PHARMACODYNAMICS OF PROLONGED TREATMENT WITH L,S-BUTHIONINE SULFOXIMINE

Purpose: To develop dosing criteria for the use of L-buthionine-S-sulf oximine (active diastereoisomer) as a glutathione depletor in the clin ic, using a pharmacodynamic and pharmacokinetic in vitro-in vivo appro ach. Methods and Materials: In vitro: L-buthionine-S-sulfoximine uptak e was determined in human glioblastoma cells (T98G) and NIH-3T3 cells using S-35-labeled drug. Dose response relationships were derived for inhibition of glutathione synthesis in CHO cells, and for depletion of glutathione in exponentially growing T98G and CHO cells, as a functio n of extracellular L-buthionine-S-sulfoximine concentration. Steady-st ate glutathione levels for CHO and NIH-3T3 cells were measured using a n enzymatic assay, while glutathione synthesis rates in CHO cells were determined using a flow cytometric assay. In vivo: L-buthionine-S-sul foximine biodistribution was determined in male nude mice carrying hum an glioblastomas (T98G) intracranialy, using S-35-labeled drug infused subcutaneously by osmotic pump. Tissue glutathione levels were measur ed using an enzymatic assay. Results and Conclusion: The observed cell ular uptake t(1/2) of approximately 55 min, coupled with a previously reported, rapid in vivo clearance of buthionine sulfoximine, suggest t hat continuous infusion would be preferable to bolus dosing. Effective concentrations of L-buthionine-S-sulfoximine (24 h exposure), require d to lower cellular glutathione content to 50% of control (EC(50)), we re under 1 mM for both cell lines. The amount of L-buthionine-S-sulfox imine in tissues (estimated from S-35 drug disposition) reached steady state within 8 h and was proportional to the rate of infusion. Brain tumors were depleted to approximately 50% of control glutathione by a infusion rate of 0.25 mu moles/h (25 g mice). At lower infusion rates an increase in glutathione content was noted in certain nude mouse tis sues including brain tumor xenografts.