Enzyme-pattern-targeted chemotherapy with tiazofurin and allopurinol in human leukemia

Abstract The hypothesis was tested that the increased IMP dehydrogenase activity in human myelocytic leukemic cells, and along with it guanylate biosynthesis, might be a sensitive target to chemotherapy by tiazofurin. 1. 1.|IMP dehydrogenase activity in normal leukocytes was 3.1 ± 0.5 (means ± S.E.)nmol/hr/mg protein and in leukemic cells it was elevated 15- to 41-fold. The activity of guanine phosphoribosyltransferase in normal leukocytes was 389 ± 27 nmol/hr/mg protein and in the leukemic cells it increased 2.8-to 6.8-fold. 2. 2.|IMP dehydrogenase was purified 4,900-fold to homogeneity from rat hepatoma 3924A with a yield of 30%. The kinetic properties of the hepatoma enzyme were similar to those of the enzyme in human myelocytic leukemic blast cells because of the similarity of the Km's for IMP (23 μ m ), NAD (44 and 65 μ m ); the Ki for TAD was 0.1 μ m in both enzymes. 3. 3.|There was a selectivity of the in vitro response to tiazofurin in human normal and leukemic leukocytes. When labeled tiazofurin was incubated with leukocytes from normal, healthy volunteers and from leukemic patients, the leukemic leukocytes made 20- to 30-fold more TAD and the GTP content decreased as compared to normal leukocytes. This procedure proved to be a suitable predictive test in a clinical setting because patients with positive tests resonded to tiazofurin whereas those with negative ones did not. 4. 4.|The National Cancer Institute approved a chemotherapeutic phase I/II trial which concentrates on treatment of refractory acute myelocytic leukemia. Tiazofurin is infused in a 60-minute period with a pump to insure uniform delivery. A novel aspect of the trial was that it was directed primarily by the biochemical impact of tiazofurin on IMP dehydrogenase activity and GTP concentration and the tiazofurin doses were to be adjusted accordingly. Patients received allopurinol as a routine precaution against possible accumulation of uric acid in the kidney. 5. 5.|In the first eight patients, there was one complete remission, two entered the chronic phase, two entered into partial remission, one did not respond, and two were not evaluable. In the five patients who responded, there was a rapid, profound decrease in IMP dehydrogenase activity of the blast cells and a gradual decline in GTP concentrations. The blast cell count followed the decrease in the GTP concentration. The white blood cell count was largely preserved. 6. 6.|Bone marrow aspirates and peripheral blood samples showed that with tiazofurin treatment there was an induced differentiation of the myelocytes. 7. 7.|Allopurinol treatment (300 mg per kg) increased the plasma hypoxanthine concentration from the normal level of about 2 to 3 μ m to 20 to 30 μ m . In presence of low concentrations of guanine in the plasma (less than 0.1 μ m ) the 10-fold elevated hypoxanthine concentration would strongly curtail the activity of the salvage enzyme, GPRT. Thus, the inhibition of blast cell IMP dehydrogenase activity by tiazofurin in presence of a markedly diminished GPRT activity caused by the allopurinol treatment resulted in a combination chemotherapeutic impact, leading to the depletion of the blast cell GTP concentration. 8. 8.|Tiazofurin treatment provided hematological remission in five patients with induced differentiation of the bone marrow. Tiazofurin therapy of refractory AML resulted in no leukopenia; therefore, this treatment avoids the morbidity and mortality encountered in conventional chemotherapy of refractory AML. 9. 9.|This clinical investigation provides evidence for the significance of the increased IMP dehydrogenase activity and the elevated capacity of the guanylate biosynthetic pathway in the expression of proliferation and differentiation of human leukocytes.