Variable baseline gamma-glutamylcysteine synthetase messenger RNA expression in peripheral mononuclear cells of cancer patients, and its induction by buthionine sulfoximine treatment.

ABSTRACT

The role of glutathione (GSH) in tumor cell resistance to alkylating agents and platinum compounds is suggested by a body of laboratory and clinical studies. The rate-limiting enzyme in GSH synthesis is gamma-glutamylcysteine synthetase (gamma-GCS), the expression of which is proportional both to GSH content and to the level of resistance in ovarian cancer cell lines. The role of this enzyme in regulating GSH levels is unclear, however. Reversal of resistance is achieved in vitro and in vivo with the use of buthionine sulfoximine (BSO), a potent inhibitor of gamma-GCS. In the course of a Phase I clinical trial of BSO and melphalan, we have measured GSH and expression of gamma-GCS mRNA in peripheral mononuclear cells before and at intervals after the initiation of treatment with BSO. Mean baseline GSH content was 6.89 nmol/mg protein. Treatment with BSO (10.5 to 17 g/m2 i.v. every 12 h for six doses) resulted in a mean nadir GSH decline to 19% of control values, most commonly on day 3. Baseline expression of gamma-GCS mRNA was measured by a reverse transcriptase polymerase chain reaction-based method. When described relative to that of beta-actin, the expression of gamma-GCS varied over 3-fold among individuals. Following GSH depletion by BSO, the level of gamma-GCS mRNA rose successively on days 3 and 5 to reach a mean increase of 2-fold on day 8. Differences were observed among patients in their capacity to respond to GSH depletion by increasing gamma-GCS steady-state mRNA levels (1.4- to 3.1-fold). These results show that the expression of gamma-GCS is variable in the population and suggest that the cellular content of GSH may be involved in the regulation of its expression.