Feasibility Study on the Use of Gold Nanoparticles as a Dose Enhancement Agent for a Superficial X-ray Therapy Applied to Melanoma

Abstract

The aim of radiation therapy is to kill tumor cells using ionizing radiation while sparing surrounding normal tissues. Recent advances in radiation therapy have resulted in the development of intensity modulated radiation therapy (IMRT) that allows the dose to conform more precisely to the three-dimensional shape of the tumor. However, the equipment-based beam delivery methods have a limitation of providing a curative radiation dose to the tumor volume without exceeding normal tissue tolerance because of the similar x-ray absorption characteristics of tumors and surrounding normal tissues. By loading high atomic number nanoparticles on tumor volume, it is possible to deliver high radiation doses to tumor volume while sparing normal tissues in kilovoltage x-ray beams. The concept of using high atomic number nanoparticles as a dose enhancement agent is premised on the high photoelectric mass absorption coefficient of high atomic number materials in kilovoltage photon energy region, compared with soft tissues. Gold nanoparticles (AuNPs) have been of particular interest to researchers in recent years because of its high photoelectric mass absorption coefficient and biocompatibility. The number of skin cancer patients has been increasing every year. AuNPs are expected to contribute to improving the efficiency of skin cancer radiation therapy as a dose enhancement agent. Prior to application of AuNPs as a dose enhancement agent in therapeutic purpose, more biological observations are required to clarify the physical predictions of the dose enhancement effect and to entirely understand the radiobiological responses of AuNPs. The ultimate goal of the present in vitro study was to investigate the potential of AuNPs as a dose enhancement agent in x-ray radiation therapy for skin cancer, especially melanoma. Three types of cell lines, skin melanoma cells, gliosarcoma cells and normal dermal fibroblast cells, and two different sizes of the spherical AuNPs, 1.9 and 50 nm in diameter, were used in this study. Cells were irradiated at room temperature in the hard x-ray beam irradiation facility (YXLON model 450-D08) at Seoul National University, with 150 kVp (superficial) and 450 kVp (orthovoltage) x-rays. The clonogenic survival assay was conducted to investigate the dose enhancement effect as functions of AuNP size and concentration, photon energy, and cell-type. Also, the cytotoxicity assay, the observation of cellular localized AuNPs, the DNA double strand break (DSB) analysis, and the cell cycle analysis were performed to support the main results. MCNP-5 (Monte Carlo N-particle-5) calculations were performed to obtain the depth dose curves and the x-ray energy spectra in depth from the skin surface. From the experiment, it was confirmed that the optimal combinations of AuNP size, concentration, and x-ray energy resulted in cells having high-linear energy transfer (LET) - like survival curve, leading to enhancing the cell radiosensitivity. The dose enhancement effect was also strongly dependent on cell-type and it was supposed to be partly contributed by the different efficiency of cellular uptake following cell type. AuNPs gave a significant dose enhancement effect on melanoma cells, which are well known as the most radioresistant cells in all types of skin cancers. The maximum dose enhancement factor was 2.29 for skin melanoma cells at 320 μM of 50 nm AuNPs with 150 kVp x-ray beams. To confirm the effect of those conditions on normal skin cells, the experiments were carried out on dermal fibroblast cells. 50 nm AuNPs had no remarkable toxicity on dermal fibroblast cells and provided lower dose enhancement effect to dermal fibroblast cells than skin melanoma cells. However, dose enhancement effect on dermal fibroblast cells should not be overlooked and this result emphasizes the importance of the accurate AuNP delivery to melanoma. By applying AuNPs to conventional fractionated radiation therapy of skin cancer, the major concerns of fractionation regimens are expected to be overcome. Although relatively high doses are deposited to tumor by applying AuNPs, the normal tissue damage would not be more significantly severe because radiation doses delivered from the equipment do not increase. Also by shortening the overall treatment time, AuNPs can contribute to resolving concern about tumor cell repopulation, which is a main cause of lowering the efficacy of the fractionated radiation therapy. In conclusion, the application of 50 nm AuNPs as a dose enhancement agent in superficial x-ray therapy could be a promising treatment method for T1 to T3 stages of melanoma. 50 nm AuNPs are preferably accumulated in melanoma by passive action. Modification of 50 nm AuNP with melanoma specific ligand would even further enhance the therapeutic effect.