Comet Assay Profiling of FLASH-Induced Damage: Mechanistic Insights into the Effects of FLASH Irradiation.

Numerous studies have demonstrated the normal tissue-sparing effects of ultra-high dose rate 'FLASH' irradiation in vivo, with an associated reduction in damage burden being reported in vitro. Towards this, two key radiochemical mechanisms have been proposed: radical-radical recombination (RRR) and transient oxygen depletion (TOD), with both being proposed to lead to reduced levels of induced damage. Previously, we reported that FLASH induces lower levels of DNA strand break damage in whole-blood peripheral blood lymphocytes (WB-PBL) ex vivo, but our study failed to distinguish the mechanism(s) involved. A potential outcome of RRR is the formation of crosslink damage (particularly, if any organic radicals recombine), whilst a possible outcome of TOD is a more anoxic profile of induced damage resulting from FLASH. Therefore, the aim of the current study was to profile FLASH-induced damage via the Comet assay, assessing any DNA crosslink formation as a putative marker of RRR and/or anoxic DNA damage formation as an indicative marker of TOD, to determine the extent to which either mechanism contributes to the "FLASH effect". Following FLASH irradiation, we see no evidence of any crosslink formation; however, FLASH irradiation induces a more anoxic profile of induced damage, supporting the TOD mechanism. Furthermore, treatment of WB-PBLs pre-irradiation with BSO abrogates the reduced strand break damage burden mediated by FLASH exposures. In summary, we do not see any experimental evidence to support the RRR mechanism contributing to the reduced damage burden induced by FLASH. However, the observation of a greater anoxic profile of damage following FLASH irradiation, together with the BSO abrogation of the reduced strand break damage burden mediated by FLASH, lends further support to TOD being a driver of the reduced damage burden plus a change in the damage profile mediated by FLASH.

In vitro assays for investigating the FLASH effect.

FLASH radiotherapy is a novel technique that has been shown in numerous preclinical in vivo studies to have the potential to be the next important improvement in cancer treatment. However, the biological mechanisms responsible for the selective FLASH sparing effect of normal tissues are not yet known. An optimal translation of FLASH radiotherapy into the clinic would require a good understanding of the specific beam parameters that induces a FLASH effect, environmental conditions affecting the response, and the radiobiological mechanisms involved. Even though the FLASH effect has generally been considered as an in vivo effect, studies finding these answers would be difficult and ethically challenging to carry out solely in animals. Hence, suitable in vitro studies aimed towards finding these answers are needed. In this review, we describe and summarise several in vitro assays that have been used or could be used to finally elucidate the mechanisms behind the FLASH effect.

Long-term neurocognitive benefits of FLASH radiotherapy driven by reduced reactive oxygen species.

Here, we highlight the potential translational benefits of delivering FLASH radiotherapy using ultra-high dose rates (>100 Gy⋅s-1). Compared with conventional dose-rate (CONV; 0.07-0.1 Gy⋅s-1) modalities, we showed that FLASH did not cause radiation-induced deficits in learning and memory in mice. Moreover, 6 months after exposure, CONV caused permanent alterations in neurocognitive end points, whereas FLASH did not induce behaviors characteristic of anxiety and depression and did not impair extinction memory. Mechanistic investigations showed that increasing the oxygen tension in the brain through carbogen breathing reversed the neuroprotective effects of FLASH, while radiochemical studies confirmed that FLASH produced lower levels of the toxic reactive oxygen species hydrogen peroxide. In addition, FLASH did not induce neuroinflammation, a process described as oxidative stress-dependent, and was also associated with a marked preservation of neuronal morphology and dendritic spine density. The remarkable normal tissue sparing afforded by FLASH may someday provide heretofore unrealized opportunities for dose escalation to the tumor bed, capabilities that promise to hasten the translation of this groundbreaking irradiation modality into clinical practice.

Palliative short-course hypofractionated radiotherapy followed by chemotherapy in esophageal adenocarcinoma: the phase II PALAESTRA trial.

