Ultra-high Dose Rate Irradiation Research Articles

Radioluminescence-based fibre-optic dosimeters in radiotherapy: a review

Since their conception in the early 1990s, radioluminescence-based fibre-optic dosimeters (FODs) have attracted great interest for dosimetric applications in radiotherapy (RT). Over the years, many scintillating materials, both organic and inorganic, have been developed and tested by multiple research groups around the world. In parallel, reliable techniques for removing or subtracting the stem effect, one of the main drawbacks of FODs, have been proposed in the literature. To date, several prototype systems are widespread, some of which prove to be viable commercial solutions. This review aims to trace the efforts over the years that have enabled FODs to become reliable tools for dosimetry in the current RT procedures and promising options for future scenarios. After a first section devoted to a thorough discussion of the stem effect issue, the use of FODs in various applications of interest to RT, primarily small-field dosimetry and in vivo dosimetry, are addressed. Their use both in external beam RT, including hadrontherapy, and in brachytherapy are considered. A special focus is given to the peculiarities of MRI LINAC dosimetry and the contributions of FODs in this context. Finally, the state of the art concerning the development and characterization of FODs for monitoring ultra-high dose rate radiation beams, typical of FLASH therapy and microbeam RT are presented, highlighting the challenges still open.

Proton-FLASH: effects of ultra-high dose rate irradiation on an in-vivo mouse ear model

FLASH-radiotherapy may provide significant sparing of healthy tissue through ultra-high dose rates in protons, electrons, and x-rays while maintaining the tumor control. Key factors for the FLASH effect might be oxygen depletion, the immune system, and the irradiated blood volume, but none could be fully confirmed yet. Therefore, further investigations are necessary. We investigated the protective (tissue sparing) effect of FLASH in proton treatment using an in-vivo mouse ear model. The right ears of Balb/c mice were irradiated with 20 MeV protons at the ion microprobe SNAKE in Garching near Munich by using three dose rates (Conv = 0.06 Gy/s, Flash9 = 9.3 Gy/s and Flash930 = 930 Gy/s) at a total dose of 23 Gy or 33 Gy. The ear thickness, desquamation, and erythema combined in an inflammation score were measured for 180 days. The cytokines TGF-β1, TNF-α, IL1α, and IL1β were analyzed in the blood sampled in the first 4 weeks and at termination day. No differences in inflammation reactions were visible in the 23 Gy group for the different dose rates. In the 33 Gy group, the ear swelling and the inflammation score for Flash9 was reduced by (57 ± 12) % and (67 ± 17) % and for Flash930 by (40 ± 13) % and (50 ± 17) % compared to the Conv dose rate. No changes in the cytokines in the blood could be measured. However, an estimation of the irradiated blood volume demonstrates, that 100-times more blood is irradiated when using Conv compared to using Flash9 or Flash930. This indicates that blood might play a role in the underlying mechanisms in the protective effect of FLASH.

Technical note: Commissioning of a linear accelerator producing ultra-high dose rate electrons.

Ultra-high dose rate radiation (UHDR) is being explored by researchers in promise of advancing radiation therapy treatments. This work presents the commissioning of Varian's Flash Extension for research (FLEX) conversion of a Clinac to deliver UHDR electrons. A Varian Clinac iX with the FLEX conversion was commissioned for non-clinical research use with 16 MeV UHDR (16H) energy. This involved addition of new hardware, optimizing the electron gun voltages, radiofrequency (RF) power, and steering coils in order to maximize the accelerated electron beam current, sending the beam through custom scattering foils to produce the UHDR with 16H beam. Profiles and percent depth dose (PDD) measurements for 16H were obtained using radiochromic film in a custom vertical film holder and were compared to 16 MeV conventional electrons (16C). Dose rate and dose per pulse (DPP) were calculated from measured dose in film. Linearity and stability were assessed using an Advanced Markus ionization chamber. Energies for 16H and 16C had similar beam quality based on PDD measurements. Measurements at the head of the machine (61.3cm SSD) with jaws set to 10×10cm2 showed the FWHM of the profile as 7.2cm, with 3.4Gy as the maximum DPP and instantaneous dose rate of 8.1E5 Gy/s. Measurements at 100cm SSD with 10cm standard cone showed the full width at half max (FWHM) of the profile as 10.5cm, 1.08Gy as the maximum DPP and instantaneous dose rate of 2.E5 Gy/s. Machine output with number of pulses was linear (R=1) from 1 to 99 delivered pulses. Output stability was measured within±1% within the same session and within±2% for daily variations. The FLEX conversion of the Clinac is able to generate UHDR electron beams which are reproducible with beam properties similar to clinically used electrons at 16 MeV. Having a platform which can quickly transition between UHDR and conventional modes (<1min) can be advantageous for future research applications.

