The biological effect of large single doses: a possible role for non-targeted effects in cell inactivation.
OBJECTIVE
Novel radiotherapy techniques increasingly use very large dose fractions. It has been argued that the biological effect of large dose fractions may differ from that of conventional fraction sizes. The purpose was to study the biological effect of large single doses.


METHODS
Clonogenic cell survival of MCF7 and MDA-MB-231 cells was determined after direct X-ray irradiation, irradiation of feeder cells, or transfer of conditioned medium (CM). Cell-cycle distributions and the apoptotic sub-G1 fraction were measured by flow cytometry. Cytokines in CM were quantified by a cytokine antibody array. γH2AX foci were detected by immunofluorescence microscopy.


RESULTS
The surviving fraction of MCF7 cells irradiated in vitro with 12 Gy showed an 8.5-fold decrease (95% c.i.: 4.4-16.3; P<0.0001) when the density of irradiated cells was increased from 10 to 50×10(3) cells per flask. Part of this effect was due to a dose-dependent transferrable factor as shown in CM experiments in the dose range 5-15 Gy. While no effect on apoptosis and cell cycle distribution was observed, and no differentially expressed cytokine could be identified, the transferable factor induced prolonged expression of γH2AX DNA repair foci at 1-12 h.


CONCLUSIONS
A dose-dependent non-targeted effect on clonogenic cell survival was found in the dose range 5-15 Gy. The dependence of SF on cell numbers at high doses would represent a "cohort effect" in vivo. These results support the hypothesis that non-targeted effects may contribute to the efficacy of very large dose fractions in radiotherapy.
Department of Radiation Oncology, Universitätsmedizin Mannheim, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany.
Marlon R
MR
Veldwijk
Department of Radiation Oncology, Universitätsmedizin Mannheim, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany.
Bo
B
Zhang
Department of Radiation Oncology, Universitätsmedizin Mannheim, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany.
Frederik
F
Wenz
Department of Radiation Oncology, Universitätsmedizin Mannheim, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany.
Carsten
C
Herskind
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects*; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D002453:Cell Cycle / Q000528:radiation effects*; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000528:radiation effects; D017077:Culture Media, Conditioned / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism*; D004307:Dose-Response Relationship, Radiation; D005260:Female; D005434:Flow Cytometry; D016193:G1 Phase / Q000528:radiation effects; D006657:Histones / Q000378:metabolism*; D006801:Humans; D061986:MCF-7 Cells; D008856:Microscopy, Fluorescence; D013997:Time Factors
D017077:Culture Media, Conditioned; D016207:Cytokines; C468783:H2AX protein, human; D006657:Histones