Background: The majority of patients with incurable esophageal adenocarcinoma suffer from dysphagia. We assessed a novel treatment strategy with initial short-course radiotherapy followed by chemotherapy with the primary aim to achieve long-term relief of dysphagia.Methods: This phase II trial included treatment-naîve patients with dysphagia due to esophageal adenocarcinoma not eligible for curative treatment. External beam radiotherapy with 20 Gy in five fractions to the primary tumor was followed by four cycles of chemotherapy (FOLFOX regimen). Dysphagia was assessed using a five-grade scale.Results: From October 2014 to May 2018 a total of 29 patients were enrolled. The rate of dysphagia improvement was 79%, median duration of improvement 6.7 months (12.2 months for responders) and median overall survival 9.9 months. In the pre-specified per protocol analysis (23 patients) the rate of dysphagia improvement was 91%, median duration of improvement 12.2 months (14.0 months for responders) and median overall survival 16.0 months. The most common grade 3-4 adverse events were neutropenia (29%), infection (25%), anorexia (11%), esophagitis (11%) and fatigue (11%).Conclusion: Initial palliative short-course radiotherapy followed by chemotherapy is a promising treatment strategy that can provide long-lasting relief of dysphagia in patients with esophageal adenocarcinoma.

Feasibility and constraints of Bragg peak FLASH proton therapy treatment planning.

Introduction: FLASH proton therapy (FLASH-PT) requires ultra-high dose rate (≥ 40 Gy/s) protons to be delivered in a short timescale whilst conforming to a patient-specific target. This study investigates the feasibility and constraints of Bragg peak FLASH-PT treatment planning, and compares the in silico results produced to plans for intensity modulated proton therapy (IMPT). Materials and method: Bragg peak FLASH-PT and IMPT treatment plans were generated for bone (n=3), brain (n=3), and lung (n=4) targets using the MIROpt research treatment planning system and the Conformal FLASH library developed by Applications SA from the open-source version of UCLouvain. FLASH-PT beams were simulated using monoenergetic spot-scanned protons traversing through a conformal energy modulator, a range shifter, and an aperture. A dose rate constraint of ≥ 40 Gy/s was included in each FLASH-PT plan optimisation. Results: Space limitations in the FLASH-PT adapted beam nozzle imposed a maximum target width constraint, excluding 4 cases from the study. FLASH-PT plans did not satisfy the imposed target dose constraints (D95% ≥ 95% and D2%≤ 105%) but achieved clinically acceptable doses to organs at risk (OARs). IMPT plans adhered to all target and OAR dose constraints. FLASH-PT plans showed a reduction in both target homogeneity (p < 0.001) and dose conformity (non-significant) compared to IMPT. Conclusion: Without accounting for a sparing effect, IMPT plans were superior in target coverage, dose conformity, target homogeneity, and OAR sparing compared to FLASH-PT. Further research is warranted in treatment planning optimisation and beam delivery for clinical implementation of Bragg peak FLASH-PT.

Beam control system and output fine-tuning for safe and precise delivery of FLASH radiotherapy at a clinical linear accelerator.

Introduction: We have previously adapted a clinical linear accelerator (Elekta Precise, Elekta AB) for ultra-high dose rate (UHDR) electron delivery. To enhance reliability in future clinical FLASH radiotherapy trials, the aim of this study was to introduce and evaluate an upgraded beam control system and beam tuning process for safe and precise UHDR delivery. Materials and Methods: The beam control system is designed to interrupt the beam based on 1) a preset number of monitor units (MUs) measured by a monitor detector, 2) a preset number of pulses measured by a pulse-counting diode, or 3) a preset delivery time. For UHDR delivery, an optocoupler facilitates external control of the accelerator's thyratron trigger pulses. A beam tuning process was established to maximize the output. We assessed the stability of the delivery, and the independent interruption capabilities of the three systems (monitor detector, pulse counter, and timer). Additionally, we explored a novel approach to enhance dosimetric precision in the delivery by synchronizing the trigger pulse with the charging cycle of the pulse forming network (PFN). Results: Improved beam tuning of gun current and magnetron frequency resulted in average dose rates at the dose maximum at isocenter distance of >160 Gy/s or >200 Gy/s, with or without an external monitor chamber in the beam path, respectively. The delivery showed a good repeatability (standard deviation (SD) in total film dose of 2.2%) and reproducibility (SD in film dose of 2.6%). The estimated variation in DPP resulted in an SD of 1.7%. The output in the initial pulse depended on the PFN delay time. Over the course of 50 measurements employing PFN synchronization, the absolute percentage error between the delivered number of MUs calculated by the monitor detector and the preset MUs was 0.8 ± 0.6% (mean ± SD). Conclusion: We present an upgraded beam control system and beam tuning process for safe and stable UHDR electron delivery of hundreds of Gy/s at isocenter distance at a clinical linac. The system can interrupt the beam based on monitor units and utilize PFN synchronization for improved dosimetric precision in the dose delivery, representing an important advancement toward reliable clinical FLASH trials.