Modeling for predicting survival fraction of cells after ultra-high dose rate irradiation

Objective. FLASH radiotherapy (FLASH-RT) with ultra-high dose rate (UHDR) irradiation (i.e. > 40 Gy s−1) spares the function of normal tissues while preserving antitumor efficacy, known as the FLASH effect. The biological effects after conventional dose rate-radiotherapy (CONV-RT) with ≤0.1 Gy s−1 have been well modeled by considering microdosimetry and DNA repair processes, meanwhile modeling of radiosensitivities under UHDR irradiation is insufficient. Here, we developed an integrated microdosimetric-kinetic (IMK) model for UHDR-irradiation enabling the prediction of surviving fraction after UHDR irradiation. Approach. The IMK model for UHDR-irradiation considers the initial DNA damage yields by the modification of indirect effects under UHDR compared to CONV dose rate. The developed model is based on the linear-quadratic (LQ) nature with the dose and dose square coefficients, considering the reduction of DNA damage yields as a function of dose rate. Main results. The estimate by the developed model could successfully reproduce the in vitro experimental dose–response curve for various cell line types and dose rates. Significance. The developed model would be useful for predicting the biological effects under the UHDR irradiation.

Comparison of Tumor Control and Skin Damage in a Mouse Model after Ultra-High Dose Rate Irradiation and Conventional Irradiation.

Recent studies suggest ultra-high dose rate radiation treatment (UHDR-RT) reduces normal tissue damage compared to conventional radiation treatment (CONV-RT) at the same dose. In this study, we compared first, the kinetics and degree of skin damage in wild-type C57BL/6 mice, and second, tumor treatment efficacy in GL261 and B16F10 dermal tumor models, at the same UHDR-RT and CONV-RT doses. Flank skin of wild-type mice received UHDR-RT or CONV-RT at 25 Gy and 30 Gy. Normal skin damage was tracked by clinical observation to determine the time to moist desquamation, an endpoint which was verified by histopathology. Tumors were inoculated on the right flank of the mice, then received UHDR-RT or CONV-RT at 1 × 11 Gy, 1 × 15, 1 × 25, 3 × 6 and 3 × 8 Gy, and time to tumor tripling volume was determined. Tumors also received 1 × 11, 1 × 15, 3 × 6 and 3 × 8 Gy doses for assessment of CD8+/CD4+ tumor infiltrate and genetic expression 96 h postirradiation. All irradiations of the mouse tumor or flank skin were performed with megavoltage electron beams (10 MeV, 270 Gy/s for UHDR-RT and 9 MeV, 0.12 Gy/s for CONV-RT) delivered via a clinical linear accelerator. Tumor control was statistically equal for similar doses of UHDR-RT and CONV-RT in B16F10 and GL261 murine tumors. There were variable qualitative differences in genetic expression of immune and cell damage-associated pathways between UHDR and CONV irradiated B16F10 tumors. Compared to CONV-RT, UHDR-RT resulted in an increased latent period to skin desquamation after a single 25 Gy dose (7 days longer). Time to moist skin desquamation did not significantly differ between UHDR-RT and CONV-RT after a 30 Gy dose. The histomorphological characteristics of skin damage were similar for UHDR-RT and CONV-RT. These studies demonstrated similar tumor control responses for equivalent single and fractionated radiation doses, with variable difference in expression of tumor progression and immune related gene pathways. There was a modest UHDR-RT skin sparing effect after a 1 × 25 Gy dose but not after a 1 × 30 Gy dose.

