Radiotherapy vs. photodynamic therapy: a comparison of antitumor effects and pulmonary toxicity in preclinical models

Introduction

Background

Interstitial lung disease (ILD) frequently co-exists with lung cancer, posing a significant treatment challenge due to the risk of life-threatening ILD exacerbations (1). Some lung cancer patients with ILD may not be suitable for surgery due to their decreased lung function. Radiotherapy (RTx), a standard alternative therapeutic modality for localized lung cancer, is often complicated by radiation pneumonitis (RP) after treatment, which can induce severe lung inflammation and respiratory failure. Pre-existing ILD is known to be a significant risk factor for developing symptomatic and severe RP in localized non-small cell lung cancer patients treated with RTx (2). Consequently, there is an urgent need for alternative treatment modalities that minimize exacerbation risks in ILD-associated localized lung cancer.

Photodynamic therapy (PDT) is a minimally invasive treatment modality for lung cancer that can be performed endoscopically (2). In PDT, a photosensitizer accumulates within the target tissue and upon exposure to light of a specific wavelength, it generates reactive species that allow for local tissue damage. We previously engineered a nanoparticle platform termed porphylipoprotein (PLP), designed to incorporate a high payload of porphyrin within high-density lipoprotein (HDL)-like nanoparticles (3). PLP features an ultrasmall nanostructure (∼20 nm) that mimics the pharmacokinetics of natural lipoproteins without requiring polyethylene glycol (PEG) coating, thereby enabling efficient tumor permeability and drug delivery. We have previously demonstrated the potent antitumor photodynamic efficacy of PLP in multiple tumor model (3-6).

Rationale and knowledge gap

Treatment options for ILD-associated localized lung cancer are limited, and the development of safe and effective modalities is urgently needed. Although alternative local therapies, such as radiofrequency ablation (RFA), microwave ablation (MWA), cryoablation (CRA), and PDT, are sometimes used for localized lung cancer cases, there is little evidence regarding their safety in ILD-associated lung cancer (2,7-10).

Objective

In this study, we investigated proof-of-concept animal studies comparing RTx vs. PDT for (I) anti-tumor effects in a mouse xenograft model; and (II) pulmonary toxicity in a rat ILD model. We present this article in accordance with the ARRIVE reporting checklist (available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-2025-662/rc).

Methods

Please see Appendix 1 for details. A protocol was prepared before the study without registration. All animal experiments were performed under a project license (No. AUP 4151, 6330) granted by the Animal Care Committee of University Health Network, Toronto, Canada, in compliance with the institutional guidelines for the care and use of animals.

Mouse xenograft model

Athymic NCr-Foxn1^nu mice underwent unilateral subcutaneous inoculation into the thigh with human A549 or H460 lung cancer cells. Tumor growth was monitored until they reached 8–12 mm in diameter. A PLP biodistribution study was conducted to evaluate the optimal timeline for treatment of the PDT group. PLP fluorescence was measured with the Xenogen-IVIS imaging system (Perkin Elmer, Massachusetts, USA). The ratio of fluorescence in the tumor to the contralateral thigh muscle was measured. Mice were then divided into three groups: control [n=8 (for A549 xenograft)/12 (for H460 xenograft)], RTx (n=10/8), and PDT (n=12/8)). The RTx group received a single 20 Gy local irradiation (10 Gray anterior to posterior, 10 Gray posterior to anterior) using the SmART+ system (Precision X-Ray, Connecticut, USA). The PDT group received an intravenous PLP (4 mg/kg) 24 hours before treatment, followed by a light dose of 100 J/cm² (at 671 nm). Mice were sacrificed 7 days after the intervention, and excised tumors were evaluated for pathology with hematoxylin and eosin (H&E) staining for morphology, Ki-67 staining for viability, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining for apoptosis.