Long-term toxicity and efficacy of FLASH radiotherapy in dogs with superficial malignant tumors.

Introduction: FLASH radiotherapy (RT) has emerged as a promising modality, demonstrating both a normal tissue sparing effect and anticancer efficacy. We have previously reported on the safety and efficacy of single fraction FLASH RT in the treatment of oral tumors in canine cancer patients, showing tumor response but also a risk of radiation-induced severe late adverse effects (osteoradionecrosis) for doses ≥35 Gy. Accordingly, the objective in this study was to investigate if single fraction high dose FLASH RT is safe for treating non-oral tumors. Methods: Privately-owned dogs with superficial tumors or microscopic residual disease were included. Treatment was generally delivered as a single fraction of 15-35 Gy 10 MeV electron FLASH RT, although two dogs were re-irradiated at a later timepoint. Follow-up visits were conducted up to 12 months post-treatment to evaluate treatment efficiency and adverse effects. Results: Fourteen dogs with 16 tumors were included, of which nine tumors were treated for gross disease whilst seven tumors were treated post-surgery for microscopic residual disease. Four treatment sites treated with 35 Gy had ulceration post irradiation, which was graded as severe adverse effect. Only mild adverse effects were observed for the remaining treatment sites. None of the patients with microscopic disease experienced recurrence (0/7), and all patients with macroscopic disease showed either a complete (5/9) or a partial response (4/9). Five dogs were euthanized due to clinical disease progression. Discussion: Our study demonstrates that single fraction high dose FLASH RT is generally safe, with few severe adverse effects, particularly in areas less susceptible to radiation-induced damage. In addition, our study indicates that FLASH has anti-tumor efficacy in a clinical setting. No osteoradionecrosis was observed in this study, although other types of high-grade adverse effects including ulcer-formations were observed for the highest delivered dose (35 Gy). Overall, we conclude that osteoradionecrosis following single fraction, high dose FLASH does not appear to be a general problem for non-oral tumor locations. Also, as has been shown previously for oral tumors, 30 Gy appeared to be the maximum safe dose to deliver with single fraction FLASH RT.

Reconfiguring a Plane-Parallel Transmission Ionization Chamber to Extend the Operating Range into the Ultra-High Dose-per-pulse Regime.

This study aims to investigate the feasibility of enhancing the charge collection efficiency (CCE) of a transmission chamber by reconfiguring its design and operation. The goal was to extend the range of dose-per-pulse (DPP) values with no or minimal recombination effects up to the ultra-high dose rate (UHDR) regime. The response of two transmission chambers, with electrode distance of 1 mm and 0.6 mm, respectively, was investigated as a function of applied voltage. The chambers were mounted one-by-one in the electron applicator of a 10 MeV FLASH-modified clinical linear accelerator. The chamber signals were measured as a function of nominal DPP, which was determined at the depth of dose maximum using EBT-XD film in solid water and ranged from 0.6 mGy per pulse to 0.9 Gy per pulse, for both the standard voltage of 320 V and the highest possible safe voltage of 1,200 V. The CCE was calculated and fitted with an empirical logistic function that incorporated the electrode distance and the chamber voltage. The CCE decreased with increased DPP. The CCE at the highest achievable DPP was 24% (36%) at 320 V and 51% (82%) at 1,200 V, for chambers with 1 mm (0.6 mm) electrode distance. For the combination of 1,200 V- and 0.6-mm electrode distance, the CCE was ∼100% for average dose rate up to 70 Gy/s at the depth of dose maximum in the phantom at a source-to-surface distance of 100 cm. Our findings indicate that minor modifications to a plane-parallel transmission chamber can substantially enhance the CCE and extending the chamber's operating range to the UHDR regime. This supports the potential of using transmission chamber-based monitoring solutions for UHDR beams, which could facilitate the delivery of UHDR treatments using an approach similar to conventional clinical delivery.

