In vitro studies suggested that sub-millisecond pulses of radiation elicit less genomic instability than continuous, protracted irradiation at the same total dose. To determine the potential of ultrahigh dose-rate irradiation in radiotherapy, we investigated lung fibrogenesis in C57BL/6J mice exposed either to short pulses (≤500 ms) of radiation delivered at ultrahigh dose rate (≥40 Gy/s, FLASH) or to conventional dose-rate irradiation (≤0.03 Gy/s, CONV) in single doses. The growth of human HBCx-12A and HEp-2 tumor xenografts in nude mice and syngeneic TC-1 Luc⁺ orthotopic lung tumors in C57BL/6J mice was monitored under similar radiation conditions. CONV (15 Gy) triggered lung fibrosis associated with activation of the TGF-β (transforming growth factor–β) cascade, whereas no complications developed after doses of FLASH below 20 Gy for more than 36 weeks after irradiation. FLASH irradiation also spared normal smooth muscle and epithelial cells from acute radiation-induced apoptosis, which could be reinduced by administration of systemic TNF-α (tumor necrosis factor–α) before irradiation. In contrast, FLASH was as efficient as CONV in the repression of tumor growth. Together, these results suggest that FLASH radiotherapy might allow complete eradication of lung tumors and reduce the occurrence and severity of early and late complications affecting normal tissue.

Purpose: One of the main limitations to anticancer radiotherapy lies in irreversible damage to healthy tissues located within the radiation field. “FLASH” irradiation at very high dose-rate is a new treatment modality that has been reported to specifically spare normal tissue from late radiation-induced toxicity in animal models and therefore could be a promising strategy to reduce treatment toxicity. Experimental Design: Lung responses to FLASH irradiation were investigated by qPCR, single-cell RNA sequencing (sc-RNA-Seq), and histologic methods during the acute wound healing phase as well as at late stages using C57BL/6J wild-type and Terc−/− mice exposed to bilateral thorax irradiation as well as human lung cells grown in vitro. Results: In vitro studies gave evidence of a reduced level of DNA damage and induced lethality at the advantage of FLASH. In mouse lung, sc-RNA-seq and the monitoring of proliferating cells revealed that FLASH minimized the induction of proinflammatory genes and reduced the proliferation of progenitor cells after injury. At late stages, FLASH-irradiated lungs presented less persistent DNA damage and senescent cells than after CONV exposure, suggesting a higher potential for lung regeneration with FLASH. Consistent with this hypothesis, the beneficial effect of FLASH was lost in Terc−/− mice harboring critically short telomeres and lack of telomerase activity. Conclusions: The results suggest that, compared with conventional radiotherapy, FLASH minimizes DNA damage in normal cells, spares lung progenitor cells from excessive damage, and reduces the risk of replicative senescence.

FLASH radiotherapy, a technique based on delivering large doses in a single fraction at the micro/millisecond timescale, spares normal tissues from late radiation-induced toxicity, in an oxygen-dependent process, whilst keeping full anti-tumor efficiency. We present a theoretical model taking into account the kinetics of formation and decay of reactive oxygen species, in particular of organic peroxyl radicals ROO. formed by addition of O2 to primary carbon-centred radicals R. and known to play a major role at the origin radio-induced complications. The model focuses on the time-dependent evolution of radiolytic products in living matter exposed to continuous irradiation at dose-rates in the range 10-3-107Gy·s-1. The 9 differential rate equations resulting from the radiolytic and enzymatic reactions network were solved using the published values of these reactions rate constants in a cellular environment. The model suggests a correlation between the area-under-the-curve of time-evolving [ROO.] and the probability of normal tissue complications. The model does not lend weight to the hypothesis of transient oxygen depletion as a main determinant of FLASH but rather suggests a major role of radical-radical recombination. The model gives support to the reduction of ROO. lifetime as the main root of FLASH and compares favorably with published experimental results. We conclude that any process - in this case radical recombination - that shortens the lifetime or limits the radiolytic yield of ROO. is likely to protect normoxic tissues against the deleterious effects of radiation.

The consequences of PARP-1 disruption or inhibition on DNA single-strand break repair (SSBR) and radio-induced lethality were determined in synchronized, isogenic HeLa cells stably silenced or not for poly(ADP-ribose) polymerase-1 (PARP-1) (PARP-1 ^KD ) or XRCC1 (XRCC1 ^KD ). PARP-1 inhibition prevented XRCC1-YFP recruitment at sites of 405 nm laser micro irradiation, slowed SSBR 10-fold and triggered the accumulation of large persistent foci of GFP-PARP-1 and GFP-PCNA at photo damaged sites. These aggregates are presumed to hinder the recruitment of other effectors of the base excision repair (BER) pathway. PARP-1 silencing also prevented XRCC1-YFP recruitment but did not lengthen the lifetime of GFP-PCNA foci. Moreover, PARP-1 ^KD and XRCC1 ^KD cells in S phase completed SSBR as rapidly as controls, while SSBR was delayed in G1. Taken together, the data demonstrate that a PARP-1- and XRCC1-independent SSBR pathway operates when the short patch repair branch of the BER is deficient. Long patch repair is the likely mechanism, as GFP-PCNA recruitment at photo-damaged sites was normal in PARP-1 ^KD cells. PARP-1 silencing elicited hyper-radiosensitivity, while radiosensitization by a PARP inhibitor reportedly occurs only in those cells treated in S phase. PARP-1 inhibition and deletion thus have different outcomes in terms of SSBR and radiosensitivity.

The development of innovative approaches that would reduce the sensitivity of healthy tissues to irradiation while maintaining the efficacy of the treatment on the tumor is of crucial importance for the progress of the efficacy of radiotherapy. Recent methodological developments and innovations, such as scanned beams, ultra-high dose rates, and very high-energy electrons, which may be simultaneously available on new accelerators, would allow for possible radiobiological advantages of very short pulses of ultra-high dose rate (FLASH) therapy for radiation therapy to be considered. In particular, very high-energy electron (VHEE) radiotherapy, in the energy range of 100 to 250 MeV, first proposed in the 2000s, would be particularly interesting both from a ballistic and biological point of view for the establishment of this new type of irradiation technique. In this review, we examine and summarize the current knowledge on VHEE radiotherapy and provide a synthesis of the studies that have been published on various experimental and simulation works. We will also consider the potential for VHEE therapy to be translated into clinical contexts.