Improvement of the therapeutic efficacy of fluoropyrimidines in breast cancer is the main goal of this application. 5-FU is one of the most active drugs used in combination chemotherapy of metastatic breast and GI carcinoma. To increase the limited therapeutic index of FU, newly found differences between tumor and normal tissue in the disposition of uridine will be exploited. Specifically, differences in the Na+-dependent uridine transporter, which may be deficient in human tumors, and uridine phosphorylase, for which a BAU-insensitive activity may be selectively found in tumors, will be pursued. Both of these actions would tend to lead to lower levels of uridine in tumor vs normal cells; since uridine has been shown to protect against FU cytotoxicity, under the appropriate conditions a selective protection of normal tissue could be achieved without compromising the antitumor effect of FU. Were this to occur, the dose of FU could then be safely escalated. To evaluate differences in the uridine transporter between normal and neoplastic tissues, the applicant will measure expression of the transporter using a cDNA probe for the rabbit kidney Na+-dependent uridine transporter and another cDNA probe for a nucleoside transporter from rat intestinal epithelium; he will also attempt to clone and characterize the cDNA for the human gene from a human kidney cDNA library. He will generate antibodies against the transporter protein to probe surgical specimens for loss of this concentrative mechanism for uridine. To further pursue his observation that as much as 50% of uridine phosphorylase activity in selected human tumors is not sensitive to BAU, a potent inhibitor of uridine phosphorylase, the applicant will purify the non-BAU sensitive form of uridine phosphorylase; kinetically characterize differences in the catalytic activity of the enzymes from normal and neoplastic human tissue specimens; complete the cloning of the cDNA for the enzyme and prepare antibodies to the proteins; assess expression in human tumors and normal tissues; and initiate structural characterization of the enzyme by defining the binding domain for BAU utilizing photoaffinity or covalent labeling probes.
In specific aim 3, the applicant will continue his preclinical and clinical evaluation of BAU in a human breast cancer xenograft model and in a Phase I-II trial to determine if the MTD of FU can be raised by a BAU regimen that has been demonstrated to cause target changes in plasma uridine levels.