The AsiDNA™ decoy mimicking DSBs protects the normal tissue from radiation toxicity through a DNA-PK/p53/p21-dependent G1/S arrest.

AsiDNA™, a cholesterol-coupled oligonucleotide mimicking double-stranded DNA breaks, was developed to sensitize tumour cells to radio- and chemotherapy. This drug acts as a decoy hijacking the DNA damage response. Previous studies have demonstrated that standalone AsiDNA™ administration is well tolerated with no additional adverse effects when combined with chemo- and/or radiotherapy. The lack of normal tissue complication encouraged further examination into the role of AsiDNA™ in normal cells. This research demonstrates the radioprotective properties of AsiDNA™. In vitro, AsiDNA™ induces a DNA-PK/p53/p21-dependent G1/S arrest in normal epithelial cells and fibroblasts that is absent in p53 deficient and proficient tumour cells. This cell cycle arrest improved survival after irradiation only in p53 proficient normal cells. Combined administration of AsiDNA™ with conventional radiotherapy in mouse models of late and early radiation toxicity resulted in decreased onset of lung fibrosis and increased intestinal crypt survival. Similar results were observed following FLASH radiotherapy in standalone or combined with AsiDNA™. Mechanisms comparable to those identified in vitro were detected both in vivo, in the intestine and ex vivo, in precision cut lung slices. Collectively, the results suggest that AsiDNA™ can partially protect healthy tissues from radiation toxicity by triggering a G1/S arrest in normal cells.

Treatment plan comparison using grading analysis based on clinical judgment.

PURPOSE: In this work we explore a method named clinical grading analysis (CGA) which is based on clinical assessments performed by radiation oncologists (ROs). The purpose is to investigate how useful the method is for treatment plan comparisons, and how the CGA results correlate with dosimetric evaluation parameters, traditionally used for treatment plan comparisons. MATERIAL AND METHODS: Helical tomotherapy (HTT) and seven-beam step-and-shoot intensity modulated radiation therapy (SS-IMRT) plans were compared and assessed by 10 experienced ROs for 23 patient cases. A CGA was performed where the plans were graded based on how the ROs thought they compared to each other. The resulting grades from the CGA were analyzed and compared to dose-volume statistics and equivalent uniform dose (EUD) data. RESULTS: For eight of the 23 cases the CGA revealed a significant difference between the HTT and the SS-IMRT plans, five cases were in favor of HTT, and three in favor of SS-IMRT. Comparing the dose-volume statistics and EUD-data with the result from the CGA showed that CGA results correlated well with dose-volume statistics for cases regarding difference in target coverage or doses to organs at risk. The CGA results also correlated well with EUD-data for cases with difference in clinical target volume (CTV) coverage but the correlation for cases with difference in planning target volume (PTV) coverage was not as clear. CONCLUSIONS: This study presents CGA as a useful method of comparing radiotherapy treatment plans. The proposed method offers a formalized way of introducing and evaluating the implementation of new radiotherapy techniques in a clinical setting. The CGA identify patients that have a clinical benefit of one or the other of the advanced treatment techniques available to them, i.e. in this study HTT and SS-IMRT, which facilitates a more optimal use of a clinics' advanced treatment resources.

Comparable survival in rats with intracranial glioblastoma irradiated with single-fraction conventional radiotherapy or FLASH radiotherapy.

