Validation of the plasmid study to relate DNA damaging effects of radionuclides to those from external beam radiotherapy

Abstract

Introduction The biological consequences of absorbed radiation doses are ill-defined for radiopharmaceuticals, unlike for external beam radiotherapy (EBRT). A reliable assay that assesses the biological consequences of any radionuclide is much needed. Here, we evaluated the cell-free plasmid DNA assay to determine the relative biological effects of radionuclides such as Auger electron-emitting [ 67 Ga]GaCl 3 or [ 111 In]InCl 3 compared to EBRT. Methods Supercoiled pBR322 plasmid DNA (1.25 or 5 ng/μL) was incubated with 0.5 or 1 MBq [ 67 Ga]GaCl 3 or [ 111 In]InCl 3 for up to 73 h or was exposed to EBRT( 137 Cs; 5 Gy/min; 0-40 Gy). The induction of relaxed and linear plasmid DNA, representing single and double strand breaks, respectively, was assessed by gel electrophoresis. Chelated forms of 67 Ga were also investigated using DOTA and THP. Topological conversion rates for supercoiled-to-relaxed or relaxed-to-linear DNA were obtained by fitting a kinetic model. Results DNA damage increased both with EBRT dose and incubation time for [ 67 Ga]GaCl 3 and [ 111 In]InCl 3 . Damage caused by [ 67 Ga]GaCl 3 decreased when chelated. [ 67 Ga]GaCl 3 proved more damaging than [ 111 In]InCl 3 ; 1.25 ng/μL DNA incubated with 0.5 MBq [ 67 Ga]GaCl 3 for 2 h led to a 70% decrease of intact plasmid DNA as opposed to only a 19% decrease for [ 111 In]InCl 3 . For both EBRT and radionuclides, conversion rates were slower for 5 ng/μL than 1.25 ng/μL plasmid DNA. DNA damage caused by 1 Gy EBRT was the equivalent to damage caused by 0.5 MBq unchelated [ 67 Ga]GaCl 3 and [ 111 In]InCl 3 after 2.05 ± 0.36 and 9.3 ± 0.77 h of incubation, respectively. Conclusions This work has highlighted the power of the plasmid DNA assay for a rapid determination of the relative biological effects of radionuclides compared to external beam radiotherapy. It is envisaged this approach will enable the systematic assessment of imaging and therapeutic radionuclides, including Auger electron-emitters, to further inform radiopharmaceutical design and application.

Nanoscopic biodosimetry using plasmid DNA in radiotherapy with metallic nanoparticles

“…They justified their results by the fact that the mean free path of free radicals decreases by increase of DNA concentrations. The similar results was reported by Verger et al 55 . They showed that for 5 ng/μL of plasmid, a higher percent of DNA damage was observed than for 1.25 ng/μL plasmid DNA.…”

“…The similar results was reported by Verger et al. 55 They showed that for 5 ng/μL of plasmid, a higher percent of DNA damage was observed than for 1.25 ng/μL plasmid DNA. Using different concentration of plasmids, no significant difference in DNA damage was observed by Brabcová et al.…”

“…They justified their results by the fact that the mean free path of free radicals decreases by increase of DNA concentrations. The similar results was reported by Verger et al 55 They showed that for 5 ng/μL of plasmid, a higher percent of DNA damage was observed than for 1.25 ng/μL plasmid DNA. Using different concentration of plasmids, no significant difference in DNA damage was observed by Brabcová et al 21 Kong et al 56 indicates that the proportion between free radical damage and direct ionizing damage is independent of DNA concentration when the DNA concentration is under a certain value (50 ng/μL).…”

Searching for a Paradigm Shift in Auger-Electron Cancer Therapy with Tumor-Specific Radiopeptides Targeting the Mitochondria and/or the Cell Nucleus

“…The influence of the proximity to DNA and accumulation in the cell nucleus on DNA damage and other radiobiological effects was thoroughly investigated for several other AE emitters, namely SPECT radiometals in clinical use such as 67 Ga, 111 In or 99m Tc or less common but more efficient AE emitters such as 195m Pt and 191 Pt [14,15]. For instance, Terry's group demonstrated how the DNA damage induced by 67 Ga and 111 In depends on their proximity to the DNA using a cell-free plasmid DNA assay [16,17]. Several classical DNA intercalators or groove binders have been studied as nuclear-targeting vectors for AE-emitting radionuclides, such as radiolabeled derivatives of acridine orange (AO) [18], pyrene [19], DAPI [20] and doxorubicin [21].…”

Status of radiobiology in molecular radionuclide therapy – Hope for the future

“…The research papers cover studies in cytogenetics after radium-223 treatment [9], the role of antioxidants in the radiosensitivity to [ 177 Lu]Lu-DOTATATE [10], relating the biological effects of radionuclides such as Auger electron-emitters gallium-67 and indium-111 to that of EBRT [11] and determining the DNA damaging capacity of Auger electron-emitter thallium-201 [12]. The future of Auger electronemitting radiopharmaceuticals was also discussed in a commentary [13].…”