Characterisation of VP-16-induced DNA cleavage in oestrogen-stimulated human breast cancer cells

Abstract

Cycling cells are recognised to be more susceptible than quiescent cells to the cytotoxic action of many commonly used cancer chemotherapeutic agents. We have found that oestrogen stimulation of T-47D human breast cancer cells is accompanied by a two-fold increase in VP-16-induced DNA cleavage as measured by alkaline DNA unwinding, and that this increase in DNA cleavage is accompanied by a corresponding enhancement of drug-induced cytostasis. The enhancement of VP-16-induced DNA cleavage seen with oestrogen exposure is antagonised both by antioestrogen treatment and by cycloheximide, an inhibitor of protein synthesis, but not by the DNA synthesis inhibitor aphidicolin. Increased c-myc protein synthesis is detectable within an hour of oestrogen exposure, while increased VP-16-induced DNA cleavage is detectable within 4h and increased DNA synthesis within 16h. Only small changes in cell-cycle distribution occur with oestrogen stimulation. In the absence of VP-16, oestrogen does not reduce DNA double-strandedness, nor does it induce changes in chromatin structure as measured by alterations in DNA superhelicity or chromatin accessibility. These findings suggest that oestrogen enhances VP-16-induced DNA damage in asynchronously growing G1-phase cells and that this enhancement may be dependent at some point upon de novo protein synthesis. Oestrogen pre-treatment of T-47D human breast cancer cells improves the therapeutic index of VP-16 without the need for cell synchronisation or highly precise drug scheduling.