Efficacy of Radiotherapy FLASH-RT: From Treating Lung Cancer to Preventing Pulmonary Fibrosis

Abstract

FLASH radiotherapy (FLASH-RT) is a radiotherapy technique that achieves ultra-high dose rates in a fraction of a second. Based on data from experimental animal models, FLASH-RT appears to protect a number of normal tissues from radiation-induced damage, including the brain, gastrointestinal tract, and lung, while conventional radiotherapy (CONV-RT) causes radiation-induced toxicity in these tissues. In this review, we provide a brief summary of the history of radiation therapy and focus on some of the most recent FLASH-RT papers and findings. It is particularly noteworthy that pulmonary fibrosis represents a common complication of radiotherapy. New evidence indicates that FLASH-RT, unlike traditional radiotherapy methods, might help protect lung cancer patients from developing pulmonary fibrosis caused by radiation. FLASH-RT will advance more quickly than anticipated, although there are still a number of unresolved concerns. FLASH-RT will be a safer and more effective option for lung cancer treatment.

Acute Lung Injury and Pulmonary Fibrosis Caused by Lung Radiotherapy

Radiotherapy is a major strategy for eliminating cancer cells and curtailing tumor development. ¹ According to some surveys, 50-70% of cancer patients receive radiation treatment as part of their treatment. ² Radiotherapy becomes the most crucial kind of treatment for patients with lung cancer who have advanced to the middle and late stages. ³ Currently, conventional radiotherapy (CONV-RT) and accelerated hyperfractionated radiotherapy are the 2 main forms of radiotherapy used for clinically diagnosed patients with middle and late-stage lung cancer. ⁴

Radiotherapy can cause inflammation and tissue damage, particularly in the lung, which is one of the most radiation-sensitive organs. Acute lung injury usually occurs within 1-3 months after radiotherapy. This injury may be associated with radiotherapy-induced lung inflammation and fibrosis, especially in patients receiving total body irradiation (TBI). ⁵ In one study, 24.8% of 335 patients undergoing TBI developed pulmonary toxicity, mainly in the form of pneumonia and pulmonary obstruction. ⁶ In addition, changes in the lungs after radiotherapy can be monitored by regular CT scans, which help to assess the evolution of radiotherapy-induced lung damage. ⁷ In the early stages after radiotherapy, patients may experience acute inflammation, which usually manifests itself as reversible lung changes within 3-6 months. ⁷ However, over time, chronic scarring may occur, leading to irreversible lung damage. ⁷ Patients may exhibit cyanosis and respiratory insufficiency in cases of severe acute injury, which can result in pulmonary hypertension and acute corpulmonale. Hemoptysis is an uncommon sign. Radiation fibrosis is a type of long-term lung injury that typically develops 6-24 months following radiation exposure. Patients may show up with different degrees of dyspnea or without any symptoms at all. Patients who have had a significant amount of lung radiation may develop chronic pulmonary insufficiency. This could develop into pulmonary hypertension and cor pulmonale. As of right now, fibrosis has no effective therapeutic intervention. ⁸ In a radiological analysis of lung injury following conventional stereotactic body radiation therapy, radiological anomalies were found in 54% of patients 6 months after irradiation and in 99% of patients 3 months later. Over a 2-year period following therapy, 47% of lesions exhibited persistent changes in either morphology or severity. ⁹ The use of CONV-RT for lung cancer is severely restricted because of the prevalence of normal tissue toxicity symptoms.

In addition, most chemotherapeutic agents used in conjunction with radiotherapy increase the risk of damage to normal tissue. Thus, reducing radiation-induced normal tissue damage is of critical importance in improving both tumor control and patient quality of life. The development of new therapies or strategies will, however, require an improved understanding of the basic molecular mechanisms that underlie the development of lung injury.

What is FLASH Radiotherapy and Why is It Different from CONV-RT?

