Cell localisation of gadolinium-based nanoparticles and related radiosensitising efficacy in glioblastoma cells

Lenka Štefančíková, Erika Porcel, Pierre Eustache, Sha Li, Daniela Salado, Sergio Marco, Jean-Luc Guerquin-Kern, Matthieu Réfrégiers, Olivier Tillement, François Lux, Sandrine Lacombe

Recently, the addition of nanoparticles (NPs) has been proposed as a new strategy to enhance the effect of radiotherapy particularly in the treatment of aggressive tumors such as glioblastoma. The physical processes involved in radiosensitisation by nanoparticles have been well studied although further understanding of its biological impact is still lacking, and this includes the localisation of these NPs in the target cells. Most studies were performed with NPs tagged with fluorescent markers. However, the presence of these markers can influence the NPs uptake and localisation. In this study, a set of methods was used to unambiguously and fully characterise the uptake of label-free NPs, their co-localisation with cell organelles, and their radiosensitising efficacy. This set was applied to the case of gadolinium-based nanoparticles (GdBN) used to amplify the radiation killing of U87 glioblastoma cells extracted from highly aggressive human tumor. For the first time, Synchrotron Radiation Deep UV (SR-DUV) microscopy is proposed as a new tool to track label-free GdBN. It confirmed the localisation of the NPs in the cytoplasm of U87 cells and the absence of NPs in the nucleus. In a second step, Transmission Electron Microscopy (TEM) demonstrated that GdBN penetrate cells by endocytosis. Third, using confocal microscopy it was found that GdBN co-localise with lysosomes but not with mitochondria. Finally, clonogenic assay measurements proved that the presence of NPs in the lysosomes induces a neat amplification of the killing of glioblastoma cells irradiated by gamma rays. The set of combined experimental protocols-TEM, SR-DUV and confocal microscopy-demonstrates a new standard method to study the localisation of label-free NPs together with their radiosensitising properties. This will further the understanding of NP-induced radiosentisation and contribute to the development of nanoagents for radiotherapy.