Radiobiological effects of alpha-particles from Astatine-211: From DNA damage to cell death

摘要

ABSTRACTIn recent years, the use of high linear energy transfer (LET) radiation for radiotherapeuticapplications has gained increased interest. Astatine-211 (211At) is an -particle emittingradionuclide, promising for targeted radioimmunotherapy of isolated tumor cells andmicroscopic clusters. To improve development of safe radiotherapy using 211At it is importantto increase our knowledge of the radiobiological effects in cells. During radiotherapy, bothtumors and adjacent normal tissue will be irradiated and therefore, it is of importance tounderstand differences in the radioresponse between proliferating and resting cells. The aimof this thesis was to investigate effects in fibroblasts with different proliferation status afterirradiation with -particles from 211At or X-rays, from inflicted DNA damage, to cellularresponses and biological consequences.Throughout this work, irradiation was performed with -particles from 211A or X-rays. Theinduction and repair of double-strand breaks (DSBs) in human normal fibroblasts wereinvestigated using pulsed-field gel electrophoresis and fragment analysis. The relativebiological effectiveness (RBE) of 211At for DSB induction varied between 1.4 and 3.1. Asmall increase of DSBs was observed in cycling cells compared to stationary cells. The repairkinetics was slower after 211At and more residual damage was found after 24 h. Comparisonbetween cells with different proliferation status showed that the repair was inefficient incycling cells with more residual damage, regardless of radiation quality. Activation of cellcycle arrests was investigated using immunofluorescent labeling of the checkpoint kinaseChk2 and by measuring cell cycle distributions with flow cytometry analysis. After -particleirradiation, the average number of Chk2-foci was larger and the cells had a more affected cellcycle progression for several weeks compared with X-irradiated cells, indicating a morepowerful arrest after 211At. Flow cytometry showed that cycling cells were arrested in G2/Mwhile stationary cells underwent a delayed entry into S phase after release of contactinhibition. Radiation-induced chromosomal damage was studied by investigating theformation of micronuclei after first mitosis post-irradiation. Alpha-particles induced 2.7 and4.1 times more micronuclei in cycling and stationary cells, respectively, compared with Xrays.Induction of DSBs and cell survival after irradiation were also investigated in synchronizedChinese hamster fibroblasts. The cells were synchronized with mimosine in G1, early, midand late S phase and in mitosis and cell survival was determined using the clonogenic assay.The radioresponse between cell cycle phases varied after both 211At and X-rays, resulting invariations of RBE for 211At between 1.8 and 3.9 for DSB induction and between 3.1 and 7.9for 37% survival. The lowest RBE was observed in mitotic cells for both DSB induction andclonogenic survival.In summary, for all endpoints studied -particles from 211At were more detrimental comparedwith X-rays. Further, the radioresponse was dependent upon the proliferation status of thecells at the time of irradiation, after both low- and high-LET radiation, resulting in variationsof the relative biological effects.