A Molecular Approach to Characterize the Effects of Fluence, Fluence Rate and LET on Radiation Damage

Abstract

Effects of fluence, fluence rate and LET on radiation damage are not resolved using traditional methods of measuring energy deposited by ionization events. The deficiency led to the use of empirical RBE factors in the clinical applications of particle therapy. The use of ionization dosimetry is similarly challenged when applied to development of radiation treatment at ultrahigh (FLASH) dose rate. This study reports a molecular method using plasmid DNA as a more comprehensive model for radiation dosimetry than ionization measurements. Aqueous solutions of purified supercoiled plasmid DNA, pUC19 (2686 bp), were prepared at different scavenging conditions and injected into 5x5x1 mm3 wells as detector elements. Irradiated samples were analyzed using base excision repair enzymes (Nth and Fpg) and gel electrophoresis to measure yields for DNA single and double strand breaks (SSB and DSB), and clustered lesions. The low LET characteristics of conventional radiation treatment was modeled using orthovoltage 150 kVp x-rays to deliver 2-110 Gy at 90 and 0.5 Gy/s. Higher LET irradiations in the range of 2 - 14 keV/μm were facilitated by measurements in the pristine Bragg peak region using synchrotron-produced 142.2 MeV protons to deliver dose at 2 - 160 Gy at 600 and 1 Gy/s. The DNA wells were inserted into a solid water equivalent phantom for proton irradiation. Only 4 wells could be positioned in the short Bragg peak region in water (∼ 2 cm). To alleviate the uncertainties due to rapidly varying dose and LET distributions, we innovated the use of a 3% water density (i.e., Styrofoam) medium to extend the Bragg peak region from 2 cm to 20 cm, enabling the placement of 20 well containers. Quantity and quality of molecular damage in the plasmid DNA model varies with fluence, fluence rate and LET of radiation. At high fluence (> 30 Gy) of low-LET radiations, the yields of DNA SSB and non-DSB clustered lesions depend on the fluence rate. These yields decrease by two times between ultrahigh and conventional dose rate irradiation. At a given fluence and fluence rate, the yields for the formation of DNA DSB and non-DSB clustered lesions increase linearly with LET. The low-density phantom allows significant (∼ 10 folds) increase in the number of sampling points and more accurate sample positioning at specific LET compared to water-equivalent phantom. Monte-Carlo track structure simulation of yields for different DNA lesions is being developed to model the molecular damage. In parallel, approaches to improve the sensitivity of the measurements to dose are being investigated. Our results suggest that a molecular-based approach can be used to differentiate the effects of fluence, fluence rate, and LET on radiation damage. The approach demonstrates the potential to improve on the modeling of radiation effects in biological systems than using measured ionization energy. Correlation of the molecular changes to biological outcome for in vitro and in vivo systems are under investigation.