Rat ILD model

Immunocompetent Sprague-Dawley rats were given a single intratracheal injection (2 mg/kg) of bleomycin (BLM) or vehicle control and were monitored for 3 weeks to track the development of interstitial lung inflammation. Rats were then divided into four groups: control [n=7 (for autopsy at week 7)/8 (at week 15)], BLM (n=5/7), BLM + RTx (n=5/6), and BLM + PDT (n=4/9). The RTx group received a single 20 Gy irradiation (10 Gray anterior to posterior, 10 Gray posterior to anterior) by SmART+ systems to the left lung base. The PDT group received PLP administration (4 mg/kg) 24 hours before they were intubated and maintained with ventilatory management. An ultrathin (0.2 mm outer diameter) optical fiber was delivered to the left lung base under computed tomography (CT) guidance followed by a light dose of 100 J/cm² (at 671 nm). Chest CT was evaluated monthly and an autopsy was done after 1 month (at week 7) or 3 months (at week 15) follow-up. Bronchoalveolar lavage (BAL) and lung tissue were sampled. Lung tissue was also measured for fluorescence intensity in Xenogen-IVIS. Lung pathology was quantified by Ashcroft fibrosis score (11) with H&E staining and Masson’s staining. For bulk RNA sequencing (RNA-seq), RNA was extracted from homogenized rat lung samples, and libraries were sequenced on a Novaseq X (Illumina, San Diego, CA, USA) to generate a minimum of 30 million paired-end 150 bp reads per sample. FASTQC (v0.12.1) was used to assess the quality control of the raw sequencing. Transcript quantification was performed using Salmon (v1.10.3). DESeq2 (v1.44.0) was used to obtain the differential expression of the quantified transcript abundance. Genes with a P-adjusted value <0.05 were considered significantly differentially expressed.

Statistical analysis

The experimental data was analyzed with software GraphPad Prism 10.3 (San Diego, CA, USA). Comparative statistical analysis was performed by Student’s t-test between two groups and by one-way analysis of variance (ANOVA) between more than two groups. P<0.05 was considered statistically significant unless otherwise indicated (***, P<0.001; **, P<0.01; *, P<0.05).

Results

Mouse xenograft model

Biodistribution of PLP in mouse xenograft model

PLP biodistribution showed the best tumor-to-contralateral background muscle ratio at 24 hours post-injection in both A549 (Figure 1A,1B) and H460 (Figure S1A,S1B) xenograft tumors. In evaluating the organ biodistribution, the fluorescence intensity in the liver was significantly higher than in the tumor, whereas the heart, kidney, and colon showed significantly lower intensities. The signal in the tumor was higher than that of the skin and lung, but the difference was not statistically significant (Figure S1C).

Figure 1 Biodistribution of PLP in the mouse tumor model. (A) Biodistribution of PLP overtime in A549 lung cancer cell line tumor [before administration (t0), 1, 6, 12, and 24 hours after administration]. (B) Tumor-to-background muscle fluorescence intensity ratio overtime in A549 lung cancer cell line tumor (n=5). PLP, porphylipoprotein.

Tumor efficacy of RTx vs. PDT in mouse xenograft model

A schematic of the study schedule is shown in Figure 2A (image A1) and the RTx and PDT setups are shown in Figure 2A (image A2). Before intervention (RTx or PDT), there was no significant difference in tumor size between the three groups: control, RTx, and PDT (Figure 2B, image B1). PDT group showed a significant reduction in A549 xenograft tumor volume compared to control or RTx group at 3 and 7 days after intervention (Figure 2B, image B1). Regarding the percent change in A549 tumor volume (day 0 → day 7), both RTx and PDT showed a reduction trend compared to the control (Figure 2B, image B2). In H460 tumors, PDT showed a significant volume reduction compared to the control group on day 3, while there was no significant difference between RTx and the control group. On day 7, both RTx and PDT showed a significant tumor volume reduction compared to the control group, while there was no significant difference between RTx and PDT (Figure S2A).

Figure 2 RTx vs. PDT: anti-tumor effect in the mouse tumor model. (A) (A1) Schematic diagram showing the schedule of the mouse xenograft model. (A2) Setups for both RTx and PDT in the mouse xenograft model. (B) A549 xenograft tumor volume (mm³) in (B1) day 0, day 3, and day 7. (B2) Tumor volume changing rate from day 0 to day 7 (%) [n=8 (control), 10 (RTx), 12 (PDT)]. (C) (C1) Histology of A549 xenograft tumor at day 7 (H&E, Ki-67, TUNEL). (C2) H-score for Ki-67 staining [n=8 (control), 10 (RTx), 12 (PDT)]. (C3) H-score for TUNEL staining [n=8 (control), 10 (RTx), 12 (PDT)]. ns, nonspecific; *, P<0.05; ***, P<0.001. A, anterior; H&E, hematoxylin and eosin; P, posterior; PDT, photodynamic therapy; PLP, porphylipoprotein; RTx, radiotherapy; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.