Background: Radiotherapy increases survival in patients with glioblastoma. However, the prescribed dose is limited by unwanted side effects on normal tissue. Previous experimental studies have shown that FLASH radiotherapy (FLASH-RT) can reduce these side effects. Still, it is important to establish an equal anti-tumor efficacy comparing FLASH-RT to conventional radiotherapy (CONV-RT). Methods: Fully immunocompetent Fischer 344 rats with the GFP-positive NS1 intracranial glioblastoma model were irradiated with CONV-RT or FLASH-RT in one fraction of 20 Gy, 25 Gy or 30 Gy. Animals were monitored for survival and acute dermal side effects. The brains were harvested upon euthanasia and tumors were examined post mortem. Results: Survival was significantly increased in animals irradiated with CONV-RT and FLASH-RT at 20 Gy and 25 Gy compared to control animals. The longest survival was reached in animals irradiated with FLASH-RT and CONV-RT at 25 Gy. Irradiation at 30 Gy did not lead to increased survival, despite smaller tumors. Tumor size correlated inversely with irradiation dose, both in animals treated with CONV-RT and FLASH-RT. Acute dermal side effects were mild, but only a small proportion of the animals were alive for evaluation of those side effects. Conclusion: The dose response was similar for CONV-RT and FLASH-RT in the present model. Tumor size upon the time of euthanasia correlated inversely with the irradiation dose.

Evaluation of intensity-modulated electron FLASH radiotherapy in a clinical setting using veterinary cases.

PURPOSE: The increased normal tissue tolerance for FLASH radiotherapy (FLASH-RT), as compared to conventional radiotherapy, was first observed in ultra-high dose rate electron beams. Initial clinical trials in companion animals have revealed a high risk of developing osteoradionecrosis following high-dose single-fraction electron FLASH-RT, which may be related to inhomogeneities in the dose distribution. In the current study, we aim to evaluate the possibilities of intensity-modulated electron FLASH-RT in a clinical setting to ensure a homogeneous dose distribution in future veterinary and human clinical trials. METHODS: Our beam model in the treatment planning system electronRT (.decimal, LLC, Sanford, FL, USA) was based on a 10-MeV electron beam from a clinical linear accelerator used to treat veterinary patients with FLASH-RT in a clinical setting. In electronRT, the beam can be intensity-modulated using tungsten island blocks in the electron block cutout, and range-modulated using a customized bolus with variable thickness. Modulations were first validated in a heterogeneous phantom by comparing measured and calculated dose distributions. To evaluate the impact of intensity modulation in superficial single-fraction FLASH-RT, a treatment planning study was conducted, including eight canine cancer patient cases with simulated tumors in the head-and-neck region. For each case, treatment plans with and without intensity modulation were created for a uniform bolus and a range-modulating bolus. Treatment plans were evaluated using a target dose homogeneity index (HI), a conformity index (CI), the near-maximum dose outside the target ( D 2 % , Body - PTV ${D_{2{\mathrm{\% }},{\mathrm{\ Body}} - {\mathrm{PTV}}}}$ ), and the near-minimum dose to the target ( D 98 % ${D_{98\% }}$ ). RESULTS: By adding intensity modulation to plans with a uniform bolus, the HI could be improved (p = 0.017). The combination of a range-modulating bolus and intensity modulation provided a further significant improvement of the HI as compared to using intensity modulation in combination with a uniform bolus (p = 0.036). The range-modulating bolus also improved the CI compared to using a uniform bolus, both with an open beam (p = 0.046) and with intensity modulation (p = 0.018), as well as increased the D 98 % ${D_{98\% }}$ (p = 0.036 with open beam and p = 0.05 with intensity modulation) and reduced the median D 2 % , Body - PTV ${D_{2\% ,{\mathrm{\ Body}} - {\mathrm{PTV}}}}$ (not significant). CONCLUSIONS: By using intensity-modulated electron FLASH-RT in combination with range-modulating bolus, the target dose homogeneity and conformity in canine patients with simulated tumors in complex areas in the head-and-neck region could be improved. By utilizing this technique, we hope to decrease the dose outside the target volume and avoid hot spots in future clinical electron FLASH-RT studies, thereby reducing the risk of radiation-induced toxicity.

Effect of Conventional and Ultrahigh Dose Rate FLASH Irradiations on Preclinical Tumor Models: A Systematic Analysis.

