Cellular irradiations with laser-driven carbon ions at ultra-high dose rates.

Objective.Carbon is an ion species of significant radiobiological interest, particularly in view of its use in cancer radiotherapy, where its large Relative Biological Efficiency is often exploited to overcome radio resistance. A growing interest in highly pulsed carbon delivery has arisen in the context of the development of the FLASH radiotherapy approach, with recent studies carried out at dose rates of 40 Gy s-1. Laser acceleration methods, producing ultrashort ion bursts, can now enable the delivery of Gy-level doses of carbon ions at ultra-high dose rates (UHDRs), exceeding 109Gy s-1. While studies at such extreme dose rate have been carried out so far using low LET particles such as electrons and protons, the radiobiology of high-LET, UHDR ions has not yet been explored. Here, we report the first application of laser-accelerated carbon ions generated by focussing 1020W cm-2intense lasers on 10-25 nm carbon targets, to irradiate radioresistant patient-derived Glioblastoma stem like cells (GSCs).Approach.We exposed GSCs to 1 Gy of 9.5 ± 0.5 MeV/n carbon ions delivered in a single ultra-short (∼400-picosecond) pulse, at a dose rate of 2 × 109Gy s-1, generated using the ASTRA GEMINI laser of the Central Laser Facility at the Rutherford Appleton Laboratory, Didcot, Oxfordshire, UK. We quantified carbon ion-induced DNA double strand break (DSB) damage using the 53BP1 foci formation assay and used 225 kVp x-rays as a reference radiation.Main Results.Laser-accelerated carbon ions induced complex DNA DSB damage, as seen through persistent 53BP1 foci (11.5 ± 0.4 foci/cell/Gy) at 24 h and significantly larger foci (1.69 ± 0.07µm2) than x-rays induced ones (0.63 ± 0.02µm2). The relative foci induction value for laser-driven carbon ions relative to conventional x-rays was 3.2 ± 0.3 at 24 h post-irradiation also confirming the complex nature of the induced damage.Significance.Our study demonstrates the feasibility of radiobiology investigations at unprecedented dose rates using laser-accelerated high-LET carbon ions in clinically relevant models.

Development of a portable hypoxia chamber for ultra-high dose rate laser-driven proton radiobiology applications.

BACKGROUND: There is currently significant interest in assessing the role of oxygen in the radiobiological effects at ultra-high dose rates. Oxygen modulation is postulated to play a role in the enhanced sparing effect observed in FLASH radiotherapy, where particles are delivered at 40-1000 Gy/s. Furthermore, the development of laser-driven accelerators now enables radiobiology experiments in extreme regimes where dose rates can exceed 109 Gy/s, and predicted oxygen depletion effects on cellular response can be tested. Access to appropriate experimental enviroments, allowing measurements under controlled oxygenation conditions, is a key requirement for these studies. We report on the development and application of a bespoke portable hypoxia chamber specifically designed for experiments employing laser-driven sources, but also suitable for comparator studies under FLASH and conventional irradiation conditions. MATERIALS AND METHODS: We used oxygen concentration measurements to test the induction of hypoxia and the maintenance capacity of the chambers. Cellular hypoxia induction was verified using hypoxia inducible factor-1α immunostaining. Calibrated radiochromic films and GEANT-4 simulations verified the dosimetry variations inside and outside the chambers. We irradiated hypoxic human skin fibroblasts (AG01522B) cells with laser-driven protons, conventional protons and reference 225 kVp X-rays to quantify DNA DSB damage and repair under hypoxia. We further measured the oxygen enhancement ratio for cell survival after X-ray exposure in normal fibroblast and radioresistant patient- derived GBM stem cells. RESULTS: Oxygen measurements showed that our chambers maintained a radiobiological hypoxic environment for at least 45 min and pathological hypoxia for up to 24 h after disconnecting the chambers from the gas supply. We observed a significant reduction in the 53BP1 foci induced by laser-driven protons, conventional protons and X-rays in the hypoxic cells compared to normoxic cells at 30 min post-irradiation. Under hypoxic irradiations, the Laser-driven protons induced significant residual DNA DSB damage in hypoxic AG01522B cells compared to the conventional dose rate protons suggesting an important impact of these extremely high dose-rate exposures. We obtained an oxygen enhancement ratio (OER) of 2.1 ± 0.1 and 2.5 ± 0.1 respectively for the AG01522B and patient-derived GBM stem cells for X-ray irradiation using our hypoxia chambers. CONCLUSION: We demonstrated the design and application of portable hypoxia chambers for studying cellular radiobiological endpoints after exposure to laser-driven protons at ultra-high dose, conventional protons and X-rays. Suitable levels of reduced oxygen concentration could be maintained in the absence of external gassing to quantify hypoxic effects. The data obtained provided indication of an enhanced residual DNA DSB damage under hypoxic conditions at ultra-high dose rate compared to the conventional protons or X-rays.