Effects of UHDR and Conventional Irradiation on Behavioral and Cognitive Performance and the Percentage of Ly6G+ CD45+ Cells in the Hippocampus.

We assessed the effects of conventional and ultra-high dose rate (UHDR) electron irradiation on behavioral and cognitive performance one month following exposure and assessed whether these effects were associated with alterations in the number of immune cells in the hippocampus using flow cytometry. Two-month-old female and male C57BL/6J mice received whole-brain conventional or UHDR irradiation. UHDR mice were irradiated with 9 MeV electrons, delivered by the Linac-based/modified beam control. The mice were irradiated or sham-irradiated at Dartmouth, the following week shipped to OHSU, and behaviorally and cognitively tested between 27 and 41 days after exposure. Conventional- and UHDR-irradiated mice showed impaired novel object recognition. During fear learning, conventional- and UHDR-irradiated mice moved less during the inter-stimulus interval (ISI) and UHDR-irradiated mice also moved less during the baseline period (prior to the first tone). In irradiated mice, reduced activity levels were also seen in the home cage: conventional- and UHDR-irradiated mice moved less during the light period and UHDR-irradiated mice moved less during the dark period. Following behavioral and cognitive testing, infiltrating immune cells in the hippocampus were analyzed by flow cytometry. The percentage of Ly6G+ CD45+ cells in the hippocampus was lower in conventional- and UHDR-irradiated than sham-irradiated mice, suggesting that neutrophils might be particularly sensitive to radiation. The percentage of Ly6G+ CD45+ cells in the hippocampus was positively correlated with the time spent exploring the novel object in the object recognition test. Under the experimental conditions used, cognitive injury was comparable in conventional and UHDR mice. However, the percentage of CD45+ CD11b+ Ly6+ and CD45+ CD11b+ Ly6G- cells in the hippocampus cells in the hippocampus was altered in conventional- but not UHDR-irradiated mice and the reduced percentage of Ly6G+ CD45+ cells in the hippocampus might mediate some of the detrimental radiation-induced cognitive effects.

Treatment planning of intracranial lesions with VHEE: comparing conventional and FLASH irradiation potential with state-of-the-art photon and proton radiotherapy

The treatment of deep-seated tumours with electrons of very high energies (VHEE, 70–150 MeV) has already been explored in the past, suggesting that a dosimetric coverage comparable with state-of-the-art proton (PT) or photon radiotherapy (RT) could be achieved with a large (&gt; 10) number of fields and high electron energy. The technical and economical challenges posed by the deployment of such beams in treatment centres, together with the expected small therapeutic gain, prevented the development of such technique. This scenario could radically change in the light of recent developments that occurred in the compact, high-gradient, electron acceleration technology and, additionally, of the experimental evidence of the sparing of organs at risk achieved in ultra-high dose rate irradiation, also referred to as FLASH. Electrons with the energy required to treat intracranial lesions could be provided, at dose rates compatible with what is needed to trigger the FLASH effect, by accelerators that are a few metres long, and the organ sparing could be exploited to significantly simplify the irradiation geometry, decreasing the number of fields needed to treat a patient. In this paper, the case of two patients affected by a chordoma and a meningioma, respectively, treated with protons in Trento (IT) is presented. The proton plans have been compared with VHEE plans and X-ray intensity-modulated radiotherapy (IMRT) plans. The VHEE plans were first evaluated in terms of physical dose distribution and then assuming that the FLASH regimen can be achieved. VHEE beams demonstrated their potential in obtaining plans that have comparable tumour coverage and organs at risk sparing when benchmarked against current state-of-the-art IMRT and PT. These results were obtained with a number of explored fields that was in the range between 3 and 7, consistent with what is routinely performed in IMRT and PT conventional irradiations. The FLASH regimen, in all cases, showed its potential in reducing damage to the organs placed nearby the target volume, allowing, particularly in the chordoma case where the irradiation geometry is more challenging, a better tumour coverage with respect to the conventional treatments.