PURPOSE: Compared with conventional dose rate irradiation (CONV), ultrahigh dose rate irradiation (UHDR) has shown superior normal tissue sparing. However, a clinically relevant widening of the therapeutic window by UHDR, termed "FLASH effect," also depends on the tumor toxicity obtained by UHDR. Based on a combined analysis of published literature, the current study examined the hypothesis of tumor isoefficacy for UHDR versus CONV and aimed to identify potential knowledge gaps to inspire future in vivo studies. METHODS AND MATERIALS: A systematic literature search identified publications assessing in vivo tumor responses comparing UHDR and CONV. Qualitative and quantitative analyses were performed, including combined analyses of tumor growth and survival data. RESULTS: We identified 66 data sets from 15 publications that compared UHDR and CONV for tumor efficacy. The median number of animals per group was 9 (range 3-15) and the median follow-up period was 30.5 days (range 11-230) after the first irradiation. Tumor growth assays were the predominant model used. Combined statistical analyses of tumor growth and survival data are consistent with UHDR isoefficacy compared with CONV. Only 1 study determined tumor-controlling dose (TCD50) and reported statistically nonsignificant differences. CONCLUSIONS: The combined quantitative analyses of tumor responses support the assumption of UHDR isoefficacy compared with CONV. However, the comparisons are primarily based on heterogeneous tumor growth assays with limited numbers of animals and short follow-up, and most studies do not assess long-term tumor control probability. Therefore, the assays may be insensitive in resolving smaller response differences, such as responses of radioresistant tumor subclones. Hence, tumor cure experiments, including additional TCD50 experiments, are needed to confirm the assumption of isoeffectiveness in curative settings.

Evaluation of single-fraction high dose FLASH radiotherapy in a cohort of canine oral cancer patients.

Background: FLASH radiotherapy (RT) is a novel method for delivering ionizing radiation, which has been shown in preclinical studies to have a normal tissue sparing effect and to maintain anticancer efficacy as compared to conventional RT. Treatment of head and neck tumors with conventional RT is commonly associated with severe toxicity, hence the normal tissue sparing effect of FLASH RT potentially makes it especially advantageous for treating oral tumors. In this work, the objective was to study the adverse effects of dogs with spontaneous oral tumors treated with FLASH RT. Methods: Privately-owned dogs with macroscopic malignant tumors of the oral cavity were treated with a single fraction of ≥30Gy electron FLASH RT and subsequently followed for 12 months. A modified conventional linear accelerator was used to deliver the FLASH RT. Results: Eleven dogs were enrolled in this prospective study. High grade adverse effects were common, especially if bone was included in the treatment field. Four out of six dogs, who had bone in their treatment field and lived at least 5 months after RT, developed osteoradionecrosis at 3-12 months post treatment. The treatment was overall effective with 8/11 complete clinical responses and 3/11 partial responses. Conclusion: This study shows that single-fraction high dose FLASH RT was generally effective in this mixed group of malignant oral tumors, but the risk of osteoradionecrosis is a serious clinical concern. It is possible that the risk of osteonecrosis can be mitigated through fractionation and improved dose conformity, which needs to be addressed before moving forward with clinical trials in human cancer patients.

Framework for Quality Assurance of Ultrahigh Dose Rate Clinical Trials Investigating FLASH Effects and Current Technology Gaps.

FLASH radiation therapy (FLASH-RT), delivered with ultrahigh dose rate (UHDR), may allow patients to be treated with less normal tissue toxicity for a given tumor dose compared with currently used conventional dose rate. Clinical trials are being carried out and are needed to test whether this improved therapeutic ratio can be achieved clinically. During the clinical trials, quality assurance and credentialing of equipment and participating sites, particularly pertaining to UHDR-specific aspects, will be crucial for the validity of the outcomes of such trials. This report represents an initial framework proposed by the NRG Oncology Center for Innovation in Radiation Oncology FLASH working group on quality assurance of potential UHDR clinical trials and reviews current technology gaps to overcome. An important but separate consideration is the appropriate design of trials to most effectively answer clinical and scientific questions about FLASH. This paper begins with an overview of UHDR RT delivery methods. UHDR beam delivery parameters are then covered, with a focus on electron and proton modalities. The definition and control of safe UHDR beam delivery and current and needed dosimetry technologies are reviewed and discussed. System and site credentialing for large, multi-institution trials are reviewed. Quality assurance is then discussed, and new requirements are presented for treatment system standard analysis, patient positioning, and treatment planning. The tables and figures in this paper are meant to serve as reference points as we move toward FLASH-RT clinical trial performance. Some major questions regarding FLASH-RT are discussed, and next steps in this field are proposed. FLASH-RT has potential but is associated with significant risks and complexities. We need to redefine optimization to focus not only on the dose but also on the dose rate in a manner that is robust and understandable and that can be prescribed, validated, and confirmed in real time. Robust patient safety systems and access to treatment data will be critical as FLASH-RT moves into the clinical trials.