FLASH radiation therapy (FLASH-RT) is a novel method that uses ultra-high dose rate irradiation (>40 Gy/s) to treat malignancies with comparable therapeutic efficacy while sparing normal tissue.^10-12 This important feature has led to current widespread interest in several scientific communities. Numerous remarkable outcomes have been demonstrated in pre-clinical investigations employing electrons and photons.^13-15 When exposed to FLASH-RT, normal tissues such as the brain, ¹⁵ gastrointestinal tract, ¹⁶ and lung ¹⁷ can be notably protected from radiation-induced damage, which is known as the FLASH effect. ¹⁸ First discovered in the 1970s by Hornsey and Bewley, they discovered that mouse skin exposed to radiation at a greater dose rate (5000 Gy/min) showed less damage than skin exposed to radiation at a lower dose rate (700 Gy/min). ¹⁹ Forty years later, Favaudon et al ¹⁷ discovered that, in comparison to conventional irradiation, ultra-high dose rate irradiation (>40 Gy/s) significantly reduced the side effects of normal tissue in mice thorax irradiation while retaining comparable tumor control. Other favorable results have been reported in the head,^20-22 skin, ²³ and abdominal tissues. ¹⁶

The clinical translation of the FLASH effect is still at its early experimental phase, despite positive preclinical findings.^24,25 This is mainly because there are a few important physical, dosimetric, and radiobiological elements that remain unclear. Before being translated into the clinic, it is necessary to clarify how the “FLASH effect” is dependent on variations in various temporal beam parameters. This can be done by designing new devices that can deliver beam fluences several orders of magnitude higher than those used for CONV-RT, as well as by making new dosimeters and dosimetric protocols available that can measure beams at the very high dose per pulse needed to trigger the effect.^26,27 Furthermore, a thorough comprehension of the fundamental radiobiological mechanisms responsible for the FLASH effect still has to be clarified.

The “oxygen depletion” concept proposed by Vozenin and colleagues, ²⁸ which holds that FLASH-RT can rapidly deplete intracellular oxygen, resulting in a temporary radiation-induced hypoxic state, has emerged as a compelling explanation for FLASH-RT. However, newer studies, such as those by Jansen et al, ²⁹ have disputed this theory, demonstrating that FLASH effects continue even when oxygen is completely depleted, implying alternative explanations. At the physicochemical level, FLASH-RT delivers ultra-high dose rates in microseconds, resulting in water ionization and excitation, as well as the formation of reactive oxygen species (ROS) such as hydroxyl radicals. ³⁰ These ROS interact with biological macromolecules, causing oxidative stress and cell damage. FLASH-RT causes DNA damage in cancer cells in 2 ways: directly and indirectly through ROS, which is its main way of working against cancer. ³¹ However, normal cells may mitigate this damage through enhanced DNA repair mechanisms. ³² Immunologically, FLASH-RT may modulate immune and inflammatory processes to enhance anti-tumor effects. For instance, FLASH irradiation can reduce pro-inflammatory cytokines like TGF-β, thereby mitigating radiation-induced chronic inflammation. ³³ Preclinical studies in animal models have shown that FLASH-RT protects healthy tissues while still effectively killing tumors, possibly by affecting the immune system. ³⁴

Recently, Khatib et al ³⁵ employed ultra-fast tracking technology to monitor oxygen dynamics during proton FLASH-RT, revealing oxygen consumption and replenishment processes during FLASH irradiation. This study provides novel insights into the oxygen kinetics underlying FLASH effects, complementing Vozenin’s hypothesis and suggesting that FLASH-RT’s unique biological effects may stem from complex interactions between oxygen depletion, ROS generation, and immune modulation. However, the whole mechanism of FLASH-RT has yet to be defined, particularly in terms of its unique activities across cell types and tissues. Future study should focus on FLASH-RT’s long-term and holistic effects, as well as strategies for optimizing therapeutic applications.

Protection from Pulmonary Fibrosis by FLASH-RT

The main features of pulmonary fibrosis are the breakdown of lung tissue and the creation of scar tissue, which makes the lungs stiff and reduces the exchange of gases between blood and air. These changes make it more challenging to enlarge the lungs. A number of symptoms and possibly even death could arise from the combination of these effects. The damage is progressive and irreversible. ⁸ Radiation therapy has been shown to influence the establishment of pulmonary fibrosis by a variety of mechanisms, including oxidative stress, elevating cytokines, causing an inflammatory response, and changing fibrotic function.^36,37The possibility that the lung irradiated with FLASH would experience the FLASH effect has been extensively studied, as pulmonary fibrosis is a harmful side effect of radiation therapy.