The pathology of the A549 tumor removed at autopsy on day 7 is shown in Figure 2C (image C1). Ki-67 staining H-scores were significantly reduced for RTx and PDT compared to the control group (Figure 2C, image C2), with no significant difference between the two treatment groups. PDT showed a depth-dependent effect, displaying stronger Ki-67-staining on the tumor side distant to the laser irradiation surface (Figure S2B, images B1 and B2). TUNEL staining H-scores were significantly elevated in the PDT group compared to the RTx and the control groups (Figure 2C, image C3). Similar to Ki-67, TUNEL staining showed a depth-dependent effect in PDT, with more positive cells on the light-incident side of the tumor (Figure S2B).

Rat ILD model

Biodistribution of PLP in rat ILD model

Regarding the lung biodistribution of PLP, significantly higher fluorescence intensity was observed on day 1 than (uninfused) control or day 7 in (BLM-untreated) control lungs (Figure 3A,3B). While no significant difference in fluorescence intensity was observed in the control- or BLM-lungs at day 1 (Figure 3C).

Figure 3 Biodistribution of PLP in the rat ILD lung model. (A) Lung distribution of PLP in rat model. (B) Fluorescence intensity of PLP in control (BLM-untreated) lung [n=6 (day 0), 7 (day 1), 6 (day 7)]. (C) Fluorescence intensity of PLP at day 1 [n=6 (control lung without PLP), 5 (BLM lung without PLP), 7 (control lung with PLP), 5 (BLM lung with PLP)]. ns, nonspecific; *, P<0.05; ***, P<0.001. BLM, bleomycin; ILD, interstitial lung disease; PLP, porphylipoprotein.

Pulmonary inflammation on CT over time

A schematic of the study schedule is shown in Figure 4A. The rat lung RTx irradiation field and PDT setup are shown in Figure S3A,S3B. The results of lung inflammation monitoring on chest CT were shown in Figure 4B. Control rats showed no evidence of lung inflammation throughout the monitoring, while the BLM only group showed inflammatory infiltrates in the bilateral lung bases, peaking at 3 weeks after BLM administration (arrows, week 3), which then spontaneously disappeared and became less noticeable at week 7. Similar to the BLM only group, rats treated with RTx 3 weeks after BLM (BLM + RTx rats) showed bilateral lung basement infiltrates at week 3 (arrows) which once attenuated at week 7, but then the lung infiltrates within the RTx irradiation site gradually increased by week 15 (3 months after RTx). In contrast, rats that underwent PDT 3 weeks after BLM administration (BLM + PDT rats) also showed bilateral lung basement infiltrates at week 3 (arrows), and further enhanced left lung basement inflammation after PDT to the left lung basement, but the pulmonary infiltrates gradually attenuated and a more reduced shade of the left lung base can be identified at week 15 (arrows, 3 months after PDT). Three-dimensional (3D)-recomposition of CT images and measurement of volume (Figure S4A) showed that the BLM + RTx rats had significantly increased infiltrate lung volume (Figure S4B, image B1) and increased percentage of the total lung volume (Figure S4B, image B2) than the BLM + PDT rats at week 15.

Figure 4 RTx vs. PDT: chest CT findings in the rat ILD lung model. (A) Schematic diagram showing the schedule of the rat ILD model. (B) Rat chest CT findings at weeks 0, 3, 7, and 15 (arrows: lung infiltration in chest CT). BAL, bronchoalveolar lavage; BLM, bleomycin; CT, computed tomography; ILD, interstitial lung disease; i.t., intratracheal; PDT, photodynamic therapy; PLP, porphylipoprotein; RTx, radiotherapy.

BAL and blood leukocyte fraction analysis at week 7

For the results of leukocyte fractionation of BAL at week 7, there were no significant differences between the control, BLM, BLM + RTx, and BLM + PDT groups for all fractions of total leukocytes, lymphocytes, neutrophils, and macrophages (Figure S5A). Similarly, there were no significant differences among the control, BLM, BLM + RTx, and BLM + PDT groups in total leukocytes, lymphocytes, and neutrophils in all fractions of blood leukocyte fractionation results in week 7 (Figure S5B).