The importance of hypoxia in radiotherapy for the immune response, metastatic potential and FLASH-RT.

PURPOSE: Hypoxia (low oxygen) is a common feature of solid tumors that has been intensely studied for more than six decades. Here we review the importance of hypoxia to radiotherapy with a particular focus on the contribution of hypoxia to immune responses, metastatic potential and FLASH radiotherapy, active areas of research by leading women in the field. CONCLUSION: Although hypoxia-driven metastasis and immunosuppression can negatively impact clinical outcome, understanding these processes can also provide tumor-specific vulnerabilities that may be therapeutically exploited. The different oxygen tensions present in tumors and normal tissues may underpin the beneficial FLASH sparing effect seen in normal tissue and represents a perfect example of advances in the field that can leverage tumor hypoxia to improve future radiotherapy treatments.

FLASH irradiation induces lower levels of DNA damage ex vivo, an effect modulated by oxygen tension, dose, and dose rate.

OBJECTIVE: FLASH irradiation reportedly produces less normal tissue toxicity, while maintaining tumour response. To investigate oxygen's role in the 'FLASH effect', we assessed DNA damage levels following irradiation at different oxygen tensions, doses and dose rates. METHODS: Samples of whole blood were irradiated (20 Gy) at various oxygen tensions (0.25-21%) with 6 MeV electrons at dose rates of either 2 kGy/s (FLASH) or 0.1 Gy/s (CONV), and subsequently with various doses (0-40 Gy) and intermediate dose rates (0.3-1000 Gy/s). DNA damage of peripheral blood lymphocytes (PBL) were assessed by the alkaline comet assay. RESULTS: Following 20 Gy irradiation, lower levels of DNA damage were induced for FLASH, the difference being significant at 0.25% (p < 0.05) and 0.5% O2 (p < 0.01). The differential in DNA damage at 0.5% O2 was found to increase with total dose and dose rate, becoming significant for doses ≥20 Gy and dose rates ≥30 Gy/s. CONCLUSION: This study shows, using the alkaline comet assay, that lower levels of DNA damage are induced following FLASH irradiation, an effect that is modulated by the oxygen tension, and increases with the total dose and dose rate of irradiation, indicating that an oxygen related mechanism, e.g. transient radiation-induced oxygen depletion, may contribute to the tissue sparing effect of FLASH irradiation. ADVANCES IN KNOWLEDGE: This paper is first to directly show that FLASH-induced DNA damage is modulated by oxygen tension, total dose and dose rate, with FLASH inducing significantly lower levels of DNA damage for doses ≥20 Gy and dose rates ≥30 Gy/s, at 0.5% O2.

Development of Ultra-High Dose-Rate (FLASH) Particle Therapy.

Research efforts in FLASH radiotherapy have increased at an accelerated pace recently. FLASH radiotherapy involves ultra-high dose rates and has shown to reduce toxicity to normal tissue while maintaining tumor response in pre-clinical studies when compared to conventional dose rate radiotherapy. The goal of this review is to summarize the studies performed to-date with proton, electron, and heavy ion FLASH radiotherapy, with particular emphasis on the physical aspects of each study and the advantages and disadvantages of each modality. Beam delivery parameters, experimental set-up, and the dosimetry tools used are described for each FLASH modality. In addition, modeling efforts and treatment planning for FLASH radiotherapy is discussed along with potential drawbacks when translated into the clinical setting. The final section concludes with further questions that have yet to be answered before safe clinical implementation of FLASH radiotherapy.