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

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. 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. 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. 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.

OC-0623 Ultra-High dose rate irradiation spares a large quantity of circulating lymphocytes in brain tumours

OC-0623 Ultra-High dose rate irradiation spares a large quantity of circulating lymphocytes in brain tumours

Absence of Tissue-Sparing Effects in Partial Proton FLASH Irradiation in Murine Intestine.

Ultra-high dose rate irradiation has been reported to protect normal tissues more than conventional dose rate irradiation. This tissue sparing has been termed the FLASH effect. We investigated the FLASH effect of proton irradiation on the intestine as well as the hypothesis that lymphocyte depletion is a cause of the FLASH effect. A 16 × 12 mm2 elliptical field with a dose rate of ~120 Gy/s was provided by a 228 MeV proton pencil beam. Partial abdominal irradiation was delivered to C57BL/6j and immunodeficient Rag1-/-/C57 mice. Proliferating crypt cells were counted at 2 days post exposure, and the thickness of the muscularis externa was measured at 280 days following irradiation. FLASH irradiation did not reduce the morbidity or mortality of conventional irradiation in either strain of mice; in fact, a tendency for worse survival in FLASH-irradiated mice was observed. There were no significant differences in lymphocyte numbers between FLASH and conventional-dose-rate mice. A similar number of proliferating crypt cells and a similar thickness of the muscularis externa following FLASH and conventional dose rate irradiation were observed. Partial abdominal FLASH proton irradiation at 120 Gy/s did not spare normal intestinal tissue, and no difference in lymphocyte depletion was observed. This study suggests that the effect of FLASH irradiation may depend on multiple factors, and in some cases dose rates of over 100 Gy/s do not induce a FLASH effect and can even result in worse outcomes.

Human enteroids as a tool to study conventional and ultra-high dose rate radiation.

Radiation therapy, one of the most effective therapies to treat cancer, is highly toxic to healthy tissue. The delivery of radiation at ultra-high dose rates, FLASH radiation therapy (FLASH), has been shown to maintain therapeutic anti-tumor efficacy while sparing normal tissues compared to conventional dose rate irradiation (CONV). Though promising, these studies have been limited mainly to murine models. Here, we leveraged enteroids, three-dimensional cell clusters that mimic the intestine, to study human-specific tissue response to radiation. We observed enteroids have a greater colony growth potential following FLASH compared with CONV. In addition, the enteroids that reformed following FLASH more frequently exhibited proper intestinal polarity. While we did not observe differences in enteroid damage across groups, we did see distinct transcriptomic changes. Specifically, the FLASH enteroids upregulated the expression of genes associated with the WNT-family, cell-cell adhesion, and hypoxia response. These studies validate human enteroids as a model to investigate FLASH and provide further evidence supporting clinical study of this therapy. Insight Box Promising work has been done to demonstrate the potential of ultra-high dose rate radiation (FLASH) to ablate cancerous tissue, while preserving healthy tissue. While encouraging, these findings have been primarily observed using pre-clinical murine and traditional two-dimensional cell culture. This study validates the use of human enteroids as a tool to investigate human-specific tissue response to FLASH. Specifically, the work described demonstrates the ability of enteroids to recapitulate previous in vivo findings, while also providing a lens through which to probe cellular and molecular-level responses to FLASH. The human enteroids described herein offer a powerful model that can be used to probe the underlying mechanisms of FLASH in future studies.

