Phase I and pharmacokinetic studies of high-dose uridine intended for rescue from 5-fluorouracil toxicity.

The clinical effects and pharmacokinetics of high-dose uridine were determined in seven patients with advanced-stage cancer and in one healthy volunteer. Uridine was also examined for its effect on 5-fluorouracil toxicity in two patients. Uridine was administered as a 1-hr i.v. infusion at doses of 1 to 12 g/sq m. Plasma and urine samples were analyzed for uridine and uracil using high-pressure liquid chromatography. In 23 courses of uridine alone, the only toxicity observed was transient shivering after one of two courses at 12 g/sq m. This side effect was also seen after administration of uridine (10 g/sq m) during combination with 5-fluorouracil. The pretreatment plasma uridine concentration was elevated from low micromolar to millimolar levels with uridine administration at doses up to 12 g/sq m. Maximal areas under the concentration-time curve were about 5 mmol/liter/hr. Both peak plasma level and area under the curve for uridine increased linearly with dose. Uridine plasma decay curves were biphasic with a terminal half-life of 118 min. Half-life, volume of distribution (634 ml/kg), and total clearance (4.98 ml/kg/min) appeared to be independent of dose. Plasma uracil concentration increased gradually after administration of uridine to plateau levels. Maximal plasma uracil concentrations were about one-tenth that of peak uridine concentrations. The plasma uracil level declined with a half-life of about 40 min after uridine levels decreased to 300 microM. Total urinary excretion of uridine was 24% of the dose, while the amount of uracil recovered in urine was 3.4%. In two patients, uridine rescue was attempted during 5-fluorouracil dose escalation. Uridine at 5 to 6 g/sq m given on 1 or on 2 days after 5-fluorouracil did not prevent myelosuppression and gastrointestinal toxicity associated with increasing plasma concentrations of 5-fluorouracil. These data show that uridine administered as a 1-hr infusion at doses which provide peak plasma uridine concentrations in the millimolar range is well tolerated. Rapid elimination of uridine primarily due to catabolism results in modest exposure to substantially elevated plasma uridine concentrations. Preliminary findings suggest that prolonged treatment with uridine may be required to test its potential to rescue patients from 5-fluorouracil toxicity.

Clinical and biochemical studies of high-dose thymidine treatment in patients with solid tumors.

In a clinical study of high-dose thymidine (TdR) treatment, toxic effects, TdR metabolism, and the influence of TdR on pyrimidine and purine metabolism were examined. Ten patients with solid tumors were treated with continuous infusion of TdR at 34-75 g/m2/day for 3 to 5 days. Hematologic toxicity occurred with 5-day TdR infusion at 75 g/m2/day but not when plasma TdR concentration failed to reach millimolar levels. In three patients who received similar TdR doses, plasma TdR levels were related to elimination rates of TdR and its metabolites from plasma. In one patient in whom urinary excretion was studied, 100% of the TdR dose given was recovered in the form of TdR, thymine (Thy), beta-aminoisobutyrate, and 5-hydroxymethyluracil (5-HMUra). The latter metabolite, which had not been previously described in high-dose TdR treatment, was also found in plasma at levels from 5% to 10% of those of TdR. No effects of high-dose TdR infusion on purine levels in plasma were observed, while a substantial increase in uracil levels was noted both in plasma and urine. These data provide further information on high-dose TdR treatment with regard to clinical, pharmacokinetic, and biochemical effects.

Differential metabolism of thymidine in human lymphoid and melanoma cells in vitro.

Four potentially key enzyme activities of thymidine metabolism were examined in 4 human cell lines differing widely in sensitivity to thymidine. Two melanoma cells showed intermediate sensitivity to thymidine compared with highly sensitive T-lymphoid cells and relatively resistant B-lymphoid cells. Thymidine kinase activity varied modestly among the 4 cell lines; while thymidine phosphorylase activity was markedly higher in extracts of both melanoma cells and B-cells. dTMP phosphatase activity was markedly higher in extracts of melanoma cells compared to both T- and B-cells. The rate of dTTP degradation in intact cells was appreciably higher in the B-cells compared to both melanoma cells and T-cells. It is possible that elevated levels of thymidine phosphorylase activity account for decreased sensitivity to thymidine, and that enhanced dTTP catabolic activity imparts additional resistance.

High uridine phosphorylase activity in human melanoma tumor.

Uridine (Urd) phosphorylase and Urd kinase activities were examined in 4 human tumor types including melanoma and human and mouse melanoma cell lines. Urd phosphorylase activity in melanoma tumor specimens was higher than in specimens of colon, ovarian, and breast tumors. Urd kinase activity levels were similar in the 4 tumor types. In 3 human melanoma cell lines examined, Urd phosphorylase activity was markedly greater than in mouse B16 melanoma cells, while Urd kinase activity did not differ appreciably in the human and mouse cell lines. Urd phosphorylase activity in crude extract preparations from different melanoma cell lines showed similar substrate affinity and sensitivity to 1-(2'-deoxy-beta-D-glucopyranosyl)-thymine, a specific inhibitor. The high Urd phosphorylase activity found in melanoma tumor tissue may be exploited in the treatment of malignant melanoma with antipyrimidine agents. Cultured human melanoma cells retain this biochemical characteristic and may serve as appropriate in vitro models for the human tumor in studies concerning pyrimidine metabolism.

