Quantifying the DNA-damaging Effects of FLASH Irradiation With Plasmid DNA

Abstract

To investigate a plasmid DNA nicking assay approach for isolating and quantifying the DNA-damaging effects of ultrahigh-dose-rate (ie, FLASH) irradiation relative to conventional dose-rate irradiation. We constructed and irradiated phantoms containing plasmid DNA to nominal doses of 20 Gy and 30 Gy using 16 MeV electrons at conventional (0.167 Gy/s) and FLASH (46.6 Gy/s and 93.2 Gy/s) dose rates. We delivered conventional dose rates using a standard clinical Varian iX linear accelerator and FLASH dose rates (FDRs) using a modified Varian 21EX C-series linear accelerator. We ran the irradiated DNA and controls (0 Gy) through an agarose gel electrophoresis procedure that sorted and localized the DNA into bands associated with single strand breaks (SSBs), double strand breaks (DSBs), and undamaged DNA. We quantitatively analyzed the gel images to compute the relative yields of SSBs and DSBs and applied a mathematical model of plasmid DNA damage as a function of dose to compute the relative biological effectiveness (RBE) of SSB and DSB (RBESSB and RBEDSB) damage for a given endpoint and FDR. Both RBESSB and RBEDSB were less than unity with the FDR irradiations, indicating FLASH sparing. With regard to the more deleterious DNA DSB damage, the DSB RBEs of FLASH beams at dose rates of 46.6 Gy/s and 93.2 Gy/s relative to the conventional 16 MeV beam dose rate were 0.54 ± 0.15 and 0.55 ± 0.17, respectively. This study demonstrated the feasibility of using a DNA-based phantom to isolate and assess the FLASH sparing effect on DNA. We also found that FLASH irradiation causes less damage to DNA compared with a conventional dose rate. This result supports the notion that the protective effect of FLASH irradiation occurs at least partially via fundamental biochemical processes.