Lung inflammation and fibrosis at week 15

Leukocyte fractionation of BAL at week 15 showed a significant increase in cell count for BLM + RTx compared to control, BLM, and BLM + PDT in total leukocyte, neutrophil, and macrophage fractions (Figure 5A). For BLM + PDT, there was no significant difference in each fraction compared to the control. On the other hand, the results of blood leukocyte fractionation during week 15 showed no significant differences among the control, BLM, BLM + RTx, and BLM + PDT groups for all fractions of total leukocytes, lymphocytes, and neutrophils (Figure S5C).

Figure 5 RTx vs. PDT: leukocyte analysis in BAL and lung pathology at week 15. (A) Total leukocyte, lymphocyte, neutrophil, and macrophage count in BAL at week 15 [n=8 (control), 7 (BLM), 6 (BLM + RTx), 9 (BLM + PDT)]. (B) (B1) Lung pathology (HE and Masson’s staining) of BLM + RTx and BLM + PDT rats at week 15. (B2) Lung Ashcroft fibrosis score for lung pathology at week 15 [n=7 (control), 8 (BLM), 6 (BLM + RTx), 7 (BLM + PDT)]. ns, nonspecific; *, P<0.05; **, P<0.01; ***, P<0.001. BAL, bronchoalveolar lavage; BLM, bleomycin; HE, hematoxylin-eosin; PDT, photodynamic therapy; RTx, radiotherapy.

Lung pathology at week 15 showed extensive inflammatory cell infiltration and fibrosis in BLM + RT group, while only localized inflammation and fibrosis were seen in BLM + PDT group (Figure 5B, image B1). In Ashcroft’s lung fibrosis score, BLM, BLM + RTx, and BLM + PDT all showed significant score increases over control, however, BLM + RTx showed a larger increase and a statistically significant increase compared to the BLM + PDT group (Figure 5B, image B2). In the bulk-RNA seq analysis in the explanted lungs at week 15, a total of 15,793 genes were used for subsequent analysis which are depicted in the volcano plot in Figure 6. Using a false discovery rate cut-off of log₂ fold change ≥1, there are 86 differentially expressed genes (DEGs) (52 up-regulated and 34 down-regulated) in the comparison of BLM + RTx lung and BLM + PDT lung samples. Of these genes, Slfn4, O6, Apold1, Svep1, and Ly6a are the top-ranked genes with log₁₀fold-change greater than 10. In the analysis for identifying the enriched Gene Ontology terms corresponding to biological processes (GO-BP), anatomical structure development and cell differentiation were enriched in both BLM + RTx or BLM + PDT lung, while cell adhesion, immune system process, and cytokine production were enriched in BLM + RTx lung (Figure S6A,S6B).

Figure 6 RTx vs. PDT: bulk RNA-seq analysis for lungs at week 15. Volcano plot for enriched gene expression in bulk RNA-seq analysis for lungs at week 15 [n=2 (BLM + RTx), 2 (BLM + PDT)]. BLM, bleomycin; FC, fold change; NS, nonspecific; PDT, photodynamic therapy; RNA-seq, RNA sequencing; RTx, radiotherapy.

Discussion

Key findings

PLP distribution showed a favorable tumor-to-background ratio (Figure 1A,1B) and no difference in distribution between healthy and ILD lungs (Figure 3A,3C). PDT showed a better antitumor effect compared to RTx in both A549 xenograft (Figure 2B) and non-inferior antitumor effect H460 xenograft (Figure S2A). In the rat ILD model, longitudinal monitoring with chest CT showed that BLM + RTx rats exhibited increased lung infiltrates at week 15 compared to BLM + PDT rats (Figure 4B, Figure S4B). PDT showed milder inflammatory findings in leukocyte cell fractionation of BAL (Figure 5A) and also a milder fibrosis profile in lung histology scoring than RTx (Figure 5B) at week 15 in the rat ILD lung model.

Strengths and comparison of similar researches

To our knowledge, this is the first report to directly compare the long-term pulmonary toxicity effects of RTx and PDT in an animal ILD model. Previously, Saito et al. compared a BLM-induced rat ILD model, with and without PDT using talaporfin sodium as a nanoparticle, and reported no difference in Ashcroft lung fibrosis score or macrophage infiltration at day 7 post-PDT (12). However, their study focused on the acute phase, where PDT was performed at 1 week after BLM administration, and dissection and analysis were performed 2 weeks after BLM administration, thus leaving long-term fibrosis effects unaddressed. In addition, in their study, they performed PDT through external illumination of the lung after opening the chest. In contrast, our study extended the observation period to 3 months, to capture potential late-stage fibrotic changes. We also introduced a head-to-head comparison with RTx, the current standard of care, to determine whether PDT offers safety advantages in an ILD setting.

