The effect of hypoxia on the induction of strand breaks in plasmid DNA by alpha-, beta- and Auger electron-emitters ²²³Ra, ¹⁸⁸Re, ^99mTc and DNA-binding ^99mTc-pyrene

Radiation-induced DNA damage occurs as a consequence of both direct and indirect effects of ionizing radiation. The induction mechanism of DNA damage is mainly influenced by the physical characteristics of the radiation quality, especially the linear energy transfer. In general, hypoxia reduces the effect of irradiation treatment in tumor cells and leads to poor patient outcomes. Emitters with high linear energy transfer (alpha- or Auger-electron-emitters) can overcome this obstacle. Our aim is to demonstrate the influence of hypoxia on the interaction between different radiation qualities with DNA by using a cell free plasmid model modulated by the free radical scavenger dimethyl sulfoxide (DMSO).
Plasmid DNA was irradiated with 223Ra, 188Re, 99mTc and DNA-binding 99mTc-pyrene in the absence or presence of DMSO and either under normoxic or hypoxic conditions. The resulting DNA damage in form of single- (SSB) and double strand breaks (DSB) was analyzed by agarose gel electrophoresis. Applied radiation doses of up to 200 Gy of 223Ra, 188Re or 99mTc or 60 Gy of 99mTc-pyrene led to maximal yields of SSB (80%) in plasmid DNA. Irradiation with 223Ra, 188Re or 99mTc at 200 Gy induced 30%, 28% and 32% linear plasmid conformations, respectively, which are associated with DSB. Hypoxia had a minor effect on SSB and DSB induction from 223Ra but a small enhancement in DSB for 188Re and 99mTc. DMSO could prevent DSB completely and SSB DNA damage from the three “free” radionuclides to comparable levels. DNA-binding 99mTc-pyrene induced less SSB and DSB compared to free 99mTcO4- due to its own radical scavenging properties. However, an additional incubation of DMSO could prevent the SSB and DSB induction only to a minor extent. Direct insults of Auger-electrons from 99mTc-pyrene are more effective than high-energy electrons or alpha particles due to the minimal distance between the radionuclide and the DNA.
We conclude that hypoxia does not limit DNA damage in plasmids induced by 223Ra, 188Re, 99mTc and 99mTc-pyrene. Dose-dependent radiation effects were comparable for alpha-emitters and both high- and low-energy electron emitters. The radioprotection by DMSO was not influenced by hypoxia. Overall, the results indicate the contribution of mainly indirect radiation effects for 99mTc, 188Re and 223Ra. 99mTc-pyrene caused direct DNA damages. The direct participation of oxygen in cell-free plasmid DNA damage induction was not proven.