THE EFFECTS OF CYCLOHEXIMIDE AND WR-1065 ON RADIATION-INDUCED REPAIR PROCESSES - A MECHANISM FOR CHEMOPREVENTION

The effects of cycloheximide (CHX) and 2-[(aminopropyl)amino]ethanethi ol (WR-1065), each alone or in combination, on radiation-induced mutat ion induction at the hypoxanthine-guanine phosphoribosyl transferase ( hprt) locus and cell killing were investigated using a Chinese hamster ovary (CHO) AAS cell system, Treatment with CHX, a potent inhibitor o f protein synthesis, at a concentration of 10 mu g/ml administered 30 min prior to irradiation with 7.5 Gy had no effect on cell survival bu t did reduce the radiation-induced mutation frequency (per 10(6) survi vors) from 106.5 +/- 8.8 (SEM) to 36.2 +/- 5.6 (SEM), Exposure of cell s to 4 mM WR-1065 reduced the mutation frequency to 44.8 +/- 4.2 (SEM) , but the combination of agents afforded no additional protection, tha t is 41.1 +/- 3.3 (SEM), The mechanism of action attributed to CHX in reducing mutation frequency is its ability to prevent the induction of an error-prone repair system, Split-dose radiation experiments, that is 8 Gy versus 4 Gy + 4 Gy separated by 3 h, were performed to evaluat e and contrast the relative abilities of CHX and WR-1065, each alone o r in combination, in affecting cell survival, Cycloheximide administer ed to cells 30 min before the first radiation dose and present through out the 3 h incubation time prior to the second dose inhibited split-d ose repair as evidenced by a reduction in surviving fraction by 60% as compared with the value obtained for non-CHX-treated cells that were exposed to two equal doses of 4 Gy, Cells exposed to 4 mM WR-1065 imme diately following the first 4 Gy radiation dose and then washed free 2 .5 h before exposure to a second 4 Gy dose, which was also followed by a 30 min exposure to WR-1065, increased the surviving fraction by 80% over the value obtained for cells not exposed to WR-1065 during their split-dose radiation treatment, When CHX treatment was combined with WR-1065 exposure, the protective effect of WR-1065 was abolished, that is surviving cell fraction was again reduced by similar to 60% as com pared with untreated control groups, These results indicate that prote in synthesis is required for WR-1065 to affect split-dose related repa ir processes, Presumably, the inhibition of the induction of an error- prone repair system by CHX would account for its effects on both resul tant decreases in mutation frequency and cell survival, In contrast, W R-1065 and/or its disulfide metabolite appear to facilitate the effica cy and fidelity of such a repair system once it is induced. This is mo st probably the result of their polyamine-like structures and properti es, which can influence the stability of DNA damaged sites as well as inhibit cell cycle progression, thus leading to longer times available in which to complete repair before damage is fixed at cell division.