Explanations of findings

First, we confirmed that the chosen PDT dose could achieve similar antitumor efficacy compared to the chosen RTx dose. Having established baseline antitumor equivalent doses between two modalities, we examined their long-term toxicity effects in a rat ILD model. Over the 15-week observation period, the BLM + RTx rats exhibited more persistent lung inflammation and fibrosis at the treatment site, whereas the BLM + PDT rats showed an initial inflammatory response that gradually resolved (Figure 4B, Figure S4B).

To further characterize these long-term differences, we performed bulk RNA-seq on the rat lung tissue at week 15. Gene expression of Rab1b, a GTPase that regulates vesicular transport, and Ly6a, a member of the lymphocyte Ag 6 family, was relatively upregulated in week 15 BLM + PDT rat lungs (Figure 6). Rab1b is involved in endoplasmic reticulum to Golgi transport, which is important in interferon beta secretion (13). Ly6a expression is induced by interferon-alpha/gamma and promotes T-cell activation (14). Interferons are known to promote cytokine secretion through the activation of macrophages and lymphocytes in wound healing, which promotes infection protection and tissue healing (15). This is in line with the known post-PDT response from the literature, which shows that the irradiated tissue is characterized by activation and re-epithelialization of epidermal stem cells, remodeling of the extracellular matrix (ECM), and promotion of angiogenesis (16,17).

Conversely, BLM + RTx rats showed an increased number of neutrophils and macrophages in the BAL than control rats at week 15 (Figure 5A). In addition, lung bulk RNA-seq at week 15 showed that the expression of Slfn4, Svep1, and Apold1 was relatively upregulated in BLM + RTx (Figure 6). GO-BP pathway analysis indicated enriched cytokine production, leukocyte migration, cell-matrix adhesion, multicellular organismal process, and cell motility in BLM + RTx lung (Figure S6A). Slfn4 is a Schlafen family protein, a myeloid cell differentiation factor, and known to be responsible for the regulation of macrophage function (18). ECM protein Svep1 is involved in airway patterning and alveolar formation during lung development (19). Apolipoprotein APOLD1 is involved in the adhesive junctions of vascular endothelial cells and the maintenance of cytoskeletal structure (20). These contexts are in line with RP pathogenesis, which involves DNA damage by radiation-produced reactive oxygen species (ROS), activation of innate immunity by damage-associated molecular patterns (DAMPs), matrix metalloproteinase secretion to accumulate ECM by neutrophil, and profibrotic cytokine secretion by macrophage (21-23).

These DEGs suggest that the type of physical injury (RTx or PDT) applied to the ILD lung may alter subsequent local lung tissue repair and immune response, and may be important targets for treatment strategies for ILD-associated lung cancer with a low risk of ILD exacerbation.

Limitations

There are some limitations in this study. First, we did not use an ILD-associated lung cancer animal model, limiting us to evaluate antitumor effects in a mouse xenograft model and lung toxicity in a rat ILD model separately. However, inoculating human cancer cells into experimental animals requires immunocompromised hosts, while evaluating the lung inflammation of ILDs requires immunocompetent animals. We believe that our model is one possible solution to these conflicting requirements. By adopting syngeneic murine lung cancer models using mouse lung cancer cell lines such as Lewis lung carcinoma and CMT167, we could consider RTx and PDT on immunocompetent mice bearing tumor (24,25). However, in the present study, we limited the scope to establish a reliable catheter-based intrapulmonary light-delivery workflow and to define a baseline pulmonary toxicity profile in healthy lung tissue. Our current instrumentation limits our access to rat-scale airway.

