Low and High Concentrations of the Topo II Inhibitor Daunorubicin in NIH3T3 Cells: Reversible G2/M Versus Irreversible G1 and S Arrest

Abstract

Daunorubicin (DNR) blocks the cell cycle by interfering with synthesis and repair of DNA. In both drug-sensitive 3T3 cells, and drug-resistant 3T3 cells, NIH-MDR-6815, (created by transfection with a human MDR1 cDNA), low concentrations of DNR (up to 80 ng/ml in sensitive cells, 1600 ng/ml in resistant cells), cells initially slowed S-phase progression for 2 to 3 hours, but the treated cells then continued in progression at a steady rate, close to that of untreated cells and accumulated in G2/M. The 2 to 3 h lag period represents the time taken for fully establishing the G2/M block. The time required to bring about cessation of proliferation is the sum of this lag period and the time taken to travel through the cell cycle. This low concentration effect is cytostatic, and fully reversible on washing out the daunorubicin. At higher drug concentrations (above 160 ng/ml in sensitive cells, 3200 ng/ml in resistant cells) the cells became blocked in both G1 and S, and did not reach G2/M. The high concentration effect was cytotoxic and irreversible, and was followed by cell death. Only cells that were in S phase were subject to this block in S, since cells that had accumulated in G2/M by using a low concentration (60 ng/ml DNR for 20 h) were not blocked in S, and did not die, when subsequently treated with high drug concentrations (320 ng/ml, 30 h). The low concentration effect occurred at the same maximal rate (4 %/h) in sensitive or resistant cells, but the external drug concentration required to produce half the maximal rate was, appropriately, twenty-fold higher in the resistant cells (20 ng/ml and 400 ng/ml, respectively).