Abstract P1-10-02: FLASH-RT, ultra-high dose rate rate radiotherapy, is as effective as conventional dose rate radiotherapy in eradicating tumor in a preclinical model of breast cancer

Abstract Problem Statement: Radiation therapy (RT) for breast cancer (BC) can induce skin and soft tissue fibrosis, raises concerns over cardiac and pulmonary injury, is associated with higher rates of lymphedema and shoulder dysfunction with regional nodal irradiation, and significantly increases complication rates in women undergoing implant-based reconstruction due to radiation toxicity. Although these toxicities are not generally associated with higher mortality, in general, they can represent significant setbacks with respect to quality of life. These toxicities dissuade some patients from breast conservation leading to unnecessary mastectomy and can lead patients to omit reconstruction after mastectomy or choose more extensive autologous breast reconstruction. Women with implant-based reconstruction and radiotherapy have known higher rates of reconstruction failure. Purpose: Ultra-high dose rate radiation (FLASH) has been shown to induce less normal tissue toxicity, therefore if tumor control of FLASH-RT would be comparable to conventional radiotherapy (CONV) then it has the potential to lower morbidity associated with radiotherapy for breast cancer and allow overall improved outcomes. At first, we aimed to determine the effectiveness of FLASH-RT compared to CONV in eradicating small breast tumors in an orthotopic BC model using single-fraction 20 or 30Gy RT to compare effectiveness of FLASH-RT vs CONV. Methods: Radiation sensitive, syngeneic mammary tumor cell line Py117, that efficiently forms non-metastatic orthotopic tumors in C57BL/6 mice, were injected (106 cells) into the left 4th mammary fat pad. 30mm3 tumors or a range of greater volumes (200-800mm3) were irradiated with single-fraction 20 or 30Gy with a 2x2cm radiation field (~17MeV beams), exposing only 5mm of the surrounding tissue. FLASH RT was delivered with 2Gy per pulse at dose rate ~200Gy/s compared to CONV dose rate of 0.13Gy/s Results: Single-fraction 20Gy suppressed 30mm3 tumor growth until ~day 15 post-RT then regrew for both FLASH and CONV, while 30mm3 tumors were eradicated with both FLASH and CONV at 30Gy. Larger tumors irradiated with 30Gy regressed until ~day 12 post-RT then regrew for both FLASH and CONV. There was no significant difference in growth suppression or tumor eradication between FLASH and CONV in any cohort. Conclusion: In this murine model of breast cancer, FLASH is as effective as CONV in controlling tumor growth. Future studies will extend the evaluation of the tumor control using clinically relevant fractionated dose schedules to be followed by comparisons of tumor control in xenograft models. Additional studies will assess normal tissue toxicity of FLASH vs CONV in murine models of implant-based breast reconstruction. We have established collaborations to understand differences in molecular pathways activated by FLASH vs CONV in tumor and normal tissue to explain the observed experimental differences in normal tissue, tumor, and cancer stem cells. Citation Format: Frederick Dirbas, Stavros Melemenidis, Bill Loo, Kathleen Horst, Edward E. Graves, Suparna Dutt, Vignesh Viswanathan, Brianna Lau, Amy Yu. FLASH-RT, ultra-high dose rate rate radiotherapy, is as effective as conventional dose rate radiotherapy in eradicating tumor in a preclinical model of breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P1-10-02.

Flash-Effect in Radiotherapy of Tumors and the Problems of its Radiobiological Substantiation

The published data regarding radiation therapy of tumors treatment with ultrahigh dose rate radiation has been analyzed. In 2014, the authors, among whom the most mentioned are Vincent Favaudon and MarieCatherine Vozenin, named it the flash effect. Note, however, that a member of our laboratory, S.V. Kozin in the collaboration with ITEP researchers, back in 1984, having studied the reaction of a graft tumors and the mice skin after the proton irradiation with the ultrahigh and ordinary dose rates, concluded that: ‘... an increase in the radiation dose rate to ultrahigh, without significantly changing radiation reactions of tumors, leads to the protection of the mice skin, which is associated with the manifestation of the effect of radiation-chemical absorption of oxygen. Thus, under certain conditions of radiotherapy, one can expect to receive a therapeutic gain due to use the radiation with a high dose rate’. Consideration of the expediency and possibility of using a single irradiation with a dose rate of tens of Gy/s in radiation therapy of tumors is now given a special attention. It can be considered as established that less damage to the skin, lungs, intestines, and some other tissues occurs in this case in comparison with the usual dose rates used for the radiation therapy. Moreover, a number of studies indicate an increase in radiation damage to the tumors, or at least a much smaller decrease in the degree of their damage. The radiobiological mechanisms remain unclear. The effect itself is of undoubted clinical and experimental interest and justifies the conduct of experimental and clinical studies, as well as the need to take it into account when developing new equipment for radiation therapy and dosimetry accompanying the treatment process.