Favaudon et al ¹⁷ employed the C57BL/6J mouse model of lung fibrosis to evaluate the incidence of fibrosis by histological and immunohistochemical methods following bilateral thorax exposure to continuous, conventional dose rate (≤0.03 Gy/s) vs pulsed, ultrahigh dose rate (≥40 Gy/s) irradiation administered as a single dose. They showed that, at similar doses, FLASH-RT stops lung fibrosis and greatly lowers the early occurrence of cell death in response to radiation. When FLASH-RT was first presented by Favaudon et al, the radiation community took an immediate interest in it.^20,23,38,39 They used human lung cells grown in vitro and C57BL/6J wild type and Terc^−/− mice harboring critically short telomeres and lacking telomerase activity exposed to bilateral thorax irradiation to examine lung responses to FLASH-RT. ³³ Research conducted in vitro has shown a decreased degree of DNA damage and mortality at the benefit of FLASH-RT. Using single-cell RNA sequencing and tracking of dividing cells, researchers found that FLASH-RT helped reduce the activation of genes that cause inflammation and slowed down the growth of progenitor cells after lung damage in mice. Lungs treated with FLASH-RT had fewer old and damaged cells compared to those treated with CONV-RT, suggesting a better chance for the lungs to heal. In line with this theory, Terc−/− mice no longer benefited from FLASH-RT. In line with this theory, Terc^−/− mice no longer benefited from FLASH-RT. Their research provides a biological hint for the advantageous FLASH impact resulting from normal tissue sparing. Gao et al ⁴⁰ found that in healthy C57BL/6 mice that got full-chest radiation, Masson’s special staining of lung sections showed that the lung structure of the FLASH group and the control group were similar, while the CONV group had more lung structure breakdown in the lung sections.

Recent studies have shown that FLASH irradiation significantly reduces the infiltration and activation of pro-inflammatory Ccrl2⁺ neutrophils and Mefv⁺ monocytes, thereby inhibiting the excessive inflammatory response they mediate. More importantly, FLASH irradiation strongly activated CD4⁺ CD40 L⁺ Th cells, which play a key role in immune regulation and inflammatory homeostasis. Additionally, FLASH irradiation enhanced the TGF-β signaling pathway and epithelial-mesenchymal transition in alveolar type I epithelial cells, thereby alleviating radiation-induced lung damage by promoting inflammation resolution and accelerating tissue repair processes. ⁴¹

In the work of Dai, ⁴² A549 cells were used to create xenografted tumor models in Balb/c-nu mice. CONV-RT had a dosage rate of 0.033 Gy/s, whereas FLASH-RT had a dose rate of 200 Gy/s with the same total dose (20 Gy). There were 2 different FLASH irradiation systems used: one pulse and ten pulses. The pathological study of the lung tissues and tumor before and after radiation, as well as changes in tumor volume, were used to evaluate the FLASH effect. The experimental data that have been reported indicate that, in the xenografted human lung tumor model, the high-energy X-rays provided by FLASH, at an average dose rate of 200 Gy/s, was able to protect the normal lung tissue from radiation-induced toxicities without affecting the efficacy of tumor management.

Wright et al ⁴³ studied how FLASH microbeam radiation treatment, which uses a special way to deliver radiation at a very high dose rate, affects normal lung tissue in rats .This paper highlights the superior performance of both ultra-high dose rate microbeam radiation (FLASH-MRT) field patterns over FLASH-RT patterns with an emphasis on fibrotic damage. Broad beam radiation caused detectable fibrotic damage over 25% of the irradiated region at a dose of 50 Gy, but 500 μm microbeams and 50 μm microbeams produced little and no fibrotic damage, respectively. Broad beam radiation caused high-grade fibrotic damage when exposed to 30 Gy of radiation. This outcome agrees with the findings of Favaudon et al ¹⁷ However, a peak entrance dose of 300 Gy with 50 μm microbeams only resulted in low-grade injury. The radioresistance of normal lung tissue is thus improved by at least an order of magnitude when comparing FLASH-MRT to FLASH-RT.

Previous research showed that a favorable predictor of radiation therapy response is T-cell infiltration into the tumor core.^44,45 Thus, Shukla et al ⁴⁶ used immunostaining for CD3, a marker of T-lymphocytes, to investigate the location of T-cells within the tumor. Radiation therapy was given to mice with lung cancers using 2 different speeds: FLASH-RT, which is very fast at over 60 Gy/s, and CONV-RT, which is much slower at 0.05 Gy/s. In contrast to traditional dose-rate-delivered protons, FLASH proton radiotherapy was found to decrease tumor burden, modify macrophage polarization towards an M1-like phenotype, enhance CD8⁺ T-cell infiltration into the tumor core, and modify the tumor microenvironment to become less immunotolerant. In a recent study, ⁴⁷ immunocompetent mice, as well as animals with moderate to severe impaired immune systems, received several mouse tumor models either subcutaneously or orthotopically. Mice were treated with radiation either in one large dose (20 Gy) or in smaller doses over several sessions (3 × 8 Gy; 2 × 6 Gy), with or without anti-CTLA-4, using FLASH-RT (≥2000 Gy/s) and CONV-RT (0.1 Gy/s). Immune profile analysis was done, and tumor progression was tracked throughout time. The findings unequivocally show that FLASH regulates the anti-tumor immune response, it may be employed as an immunomodulatory drug, and the tumor responses in a variety of immunocompetent and immunodeficient mice models are essentially dose-rate independent.

