Effects of Streptozotocin/Bis-chloroethylnitrosourea Combination Therapy on {T'-Methylguanine DNA Methyltransferase Activity and mRNA Levels in HT-29 Cells in Vitro1

Treatment of chloroethylnitrosourea-resistent cells with streptozotocin (STZ) prior to bis-chloroethylnitrosourea (BCNU) exposure has been shown to result in a depletion of 06-methylguanine DNA methyltransferase (MGMT) activity, increased BCNU-induced interstrand cross-link ing, and a 2-3 log enhancement of BCNU cytotoxicity in vitro. The current study was undertaken to define the kinetics of repletion of MGMT activity following the STZ/BCNL1 combination and to assess at the molecular level the effects of the combination on MGMT mRNA expres sion. Results demonstrate that MGMT activity can be depleted by >90% relative to untreated controls using an optimized STZ/BCNU combination regimen and that >50% depletion can be maintained for at least 24 h. This depletion appears to be independent of effects at the mRNA level because neither STZ alone nor the STZ/BCNU combination significantly altered steady state levels of MGMT mRNA. Cytotoxicity studies are consistent with MGMT repletion data and demonstrate that, as the interval between STZ and BCNU exposures increases, the degree of enhanced cytotoxicity induced by the combination relative to BCNU alone decreases. These results suggest that the enhanced cytotoxicity induced by the STZ/BCNU combination over BCNU treatment alone is favored by both the lack of induction of expression of MGMT mRNA and by slow reappearance of MGMT activity.