Recent progress in pencil beam scanning FLASH proton therapy: a narrative review

Background and Objective: Recent experimental studies using ultra-high dose rate radiation therapy (FLASH-RT) have shown improved normal tissue sparing and comparable tumor control compared to conventional dose rate radiotherapy (RT). Pencil beam scanning (PBS) proton therapy with superior dosimetry characteristics has begun to draw attention to the delivery of conformal FLASH-RT for preclinical studies. This review aims to provide recent updates on the development of PBS FLASH-RT.

TMIC-68. EVALUATING FLASH AND CONVENTIONAL DOSE-RATE RADIATION AND IMMUNE RESPONSE WITH SINGLE-CELL SEQUENCING IN DIFFUSE MIDLINE GLIOMA (DMG)

Abstract Diffuse Midline Glioma – H3K27M mutant (DMG), is a fatal and inoperable pediatric brain tumor with limited treatment options as radiation provides only temporary reprieve, leaving the median survival between 9-15 months. Conventional dose-rate radiation (2Gray/minute, CONV) has been shown in other cancers to recruit an immune component, however, this has not been studied in DMG. Ultra-high dose-rate radiation given at 90 Gray/second (FLASH), is a novel technique associated with decreased toxicity and effective tumor control. Using a syngeneic model of brainstem DMG, we performed single-cell RNA sequencing on CD45+ immune cells isolated from tumors irradiated with 15Gray using FLASH or CONV, and compared to unirradiated tumor and normal brainstem. Isolation of 33,308 immune cells revealed 17 unique clusters, most abundant of which was microglia (73.8%), present in four distinct subtypes representing a spectrum from homeostatic to activated. In the most activated microglia, both FLASH and CONV showed an enrichment in type 1 interferon (IFN1) pathway scores compared to untreated tumors (p&amp;lt; 0.001 and p&amp;lt; 0.001, respectively). The most differential response was found in macrophages (MAC) and dendritic cells (DC) with a robust enrichment of IFN1 pathway scores for CONV compared to FLASH (p&amp;lt; 0.001, MAC and p&amp;lt; 0.001 DC). FLASH showed an increase in anti-inflammatory MAC markers such as Mrc1, Cd163, and Maf and an enrichment of myeloid-derived suppressor cell (MDSC) signature in monocytes, not seen in CONV (p&amp;lt; 0.001). Finally, we correlated our data with publicly available single-cell data taken from the cerebrospinal fluid of DMG patients treated with anti-GD2 CAR T Cell therapy and found similar inflammatory markers characteristic of our unirradiated murine tumors. In summary, our work is the first to study immune alterations comparing different dose-rates of radiation with single-cell resolution in DMG, highlighting the potential for combining radiation and immunotherapy in these tumors.

Ultra-High Dose Rate FLASH Effect on Head and Neck Cancer Cells and Normal Salivary Gland in Mice