Very high-energy electrons (VHEE) offer a better balance in radiation treatment compared to regular photons and protons, making up for the shallow treatment depth of low-energy electrons. By combining FLASH and VHEE, we conducted an analysis to quantitatively compare various energies in terms of plan quality, dose rate distribution (in PTV and OAR), and total treatment duration (beam-on time). A comparative analysis was conducted between various energies in terms of treatment plan quality, irradiation time, and dose-averaged dose rate (DADR). According to Zhang et al, ⁴⁸ a high-quality treatment strategy may be able to maximize the benefits of the FLASH effect, leading to a notable increase in the tumor therapeutic gain ratio. The minimum beam intensity required for lung cases must be approximately 9.375 × 10¹¹ electrons/s in order to permit more than 90% of the PTV to attain the DADR of more than 40 Gy/s. The total irradiation time for the lung cases at this beam intensity (fraction dose: 10 Gy) is 1034.25 ms (100 MeV), 981.55 ms (120 MeV), and 928.15 ms (140 MeV), including 298.75 ms for scanning. The research successfully created a standard reference for how well VHEE radiation performs at FLASH dose rates by carefully studying the already known FLASH parameters.

A study used a 3D-printed mouse model that accurately represents its anatomy to check the differences in radiation doses between FLASH-RT and CONV-RT electron treatments at various hospitals. ⁴⁹ A dual-nozzle 3D printer was utilized to print the mouse phantom. Hounsfield units and densities were compared with the reference CT image of the living mouse after the lungs were printed individually using lightweight polylactic acid (∼0.64 g/cm³). Radiochromic film was used to check the average difference in radiation doses between FLASH-RT and CONV-RT, as well as how much they differed from the suggested dose. This audit demonstrated that FLASH-RT and CONV-RT electron irradiation can be supplied uniformly between collaborating institutions applying a 3D made mouse phantom.

Lipid peroxidation products have been linked to aging and human diseases. The expression of 37 oxylipins was measured in the lung of a mouse and in normal or cancerous cells exposed to CONV-RT or FLASH-RT. ⁵⁰ Fourteen to sixteen of the 37 isomers that were analyzed were present in sufficient amounts for quantitative analysis. The outcomes included the expression of oxylipins in relation to radiation dose, normoxia vs hypoxia, temperature, and post-irradiation duration. The findings demonstrated that oxylipin down-regulation was a characteristic of FLASH irradiation that was unique to normal cells. Temperature effects show that a main type of radical interacts with each other before creating peroxyl radicals, and this suggests that the flexibility and makeup of the membrane change in response to the FLASH method.

Future Prospects

In the field of radiotherapy, FLASH-RT has emerged as a popular study topic. FLASH-RT offers several benefits over regular radiotherapy, including reduced side effects, a shorter treatment duration, and a higher radiation dosage. Conventional radiation therapy often leads to a decrease in the number and function of immune cells due to their high radiation sensitivity, thereby affecting the efficacy of immunotherapy. FLASH radiation therapy, with its rapid delivery of radiation doses, has a relatively minor impact on the function and number of circulating immune cells, thereby preserving the integrity of the immune system. Studies have shown that this protective effect enables FLASH radiation therapy to achieve superior antitumor efficacy when used in combination with immune checkpoint inhibitors (such as PD-1/PD-L1 inhibitors and CTLA-4 inhibitors). ⁵¹ Despite FLASH-RT’s numerous advantages, it still faces certain challenges that require attention. For example, while it is currently thought that the difference in effect between normal and tumor tissues can guarantee anti-tumor effects while sparing the surrounding tissues, the exact mechanism of action is still unknown, and research is still needed to determine how different tissues respond to the FLASH effect in terms of dose rates. Secondly, FLASH-RT needs to be performed at very high dose rates, which cannot be met by traditional gas pedals, and the cost of clinicalization of these techniques is also a major reason limiting the development of FLASH-RT. Furthermore, the precision of standard radiotherapy image guidance is lacking in the present FLASH-RT irradiation trials. When combined with its brief pulse and high dose rate features, inaccurate positioning might have a significant negative impact on human health. If these issues remain unresolved, it will be difficult to widely deploy FLASH-RT in the clinic.