Second, we only investigated the single dosage variation (20 Gy/single fraction for RTx and 100 J/cm² for PDT). The treatment doses in both RTx and PDT were not curative doses and were single-fractioned. The 20 Gy irradiation used in the RTx in this study is close to a clinical dose for cancer symptom palliation (26,27). In fact, Ki67 staining of tumors in the mouse Xenograft model (A549) is not completely negative for either RTx or PDT (Figure 2C, image C2). The tumor treated with PDT also showed minimal Ki-67 staining at sites closer to the laser irradiation, whereas Ki-67-positive cells were more prominent at more distal sites (Figure S2B); this is consistent with the known technical limitation of PDT, in which the lack of light reach resulting in difficulty for the treatment to penetrate deep into the irradiated area (28,29). In practice, the fact that lung cancers located far from the bronchus cannot be treated represents a significant limitation in the clinical application of this research. On the other hand, peripheral lung cancer located near the bronchi is also a common condition. Although it is important to understand the limitations of PDT’s applicability, this does not significantly diminish its clinical usefulness. Also, both RTx and PDT have abundant evidence of application for symptom palliation (2,27,30,31). Since there is a practical clinical need for palliative lung cancer treatment for ILD-associated lung cancer, this study has a sufficient rationale. Furthermore, in the RTx group, although a single 20 Gy dose is well-established and widely used in the field of radiation-induced lung injury research (32,33), it also represents a high radiation dose for the lungs especially in the setting of BLM-induced ILD. In this study, rats were anesthetized with isoflurane inhalation and maintained in a spontaneously breathing setup, allowing us to successfully ensure precise irradiation fields with a single dose. However, it would be beneficial to conduct a comparative study of lung toxicity between RTx and PDT using fractionated therapy, which is expected to reduce toxicity, in the future.

Third, the treatment delivery methods differ between the two models: RTx was delivered via external beam, whereas PDT was delivered through an ultrafine fiber. Each approach presents unique limitations. In RTx, respiratory motion causes displacement of the lung tissue during irradiation, leading to potential variation in dose distribution. It would be desirable to use a catheter-based treatment, such as brachytherapy, to create a more comparable model for lung toxicity in future studies. PDT is subject to similar motion-related variability, compounded by a more limited treatment field, which we attempted to address by incremental repositioning of the fiber. Additionally, due to anatomical constraints of rat bronchus, the fiber tip was positioned in close proximity to the bronchial mucosa, likely resulting in a high local power density that may have induced thermal damage. We did not monitor the local tissue temperature during PDT, and therefore any potential thermal contribution to the lung injury remains uncertain (34).

Lastly, while the sample size of n=2 per group in the bulk RNA-seq analysis is too small to ensure statistical power and reliability, the transcriptome profiles of lung tissue at week 15 differed significantly between the RTx group and the PDT group, revealing a stronger inflammatory and fibrotic profile in the RTx group. This result was intriguing and consistent with the BAL analysis and lung pathology findings. However, definitive conclusions cannot be drawn based solely on the analyses in this study, and more conclusive findings should be pursued in future studies with larger sample sizes.

Despite these limitations, PDT demonstrated a more favorable pulmonary toxicity profile in ILD tissue compared to RTx, supporting its potential as a safer therapeutic alternative for ILD-associated lung cancer. As this study serves as a proof-of-concept in a small animal model, further investigations are required to validate the results. Future studies should incorporate validated dosing parameters and refined treatment delivery methods for both RTx and PDT. Temperature monitoring and other dosage delivery methods should also be considered to confirm that any unexpected damage to the lung remains minimal.

Conclusions

We established two preclinical animal models comparing RTx and PDT. First, we established a dose of PLP-mediated PDT that achieved comparable antitumor efficacy to RTx in a mouse xenograft model. Second, in the rat ILD model, although consideration must be given to the differences in dose delivery between RTx and PDT, PDT was associated with a safer pulmonary toxicity profile. Lung pathology and bulk RNA-seq analysis supported these findings, revealing less immune-cell infiltration and milder fibrotic changes in PDT-treated lungs compared to RTx. These results suggest that PDT is possibly a promising alternative treatment modality for ILD-associated lung cancer cases.

Acknowledgments

We would like to thank Judy McConnell (Toronto General Hospital) for laboratory management, Teesha Komal (University Health Network-STTARR) for technical assistance with imaging, Napoleon Law (University Health Network-STTARR pathology) for technical assistance with histology, as well as Maria Monroy and the UHN Animal Resources Centre.