<h3>Purpose/Objective(s)</h3> Ultra-high dose rate irradiation (FLASH) is a novel therapeutic modality that has been reported to spare tissues compared to conventional dose rate. We aimed to investigate the effect of FLASH on head and neck (H&N) cancer cells and salivary gland dysfunction caused by radiotherapy. <h3>Materials/Methods</h3> H&N cancer cells (HSG and FaDu) were irradiated with 10 MeV electron beam at a dose rate of 132.2Gy/s (FLASH) or 0.1Gy/s (CONV), by a modified clinic linear accelerator. The irradiated dose was confirmed with EBT-XD film, and 3, 7, and 11 Gy were irradiated. We evaluated the cell cycle and the surviving fraction of cells with clonogenic assays. We administered single dose head and neck irradiation to mice with a 6 MeV electron beam at dose rate of 105.6-126 Gy/s (FLASH) or 0.03Gy/s (CONV), using a modified linear accelerator. The irradiated dose was confirmed with EBT3 film, and 9-76 Gy and 10-30 Gy were irradiated for FLASH and CONV, respectively. Parotid and submandibular glands were obtained from mice at 3 and 30 days after irradiation. Immunohistological examination and apoptotic assays were conducted to assess the radiation response. <h3>Results</h3> The proportion of sub G0/G1 was higher in the FLASH group, and the surviving fraction was significantly decreased in FLASH through every dose range. At three days post-irradiation, both 10 Gy and 30 Gy irradiated groups showed a greater number of salivary cells and increased expression of AQP5 in FLASH compared to conventional irradiated mice. As a late toxicity (30 days of irradiation), fibrosis did not show any significant difference, although FLASH group received much higher radiation dose. There was no difference in apoptosis between groups on post 3 days, but FLASH showed significantly less degree of apoptosis on 30 days after irradiation. <h3>Conclusion</h3> This study demonstrates that FLASH irradiation strengthens the therapeutic effect for H&N cancers and attenuates radiation-induced salivary gland damage. Further research is needed on the biological mechanisms involved.

Modification of Commercial Radiotherapy System for Ultra-High Dose Rate Electron Beam Irradiation

<h3>Purpose/Objective(s)</h3> Because electron generation in X-ray mode needs to be about 1000 times more than electron beam mode, modifying commercial radiotherapy system make it possible to irradiate with ultra-high dose rate. This study aimed to show the feasibility of ultra-high dose rate irradiation using a commercial linear accelerator. <h3>Materials/Methods</h3> Removing targets and flattening filters, adjusting source-surface-distance (SSD), and controlling RF driver and electron gun were performed in Simens Oncor® and Varian Clinac iX® radiotherapy system. The irradiation time was adjusted in microseconds through the Arduino-based gating system, and dose rate was confirmed with EBT3 and EBT-XD films. <h3>Results</h3> In 6 MV X-ray mode of Siemens® system, 100.9±3.78 Gy/s electron beam was irradiated at SSD of 60cm. After modifying 10MV X-ray mode in Varian® radiotherapy system, dose rate was 43.1Gy/s at SSD of 100cm. By manually controlling the parameters of the RF driver and electron gun, 318 Gy/s rate of electron beam was irradiated within 15 × 15 cm² of irradiation field at SSD of 100cm. Acquired ultra-high dose rate electron beam exhibits the following characteristics at SSD of 100cm with full opened jaw irradiation: R₅₀ 4.1cm, W₈₀ 9.8cm (vertical) and 9.3cm (horizontal). <h3>Conclusion</h3> We confirmed that FLASH irradiation was feasible by modifying a commercial radiotherapy system, which made further research possible.

