Room: Karl Dean Ballroom A1
Purpose: To measure the effects of LET on DNA double-strand break (DSB) repair along a clinical scanned proton beam and correlate repair markers with clonogenic outcomes.
Methods: H460 cells were irradiated with a scanning proton beam using an established high-throughput setup. The cells were fixed along a 24-hour time course. DSB repair markers were examined by immunofluorescent foci staining. Plates were automatically imaged using the Cytation5 imaging platform. Automated image and data analysis were carried out using CellProfiler and R respectively. Graphpad Prism was used for data plotting and statistical analysis.
Results: Dose-normalized persistent DSBs per cell vs LET correlated well with clonogenic RBE. 53bp1 foci induction saturated in an LET-dependent manner with the saturation occurring at lower doses for higher LETs, indicative of increased biological efficacy. The number of persistent 53bp1 foci scaled quadratically with dose and demonstrated an LET-independent relationship with clonogenic SF.Both non-homologous end joining (DNA-PKcs foci) and homologous recombination (Rad51 foci) were found to contribute to DSB resolution. The number of persistent DNA-PKcs foci scaled linearly with dose while Rad51 foci trended quadratically. The rates of both DNA-PKcs and Rad51 foci accumulation were observed to increase with LET. For the lowest LETs, 0.97 and 2.6keV/Î¼m, no persistent Rad51 foci above background were observed even at the maximum dose of 4.2Gy. For LETs sampled within the regions proximal to the Bragg peak, 4-6keV/Î¼m, persistent Rad51 foci were found for doses above 5Gy. A similar trend was measured for regions associated with the Bragg peak. Rad51 foci persisting 24-hours post-irradiation were found to accumulate even for low doses in the distal falloff region.
Conclusion: At the DNA repair level, proton LET appears to behave as a dose multiplier resulting in separable biological endpoints. The observed increase in persistent Rad51 foci could be indicative of complex DNA damage.