Out-of-Field Cell Survival Following Exposure to Intensity-Modulated Radiation Fields

Abstract

Purpose-To determine the in-field and out-of-field cell survival of cells irradiated with either primary field or scattered radiation in the presence and absence of intercellular communication.Methods and Materials-Cell survival was determined by clonogenic assay in human prostate cancer (DU145) and primary fibroblast (AGO1552) cells following exposure to different field configurations delivered using a 6-MV photon beam produced with a Varian linear accelerator.Results-Nonuniform dose distributions were delivered using a multileaf collimator (MLC) in which half of the cell population was shielded. Clonogenic survival in the shielded region was significantly lower than that predicted from the linear quadratic model. In both cell lines, the outof-field responses appeared to saturate at 40%-50% survival at a scattered dose of 0.70 Gy in DU-145 cells and 0.24 Gy in AGO1522 cells. There was an approximately eightfold difference in the initial slopes of the out-of-field response compared with the α-component of the uniform field response. In contrast, cells in the exposed part of the field showed increased survival. These observations were abrogated by direct physical inhibition of cellular communication and by the addition of the inducible nitric oxide synthase inhibitor aminoguanidine known to inhibit intercellular bystander effects. Additional studies showed the proportion of cells irradiated and dose delivered to the shielded and exposed regions of the field to impact on response.Conclusions-These data demonstrate out-of-field effects as important determinants of cell survival following exposure to modulated irradiation fields with cellular communication between differentially irradiated cell populations playing an important role. Validation of these observations in additional cell models may facilitate the refinement of existing radiobiological models and the observations considered important determinants of cell survival.

Bystander Signalling: Exploring Clinical Relevance Through New Approaches and New Models

“…These effects are cell signal mediated either through direct physical cell contact via gap junction intercellular communication (GJIC) (4) or through secreted, diffusible signalling molecules into the surrounding media (5)(6)(7). A number of candidate signalling molecules have been identified in mediating RIBEs such as reactive oxygen and nitrogen species (ROS/NOS) including nitric oxide (NO), and cytokines such as transforming growth factor-β (TGF-β) and interleukin-8 (IL-8).…”

“…This has recently been addressed in several studies which have delivered modulated 6 MV radiation fields using clinical linear accelerators to more accurately replicate exposure conditions in vivo (7,23,24). Work from our laboratory determined cell survival responses occurring in-and out-of-field using a modulated beam profile generated using a multi leaf collimator (MLC) and demonstrated significantly reduced out-offield cell survival following irradiation compared to the level of response predicted on scattered dose alone (7).…”

“…These results may have significant implications for the interpretation of a large portion of radiobiology data as they suggest the traditional paradigm of independent cellular responses may not be valid, with further consequences for the evaluation of many clinical conditions. Whilst classically defined bystander effects were interpreted as a low dose phenomenon, recent findings have highlighted the potential significance of radiation induced signalling in cohort and bystander effects following modulated dose distributions suggesting these may have a role in directly irradiated regions (7,23,24). Signalling from cells irradiated to high doses within the planning target volume (PTV) may contribute to increased tumour cell killing and improved outcomes.…”

Modelling responses to spatially fractionated radiation fields using preclinical image-guided radiotherapy

“…Using more simplistic approaches to generate non-uniform dose distributions in vitro, focused on the impact of steep dose gradients, our group has shown important contributions of non-targeted radiobiological effects driven by intercellular signalling, commonly known as bystander effects. 8,9 Although classically associated with low dose exposures, recent results have shown large contributions of signalling effects between cells exposed to high doses 10 which remain in contact for extended periods, [11][12][13][14] with reports of bystander cell killing of .50% and saturation doses .4 Gy. 13 These observations have been supported by mathematical modelling studies which suggest that intercellular communication may have a significant role in cell death even in populations exposed to doses up to several Gy.…”

“…8,9 Although classically associated with low dose exposures, recent results have shown large contributions of signalling effects between cells exposed to high doses 10 which remain in contact for extended periods, [11][12][13][14] with reports of bystander cell killing of .50% and saturation doses .4 Gy. 13 These observations have been supported by mathematical modelling studies which suggest that intercellular communication may have a significant role in cell death even in populations exposed to doses up to several Gy. [15][16][17][18] Further evidence for these effects has been obtained in small animal models, showing effects of cell signalling in vivo for both tumour 19 and normal tissue response.…”