Footnote

Reporting Checklist: The authors have completed the ARRIVE reporting checklist. Available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-2025-662/rc

Data Sharing Statement: Available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-2025-662/dss

Peer Review File: Available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-2025-662/prf

Funding: This work was supported by the William Coco Chair in Surgical Innovation for Lung Cancer, a joint hospital-university chair between University Health Network, the University of Toronto, Toronto General and Western Hospital Foundation, the National Sanitarium Association 2017 Innovative Research Program (to K.Y.), the Canadian Cancer Society Impact Grant (No. #704123, to K.Y.), and the Terry Fox Research Institute New Frontiers Program (No. #1137).

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-2025-662/coif). Y.H. was financially supported by Uehara foundation overseas research fellowships (No. 201940067). K.Y. received funding from the National Sanitarium Association 2017 Innovative Research Program and the Canadian Cancer Society Impact Grant (No. #704123). The other authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All animal experiments were performed under a project license (No. AUP 4151, 6330) granted by the Animal Care Committee of University Health Network, Toronto, Canada, in compliance with the institutional guidelines for the care and use of animals.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

References

1. Ogura T, Takigawa N, Tomii K, et al. Summary of the Japanese Respiratory Society statement for the treatment of lung cancer with comorbid interstitial pneumonia. Respir Investig 2019;57:512-33. [Crossref] [PubMed]
2. Crous A, Abrahamse H. Photodynamic therapy of lung cancer, where are we? Front Pharmacol 2022;13:932098. [Crossref] [PubMed]
3. Cui L, Lin Q, Jin CS, et al. A PEGylation-Free Biomimetic Porphyrin Nanoplatform for Personalized Cancer Theranostics. ACS Nano 2015;9:4484-95. [Crossref] [PubMed]
4. Muhanna N, Cui L, Chan H, et al. Multimodal Image-Guided Surgical and Photodynamic Interventions in Head and Neck Cancer: From Primary Tumor to Metastatic Drainage. Clin Cancer Res 2016;22:961-70. [Crossref] [PubMed]
5. Ujiie H, Ding L, Fan R, et al. Porphyrin-High-Density Lipoprotein: A Novel Photosensitizing Nanoparticle for Lung Cancer Therapy. Ann Thorac Surg 2019;107:369-77. [Crossref] [PubMed]
6. Lou J, Aragaki M, Bernards N, et al. Repeated porphyrin lipoprotein-based photodynamic therapy controls distant disease in mouse mesothelioma via the abscopal effect. Nanophotonics 2021;10:3279-94. [Crossref] [PubMed]
7. Zhao ZR, Lau RWH, Ng CSH. Catheter-based alternative treatment for early-stage lung cancer with a high-risk for morbidity. J Thorac Dis 2018;10:S1864-70. [Crossref] [PubMed]
8. Brace CL. Radiofrequency and microwave ablation of the liver, lung, kidney, and bone: what are the differences? Curr Probl Diagn Radiol 2009;38:135-43. [Crossref] [PubMed]
9. Matthews W, Rizzoni W, Mitchell J, et al. In vitro photodynamic therapy of human lung cancer. J Surg Res 1989;47:276-81. [Crossref] [PubMed]
10. Erinjeri JP, Clark TW. Cryoablation: mechanism of action and devices. J Vasc Interv Radiol 2010;21:S187-91. [Crossref] [PubMed]
11. Ashcroft T, Simpson JM, Timbrell V. Simple method of estimating severity of pulmonary fibrosis on a numerical scale. J Clin Pathol 1988;41:467-70. [Crossref] [PubMed]
12. Saito Y, Imai K, Furumoto H, et al. Effect of photodynamic therapy (PDT) on a rat model of bleomycin-induced interstitial pneumonia. Photodiagnosis Photodyn Ther 2022;37:102659. [Crossref] [PubMed]
13. Beachboard DC, Park M, Vijayan M, et al. The small GTPase RAB1B promotes antiviral innate immunity by interacting with TNF receptor-associated factor 3 (TRAF3). J Biol Chem 2019;294:14231-40. [Crossref] [PubMed]
