Effects of streptozotocin/bis-chloroethylnitrosourea combination therapy on O6-methylguanine DNA methyltransferase activity and mRNA levels in HT-29 cells in vitro.

Treatment of chloroethylnitrosourea-resistant cells with streptozotocin (STZ) prior to bis-chloroethylnitrosourea (BCNU) exposure has been shown to result in a depletion of O6-methylguanine DNA methyltransferase (MGMT) activity, increased BCNU-induced interstrand cross-linking, 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/BCNU combination and to assess at the molecular level the effects of the combination on MGMT mRNA expression. Results demonstrate that MGMT activity can be depleted by greater than 90% relative to untreated controls using an optimized STZ/BCNU combination regimen and that greater than 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.