 28:494-504 |
| Pike, D C; Hora, M T; Bailey, S W et al. (1986) Pyrimidodiazepine, a ring-strained cofactor for phenylalanine hydroxylase. Biochemistry 25:4762-71 |
