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Gold nanoparticles (GNPs) emerged as promising antitumor radiosensitizers. However, the complex dependence of GNPs radiosensitization on the irradiation conditions remains unclear. In the present study, we investigated the impacts of the dose rate and photon energy on damage of the pBR322 plasmid DNA exposed to X-rays in the presence of 12 nm, 15 nm, 21 nm, and 26 nm GNPs. The greatest radiosensitization was observed for 26 nm GNPs. The sensitizer enhancement ratio (SER) 2.74 ± 0.61 was observed at 200 kVp with 2.4 mg/mL GNPs. Reduction of X-ray tube voltage to 150 and 100 kVp led to a smaller effect. We demonstrate for the first time that the change of the dose rate differentially influences on radiosensitization by GNPs of various sizes. For 12 nm, an increase in the dose rate from 0.2 to 2.1 Gy/min led to a ~1.13-fold increase in radiosensitization. No differences in the effect of 15 nm GNPs was found within the 0.85–2.1 Gy/min range. For 21 nm and 26 nm GNPs, an enhanced radiosensitization was observed along with the decreased dose rate from 2.1 to 0.2 Gy/min. Thus, GNPs are an effective tool for increasing the efficacy of orthovoltage X-ray exposure. However, careful selection of irradiation conditions is a key prerequisite for optimal radiosensitization efficacy.

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