Preclinical and clinical research with FLASH-RT has produced encouraging results; a recent study revealed that treating juvenile brain tumors and potentially other malignant solid tumors with a combination of FLASH and CART radioimmunotherapy may present interesting options. ⁵² As a result, FLASH-RT is anticipated to develop into a safer and more efficient lung cancer therapy option. Consequently, investigating the impact of FLASH-RT in comparison to conventional dosage radiation treatment on pulmonary fibrosis holds significant practical significance in mitigating the pulmonary side effects of radiation therapy (Figure 1). Moreover, the degree of pulmonary fibrosis and the treatment plan selected for each patient should be taken into account when comparing the effects of FLASH-RT against conventional dosage radiation on pulmonary fibrosis. To validate the safety and efficacy of FLASH-RT, additional clinical trials and preclinical models are warranted. Table 1 provides a systematic summary of preclinical experimental models for lung research. FLASH-RT is predicted to develop into a more sophisticated and potent lung cancer treatment in the future as a result of ongoing advancements in radiation oncology as well as the collection of clinical big data, which will assist lung cancer patients even more.

Figure 1.

Summary of the Mechanisms by which Radiation-Induced Pulmonary Fibrosis and Protection by FLASH-RT

Table 1.

FLASH Radiotherapy Preclinical Research (Lung)

References	Year	Animal model	Tumor type	Irradiation particle	Total dose (Gy)	Conventional RT dose rate (Gy/s)	FLASH RT dose rate (Gy/s)	Biological effects on normal tissues
17	2014	Mice (lung)	Orthotopic lung cancer	Electron	15-17	≤0.03	≥40	Reduced pulmonary fibrosis
33	2019	Mice (lung)	Orthotopic lung cancer	Electron	17	≤0.03	>40	Mitigated DNA damage, preserved lung progenitor cells
40	2022	Healthy mice	/	Electron	30	0.1	1200	Reduced alveolar fibrosis
42	2023	Mice (lung)	Lung cancer xenograft	Proton	20	0.033	200	Attenuated acute radiation pneumonitis
46	2023	Mice (lung)	Orthotopic lung cancer	Proton	18	0.05	60	Modified immune microenvironment, immunomodulatory effects
50	2024	Mice (subcutaneous)	Lung cancer xenograft	Electron	20	/	≥100	Preserved normal pulmonary architecture
41	2025	Healthy mice	/	Electron	17.8	≤0.04	≥40	Alleviate acute injury and pulmonary fibrosis

Conclusion

FLASH-RT represents a paradigm shift in lung cancer treatment, offering a novel approach that balances tumor control with unprecedented normal tissue preservation. By delivering ultra-high dose rates (>40 Gy/s) in microseconds, FLASH-RT demonstrates dual advantages: equivalent antitumor efficacy to CONV-RT while significantly reducing radiation-induced normal tissue toxicity, particularly in mitigating acute lung injury and pulmonary fibrosis. The protective effects of FLASH-RT may come from a temporary lack of oxygen, better DNA repair in normal cells, and its ability to reduce long-term inflammation.

Comparative studies show that FLASH-RT is better at keeping lung structure intact than CONV-RT, with early research indicating significant decreases in signs of lung scarring and inflammation. Clinically, this translates to the potential for dose escalation without increasing toxicity, which could revolutionize treatment protocols for inoperable lung adenocarcinoma patients.

While promising, challenges remain. The exact molecular mechanisms underlying the FLASH effect require further elucidation through advanced omics technologies. Translational efforts must prioritize large-scale preclinical validation and first-in-human trials to establish optimal dose-fractionation parameters. Technical hurdles in equipment development—particularly compact proton FLASH systems—need resolution for clinical integration. Future directions should also explore synergies with immunotherapy and targeted agents to create multimodal treatment platforms.

In summary, FLASH-RT emerges as a transformative modality with the potential to redefine radiotherapy standards in lung cancer. Its unique ability to spare normal tissues while maintaining oncologic potency suggests that it requires continued interdisciplinary research to accelerate clinical adoption and improve patient outcomes.