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]ethanethiol (WR-1065), each alone or in combination, on radiation-induced mutation induction at the hypoxanthine-guanine phosphoribosyl transferase (hprt) locus and cell killing were investigated using a Chinese hamster ovary (CHO) AA8 cell system. Treatment with CHX, a potent inhibitor of protein synthesis, at a concentration of 10 micrograms/ml administered 30 min prior to irradiation with 7.5 Gy had no effect on cell survival but did reduce the radiation-induced mutation frequency (per 10(6) survivors) from 106.5 +/- 8.8 (SEM) to 36.2 +/- 5.6 (SEM). Exposure of cells to 4 mM WR-1065 reduced the mutation frequency to 44.8 +/- 4.2 (SEM), but the combination of agents afforded no additional protection, that 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 evaluate and contrast the relative abilities of CHX and WR-1065, each alone or in combination, in affecting cell survival. Cycloheximide administered to cells 30 min before the first radiation dose and present throughout the 3 h incubation time prior to the second dose inhibited split-dose repair as evidence 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 immediately following the first 4 Gy radiation dose and then washed free 2.5 h before exposure to a second 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 was abolished, that is surviving cell fraction was again reduced by approximately 60% as compared with untreated control groups. These results indicate that protein synthesis is required for WR-1065 to affect split-dose related repair processes. Presumably, the inhibition of the induction of an error-phone repair system by CHX would account for its effects on both resultant decreases in mutation frequency and cell survival. In contrast, WR-1065 and/or its disulfide metabolite appear to facilitate the efficacy and fidelity of such a repair system once it is induced.(ABSTRACT TRUNCATED AT 400 WORDS)
Murley
J S
JS
Center for Mechanistic Biology and Biotechnology, Argonne National Laboratory, IL 60439-4833, USA.
Grdina
D J
DJ
D000818:Animals; D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D016466:CHO Cells; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D003513:Cycloheximide / Q000494:pharmacology*; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects*; D008624:Mercaptoethylamines / Q000494:pharmacology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D011837:Radiation-Protective Agents / Q000494:pharmacology*
D016588:Anticarcinogenic Agents; D008624:Mercaptoethylamines; D011500:Protein Synthesis Inhibitors; D011837:Radiation-Protective Agents; C020174:N-(2-mercaptoethyl)-1,3-diaminopropane; D003513:Cycloheximide