FLASH vs. Conventional Dose Rate Proton Radiation in Mouse Abdomen

<h3>Purpose/Objective(s)</h3> The mechanisms underlying normal tissue toxicity sparing and survival benefits of ultra-high dose rate radiation (FLASH) are still unclear. We used a low-energy proton system (50 MeV) optimized for small animal radiobiological research to study the biological effects after FLASH and conventional dose rate (CONV) proton radiation to mouse abdomen. <h3>Materials/Methods</h3> We radiated the abdomen of 6–7-week-old female C57BL/6 mice using the plateau region of a continuous (unpulsed) cyclotron-generated 50 MeV preclinical proton beam, transmitting through the abdomen, with 1.5cm (for survival study) or 3cm (for EdU assay) width of beam via customized vertical and horizontal collimators. Mice survival and regenerated crypts in colon tissues using EdU assay were studied. Mice were stratified into 3 groups: 1) control/sham radiation (n=8 for survival, n=4 for EdU); 2) FLASH (20Gy at 48-93Gy/sec, n=22 for survival and n=13 for EdU); 3) CONV (20Gy at ∼1Gy/sec, n=19 for survival, n=11 for EdU). For survival study, mice were observed for survival post radiation. For EdU assay, the mice were injected with EdU 3-4 hours prior to euthanasia, and colon tissues were collected at 24 hours and 96 hours post-radiation for sectioning and then EdU staining. <h3>Results</h3> FLASH and CONV groups showed different survival: FLASH (13 days post radiation, 36.4% survival); CONV (15.6% survival, P = 0.04). FLASH group had more EdU staining than the CONV group, though both FLASH and CONV groups showed reduced EdU staining compared to the control group. <h3>Conclusion</h3> Preliminary results of mouse abdomen FLASH and CONV proton radiation from a 50 MeV beam suggest FLASH proton radiation leads to better survival and less normal tissue damage than CONV radiation.

FLASH with carbon ions: Tumor control, normal tissue sparing, and distal metastasis in a mouse osteosarcoma model.

The FLASH effect is a potential breakthrough in radiotherapy because ultra-high dose-rate irradiation can substantially widen the therapeutic window. While the normal tissue sparing at high doses and short irradiation times has been demonstrated with electrons, photons, and protons, so far evidence with heavy ions is limited to in vitro cell experiments. Here we present the first in vivo results with high-energy 12C-ions delivered at an ultra-high dose rate. LM8 osteosarcoma cells were subcutaneously injected in the posterior limb of female C3H/He mice 7days before radiation exposure. Both hind limbs of the animals were irradiated with 240MeV/n 12C-ions at ultra-high (18Gy in 150ms) or conventional dose rate (∼18Gy/min). Tumor size was measured until 28days post-exposure, when animals were sacrificed and lungs, limb muscles, and tumors were collected for further histological analysis. Irradiation with carbon ions was able to control the tumour both at conventional and ultra-high dose rate. FLASH decreases normal tissue toxicity as demonstrated by the reduced structural changes in muscle compared to conventional dose-rate irradiation. Carbon ion irradiation in FLASH conditions significantly reduced lung metastasis compared to conventional dose-rate irradiation and sham-irradiated animals. We demonstrated the FLASH effect in vivo with high-energy carbon ions. In addition to normal tissue sparing, we observed tumor control and a substantial reduction of lung metastasis in an osteosarcoma mouse model.

Radiobiological Aspects of FLASH Radiotherapy.

Radiotherapy (RT) is one of the primary treatment modalities for cancer patients. The clinical use of RT requires a balance to be struck between tumor effect and the risk of toxicity. Sparing normal tissue is the cornerstone of reducing toxicity. Advances in physical targeting and dose-shaping technology have helped to achieve this. FLASH RT is a promising, novel treatment technique that seeks to exploit a potential normal tissue-sparing effect of ultra-high dose rate irradiation. A significant body of in vitro and in vivo data has highlighted a decrease in acute and late radiation toxicities, while preserving the radiation effect in tumor cells. The underlying biological mechanisms of FLASH RT, however, remain unclear. Three main mechanisms have been hypothesized to account for this differential FLASH RT effect between the tumor and healthy tissue: the oxygen depletion, the DNA damage, and the immune-mediated hypothesis. These hypotheses and molecular mechanisms have been evaluated both in vitro and in vivo. Furthermore, the effect of ultra-high dose rate radiation with extremely short delivery times on the dynamic tumor microenvironment involving circulating blood cells and immune cells in humans is essentially unknown. Therefore, while there is great interest in FLASH RT as a means of targeting tumors with the promise of an increased therapeutic ratio, evidence of a generalized FLASH effect in humans and data to show that FLASH in humans is safe and at least effective against tumors as standard photon RT is currently lacking. FLASH RT needs further preclinical investigation and well-designed in-human studies before it can be introduced into clinical practice.