14. Maliah A, Santana-Magal N, Parikh S, et al. Crosslinking of Ly6a metabolically reprograms CD8 T cells for cancer immunotherapy. Nat Commun 2024;15:8354. [Crossref] [PubMed]
15. Zhang LJ. Type1 Interferons Potential Initiating Factors Linking Skin Wounds With Psoriasis Pathogenesis. Front Immunol 2019;10:1440. [Crossref] [PubMed]
16. Grandi V, Corsi A, Pimpinelli N, et al. Cellular Mechanisms in Acute and Chronic Wounds after PDT Therapy: An Update. Biomedicines 2022;10:1624. [Crossref] [PubMed]
17. Allison RR, Moghissi K. Photodynamic Therapy (PDT): PDT Mechanisms. Clin Endosc 2013;46:24-9. [Crossref] [PubMed]
18. van Zuylen WJ, Garceau V, Idris A, et al. Macrophage activation and differentiation signals regulate schlafen-4 gene expression: evidence for Schlafen-4 as a modulator of myelopoiesis. PLoS One 2011;6:e15723. [Crossref] [PubMed]
19. Foxworth N, Wells J, Ocaña-Lopez S, et al. Svep1 orchestrates distal airway patterning and alveolar differentiation in murine lung development. eLife 2024;13:RP100443.
20. Stritt S, Nurden P, Nurden AT, et al. APOLD1 loss causes endothelial dysfunction involving cell junctions, cytoskeletal architecture, and Weibel-Palade bodies, while disrupting hemostasis. Haematologica 2023;108:772-84. [Crossref] [PubMed]
21. Rahi MS, Parekh J, Pednekar P, et al. Radiation-Induced Lung Injury-Current Perspectives and Management. Clin Pract 2021;11:410-29. [Crossref] [PubMed]
22. Giuranno L, Ient J, De Ruysscher D, et al. Radiation-Induced Lung Injury (RILI). Front Oncol 2019;9:877. [Crossref] [PubMed]
23. Yan Y, Fu J, Kowalchuk RO, et al. Exploration of radiation-induced lung injury, from mechanism to treatment: a narrative review. Transl Lung Cancer Res 2022;11:307-22. [Crossref] [PubMed]
24. Li HY, McSharry M, Bullock B, et al. The Tumor Microenvironment Regulates Sensitivity of Murine Lung Tumors to PD-1/PD-L1 Antibody Blockade. Cancer Immunol Res 2017;5:767-77. [Crossref] [PubMed]
25. Weiser-Evans MC, Wang XQ, Amin J, et al. Depletion of cytosolic phospholipase A2 in bone marrow-derived macrophages protects against lung cancer progression and metastasis. Cancer Res 2009;69:1733-8. [Crossref] [PubMed]
26. Kepka L, Socha J. Dose and fractionation schedules in radiotherapy for non-small cell lung cancer. Transl Lung Cancer Res 2021;10:1969-82. [Crossref] [PubMed]
27. Jumeau R, Vilotte F, Durham AD, et al. Current landscape of palliative radiotherapy for non-small-cell lung cancer. Transl Lung Cancer Res 2019;8:S192-201. [Crossref] [PubMed]
28. Gunaydin G, Gedik ME, Ayan S. Photodynamic Therapy-Current Limitations and Novel Approaches. Front Chem 2021;9:691697. [Crossref] [PubMed]
29. Huis In 't Veld RV, Heuts J, Ma S, et al. Current Challenges and Opportunities of Photodynamic Therapy against Cancer. Pharmaceutics 2023;15:330. [Crossref] [PubMed]
30. Diaz-Jiménez JP, Martínez-Ballarín JE, Llunell A, et al. Efficacy and safety of photodynamic therapy versus Nd-YAG laser resection in NSCLC with airway obstruction. Eur Respir J 1999;14:800-5. [Crossref] [PubMed]
31. Moghissi K, Dixon K, Stringer M, et al. The place of bronchoscopic photodynamic therapy in advanced unresectable lung cancer: experience of 100 cases. Eur J Cardiothorac Surg 1999;15:1-6. [Crossref] [PubMed]
32. Jin H, Yoo Y, Kim Y, et al. Radiation-Induced Lung Fibrosis: Preclinical Animal Models and Therapeutic Strategies. Cancers (Basel) 2020;12:1561. [Crossref] [PubMed]
33. Wang J, Zhou F, Li Z, et al. Pharmacological targeting of BET proteins attenuates radiation-induced lung fibrosis. Sci Rep 2018;8:998. [Crossref] [PubMed]
34. Overchuk M, Weersink RA, Wilson BC, et al. Photodynamic and Photothermal Therapies: Synergy Opportunities for Nanomedicine. ACS Nano 2023;17:7979-8003. [Crossref] [PubMed]