Ionizing radiation induced phosphatidylserine externalization on endothelial cell surface - a potential target for vascular targeting

Background: Vascular targeting agents can deliver anti-cancer drugs specifically to tumor sites by binding to unique markers/targets on endothelial surface of tumors. The discriminating power of the markers/targets relative to normal tissues determines the specificity of the technique. Stereotactic RadioSurgery (SRS) can precisely deliver focused ionizing radiation to a target tumor site. Radiation-induced molecular changes should be restricted within the tumor tissue, and be good targets for vascular targeting. This study investigated radiation-induced externalization of phosphatidylserine (PS) in endothelial cells, a potential target for vascular targeting. Materials and Methods: An immortalized cell line generated from mouse brain endothelium, bEnd3 cells, were cultured and irradiated at different radiation doses using a linear accelerator (LINAC) Elekta Synergy. Then PS externalization in the cells was visualized using pSIVA-IANBD, a polarity sensitive probe for PS. Live cell imaging was used to monitor the PS externalization in real time. Results: Ionizing radiation has remarkable effects on the cells and the effects are found to be dose dependent. The cell proliferation rate decreased after exposure to 5 Gy radiation whereas higher radiation doses (15 Gy and 25 Gy) totally inhibited proliferation. In comparison with sham-radiation treated cells, the irradiated cells showed distinct pseudopodial elongation with little or no spreading of the cell body. The percentages of pSIVA positive cells were significantly higher in the cells that received 25 Gy and 15 Gy radiation 24 hours after treatment. This effect sustained until the end of the experiment (3 days). Radiation at 5 Gy did not induce significant PS externalization compared with the sham-radiation control at any time points. Conclusions: Ionizing radiation can cause remarkable cellular changes in the endothelial cells. Significant PS externalization can be induced by the radiation at dose levels 15 Gy and above. Given the precise focusing of the radiation beams, the radiation-induced markers/targets may have high discriminating power to be harnessed in vascular targeting for cancer treatment.