In vitro and in vivo studies on the combination of Brequinar sodium (DUP-785; NSC 368390) with 5-fluorouracil; effects of uridine.

Brequinar sodium (DUP-785; Brequinar) is a potent inhibitor of the pyrimidine de novo enzyme dihydroorotate dehydrogenase (DHO-DH), leading to a depletion of pyrimidine nucleotides, which could be reversed by uridine. In in vitro studies we investigated the effect of different physiological concentrations of uridine on the growth-inhibition by Brequinar, the effect of the nucleoside transport inhibitor, dipyridamole, and the combination of Brequinar and 5-fluorouracil (5FU). Uridine at 1 microM slightly reversed the growth inhibition by Brequinar, while the effect of 5-500 microM was greater. However, at Brequinar concentrations greater than 30 microM, uridine could not reverse the growth-inhibitory effects. Addition of dipyridamole could only partially prevent the reversing effects of uridine. The combination of Brequinar and 5FU was more than additive in the absence of uridine in the culture medium, but not in the presence of uridine. The combination of Brequinar and 5FU was tested in vivo in two murine colon tumour models, Colon 26 and Colon 38. Scheduling of both compounds appeared to be very important. In Colon 38 no potentiating effect of Brequinar could be observed. In contrast in Colon 26 a more than additive effect could be observed. Since uridine concentrations are considerably different in these tumours (higher in Colon 38), it was concluded from both the in vitro and in vivo experiments that uridine is an important determinant in combinations of Brequinar and 5FU.

Clinical and pharmacokinetic studies of prolonged administration of high-dose uridine intended for rescue from 5-FU toxicity.

A clinical and pharmacokinetic investigation of prolonged administration of high-dose uridine was performed in seven patients with advanced-stage cancer. Uridine administration was examined as a continuous infusion at 1 and 2.5 g/m2/hr (two patients) and as a series of intermittent infusions during 72 hrs at doses of 1-3 g/m2/hr, whereby 3-hr uridine administration was alternated with a 3-hr treatment-free interval (six patients). Continuous infusions of uridine resulted in plasma uridine concentrations of 0.5-1 mM, but was discontinued due to rapid increase in body temperature. Further studies focused on the intermittent schedule in an attempt to avoid the development of fever. Intermittent uridine infusion resulted in markedly elevated plasma uridine levels in the millimolar range. However, during the treatment-free period, rapid elimination of uridine was observed, resulting in plasma levels of 138-335 microM for 3 g/m2/hr. Plasma uracil concentrations also increased markedly, but smaller fluctuations compared to uridine were seen. Total urinary excretion of uridine was 15%-40% of the dose, while uracil excretion in urine was 2%-17%. Intermittent uridine infusion resulted in little or no rise in body temperature (less than or equal to 1.0 degrees C) in ten of 12 courses, and fever of greater than 39 degrees C in two courses. Both intermittent and continuous infusion of uridine gave rise to phlebitis, which necessitated central venous administration. These data show that using an intermittent infusion schedule, long-term administration of uridine is tolerable, with fever being dose-limiting. Intermittent infusion provides for the maintenance of markedly elevated plasma uridine levels and long-term uridine exposure to the tissues, and may be useful in further studies aimed at testing the potential of uridine to rescue patients from 5-FU toxicity.

Effect of pyrimidine nucleosides on body temperatures of man and rabbit in relation to pharmacokinetic data.

The effect of high-dose uridine on body temperatures of rabbits and man has been studied in relation to plasma concentrations of uridine and its catabolite uracil. Uridine induced fever in both rabbits and man. High-dose cytidine had no influence on body temperature in rabbits. Plasma concentrations of uridine were between 1 and 1.5 mM at 30 min after an iv bolus injection of 400 mg uridine/kg in rabbits and reached peak levels of 2 mM after a 1-hr infusion of 12 g uridine/m2 in man. The plasma concentration of cytidine in rabbits was about 0.5 mM and that of uridine was 0.30 mM at 30 min after an iv bolus injection of 400 mg cytidine/kg. The mean residence time for uridine in patients and rabbits varied between 80 and 195 min. The area under the plasma concentration-time curve (AUC) for uridine in rabbits was 2.0 mmol.hr/liter, and that for cytidine was 0.6 mmol.hr/liter. A large AUC for uridine indicates a prolonged exposure of tissues to uridine, which might lead to extensive formation of degradation products. The administration of some of these catabolites, dihydrouracil (at 20-40 mg/kg), carbamyl-beta-alanine (at 60 mg/kg), and beta-alanine (at 300-400 mg/kg), resulted in a significant increase in body temperature. It is concluded that the change in body temperature associated with uridine administration was not due to bacterial pyrogens but that one of the degradation products might be involved in thermoregulation.