In vitroFLASH irradiation of A549 lung cancer cells and IMR90 healthy human fibroblasts in the synchrocyclotron room of a clinical proton therapy system

Abstract

Purpose: This study aims to investigate the FLASH irradiation effect on lung tumor (A549) and healthy fibroblast (IMR90) cell lines using an irradiation station installed at the synchrocyclotron room of a clinical proton facility. Methods and Materials: An irradiation system composed of a lead scatterer and 3D-printed positioning system was designed and fabricated to operate within the beamline gap of the IBA Proteus One proton therapy facility. A dosimetric analysis of the produced irradiation field was carried out using radiochromic films. FLASH and conventional-rate irradiations were conducted on relevant cell lines for lung cancer. Biological assessments post-irradiation included clonogenic and viability assays for cell survival, immunofluorescence analysis of p21 protein expression, and flow cytometry analysis for cell cycle arrest evaluation. Results: The irradiation system successfully delivered homogeneous and repeatable FLASH dose rates (>900 Gy/s) with a positioning accuracy of 1 mm and dose uniformity within 10%. Clonogenic assays revealed no statistically significant differences in survival between FLASH and conventional dose rates for both A549 and IMR90 cell lines, although a trend towards higher viability was observed in IMR90 cells under FLASH conditions. Flow cytometry demonstrated significant differences in cell cycle arrest patterns at doses above 8 Gy, with FLASH-irradiated cells exhibiting an increase in G0/G1 arrest and a decrease in G2/M phases compared to conventional rates. Immunofluorescence analysis of p21 expression showed no significant differences between irradiation modalities. Conclusions: The developed irradiation station effectively facilitates FLASH radiotherapy experiments in a clinical proton facility, achieving the necessary dose rates without costly hardware modification. Our analysis reported notable alterations in cell cycle dynamics suggesting distinct biological responses between FLASH and conventional rates in both healthy and tumor cells. These findings contribute to the emerging understanding of the FLASH effect and support the potential for its differential impact on cancerous versus healthy tissues.