Comparable Long-Term Tumor Control for Hypofractionated FLASH Versus Conventional Radiation Therapy in an Immunocompetent Rat Glioma Model.

Purpose: To ensure a clinical translation of FLASH radiation therapy (FLASH-RT) for a specific tumor type, studies on tumor control and toxicity within the same biological system are needed. In this study, our objective was to evaluate tumor control and toxicity for hypofractionated FLASH-RT and conventional radiation therapy (CONV-RT) in an immunocompetent rat glioma model. Methods and Materials: Fisher 344 rats (N = 68) were inoculated subcutaneously with NS1 glioma cells and randomized into groups (n = 9-10 per group). CONV-RT (∼8 Gy/min) or FLASH-RT (70-90 Gy/s) was administered in 3 fractions of either 8 Gy, 12.5 Gy, or 15 Gy using a 10-MeV electron beam. The maximum tumor diameter was measured weekly, and overall survival was determined until day 100. Long-term tumor control was defined as no evident tumor on day 100. Animals were evaluated for acute dermal side effects at 2 to 5 weeks after completed RT and for late dermal side effects at 3 months after initiation of treatment. Results: Survival was significantly increased in all irradiated groups compared with control animals (P < .001). In general, irradiated tumors started to shrink at 1 week post-completed RT. In 40% (23 of 58) of the irradiated animals, long-term tumor control was achieved. Radiation-induced skin toxic effects were mild and consisted of hair loss, erythema, and dry desquamation. No severe toxic effect was observed. There was no significant difference between FLASH-RT and CONV-RT in overall survival, acute side effects, or late side effects for any of the dose levels. Conclusions: This study shows that hypofractionated FLASH-RT results in long-term tumor control rates similar to those of CONV-RT for the treatment of large subcutaneous glioblastomas in immunocompetent rats. Neither treatment technique induced severe skin toxic effects. Consequently, no significant difference in toxicity could be resolved, suggesting that higher doses may be required to detect a FLASH sparing of skin.

Beam commissioning and measurements validating the beam model in a new TPS that converts helical tomotherapy plans to step-and-shoot IMRT plans.

PURPOSE: A new type of treatment planning system called SHAREPLAN has been studied, which enables the transfer of treatment plans generated for helical tomotherapy delivery to plans that can be delivered on C-arm linacs. The purpose is to ensure continuous patient treatment during periods of unscheduled downtime for the TomoTherapy unit, particularly in clinics without a backup unit. The purpose of this work was to verify that the plans generated in this novel planning system are deliverable and accurate. The work consists primarily of beam commissioning, verification of the beam model, and measurements verifying that generated plans are deliverable with sufficient accuracy. METHODS: The beam commissioning process involves input of general geometric properties of the modeled linac, profiles and depth dose curves for a specific photon nominal energy (6 MV), and the automated modeling of other beam properties. Some manual tuning of the beam model is required. To evaluate its accuracy, the confidence limit concept [J. Venselaar et al., "Tolerances for the accuracy of photon beam dose calculations of treatment planning systems," Radiother. Oncol. 60, 191-201 (2001)] was used, which is a method supported by ESTRO. Measurements were conducted with a 2D diode array at the commissioned linac as a final check of the beam model and to evaluate whether the generated plans were deliverable and accurate. RESULTS: The comparison and evaluation of calculated data points and measured data according to the method applied confirmed the accuracy of the beam model. The profiles had a confidence limit of 1.1% and the depth dose curves had a confidence limit of 1.7%, both of which were well below the tolerance limit of 2%. Plan specific QC measurements and evaluation verified that different plans generated in the TPS were deliverable with sufficient accuracy at the commissioned linac, as none of the 160 beams for the 20 different plans evaluated had a fraction of approved data points below 90%, the local clinical approval criterion for delivery QA measurements. CONCLUSIONS: This study is a validation of the new TPS as it verifies that the generated plans are deliverable at a commissioned linac with adequate accuracy. A thorough investigation of the treatment plan quality will require a separate study. The TPS is proving to be a useful and time-saving complement, especially for clinics having a single unit for helical delivery among its conventional linacs.