Ponente
Descripción
Purpose
FLASH radiotherapy (RT) is a promising technique in radiotherapy, where ultra-high dose rates (>40 Gy/s) have demonstrated, in in-vitro and animal studies, a protective effect on healthy tissues, while maintaining isoefficacy with conventional (CONV) RT in treating tumors. We performed several biological essays with proton beams at FLASH vs CONV dose rates, using healthy lung fibroblasts and lung cancer cells to study potential biological mechanisms that could be at the root of the FLASH effect.
Materials and methods
Healthy lung fibroblasts (IMR90) and lung adenocarcinoma cells (A549) were cultured in DMEM supplemented with 10% FBS under standard culturing conditions. Before irradiation, 9.6M cells were collected and pellet in eppendorf vials and irradiated with proton beams at the IBA proteusOne in Quironsalud.
FLASH irradiations (>800 Gy/s average dose rate, dose range 0-12 Gy) with a 230-MeV beam were performed at the synchrocyclotron room of the facility using a portable 3D-printed system to degrade the beam and position the samples. CONV rate irradiations (<0.2 Gy/s, dose range 0-16 Gy) were performed in the treatment room using a SOBP (depth 2 cm, modulation 1 cm).
In-vitro clonogenic study (A549) and viability assay (IMR90) were performed by seeding 300 irradiated cells. 7 days later, cells were fixed, stained with crystal violet and number of colonies/viable cells quantified. Data was analyzed with the linear quadratic model. Irradiation-induced cell cycle arrest was studied via flow cytometry 48h post-radiation. Dose-dependent expression of p21 protein (associated with cell cycle arrest) was studied via immunofluorescence (IF) on irradiated cells fixed and permeabilized 48h after irradiation.
Results
Preliminary analysis of the results suggests little to no differential biological effects related with dose rate. Clonogenic assays for the A549 cells and viability study for the IMR90 cells showed no differential dose-rate effect in the biological response (Fig. 1). Cell cycle analysis via flow cytometry and IF showed G1 arrest rates and p21 induction probability both increase with dose for healthy and tumor cells, with no significant dose-rate dependence observed (Fig. 2).
Conclusions
The irradiation system, designed and fabricated in-house, made FLASH irradiation of biological samples possible in a clinical proton therapy center without any hardware or software system modifications. Equivalent biological results were obtained in an in-vitro study at normoxic conditions between conventional and FLASH dose rate protons for both healthy fibroblasts and lung cancer cells.
