Toluidine Blue-Loaded Nanomicelles for Photodynamic Therapy of Lung Cancer: An In-vitro Study
-
Bahareh Khalili Najafabad
,
Mehdi Hoseini
,
Amirhossein Rashnoodi
,
Saleh Rouhi
,
Soroush Najafisarvestani
,
Zeynab Seraj
,
Ghazal Sadraei
Abstract
Abstract
Introduction: Photodynamic therapy (PDT) has attracted considerable attention as a non-invasive and selective method for cancer treatment. In this study, the photodynamic effects of nanomicelles loaded with the dye toluidine blue (TB) were evaluated on human lung cancer (QU-DB) and non-cancerous (MRC-5) cell lines. Nanomicelles, as biocompatible carriers, possess the ability to improve solubility, stability, and targeted accumulation of a photosensitizer (PS), while showing enhanced tumor tissue accumulation through the enhanced permeability and retention (EPR) effect.
Methods: The TB-loaded nanomicelles were synthesized using the direct dissolution method and underwent physicochemical characterization. Then, cells were irradiated with laser red light (λ = 655 nm) at doses of 10, 20, and 40 J/cm². Post-irradiation, cell viability was quantified via the MTT assay, enabling the calculation of key therapeutic indices such as IC₅₀, ED₅₀, and the synergy index (SI).
Results: The results demonstrated that encapsulating TB in nanomicelles increased optical absorption, reduced dye aggregation, and consequently enhanced reactive oxygen species (ROS) generation efficiency. The photodynamic effect of TB-loaded nanomicelles was significantly stronger in the QU-DB cancer cell line compared with free TB, while inducing lower toxicity in the non-tumor MRC-5 cells. Thus, the selective distinction between tumor and non- cancerous cell lines was confirmed. Furthermore, increasing light dose resulted in greater cytotoxicity, with dose-dependent reductions in cell survival. However, no favorable selectivity index was obtained when comparing micellar drug cytotoxicity with the free PS (p > 0.05).
Conclusion: TB-loaded nanomicelles represent a promising drug delivery system to enhance the efficacy and selectivity of PDT in lung cancer while reducing its off-target effects.
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29- Xiaoxiao Gu, Haoyang Yuan, Cailing Li, Lixian Xu, Shuang Li, and Dinghua Yu, "Toluidine blue O photosensitizer combined with caffeic acid improves antibacterial performance by increasing the permeability of cell membrane." Colloids and Surfaces B: Biointerfaces, Vol. 233p. 113657, (2024).
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31- Parinaz Mehnati, Reza Malekzadeh, Mohammad Yousefi Sooteh, and Soheila Refahi, "Assessment of the efficiency of new bismuth composite shields in radiation dose decline to breast during chest CT." The Egyptian Journal of Radiology and Nuclear Medicine, Vol. 49 (No. 4), pp. 1187–89, (2018).
32- Nikan Asadpour, Reza Malekzadeh, Saeed Rajabpour, Soheila Refahi, Parinaz Mehnati, and Ahmad Shanei, "Shielding performance of multi-metal nanoparticle composites for diagnostic radiology: an MCNPX and Geant4 study." Radiological Physics and Technology, Vol. 16 (No. 1), pp. 57–68, (2023).
33- Behnaz Babaye Abdollahi et al., "Main approaches to enhance radiosensitization in cancer cells by nanoparticles: A systematic review." Advanced pharmaceutical bulletin, Vol. 11 (No. 2), p. 212, (2020).
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35- Junjie Liu, Jinjin Shi, Weimin Nie, Sijie Wang, Genhua Liu, and Kaiyong Cai, "Recent progress in the development of multifunctional nanoplatform for precise tumor phototherapy." Advanced healthcare materials, Vol. 10 (No. 1), p. 2001207, (2021).
36- Lama Misba, Hayder Abdulrahman, and Asad U Khan, "Photodynamic efficacy of toluidine blue O against mono species and dual species bacterial biofilm." Photodiagnosis and Photodynamic Therapy, Vol. 26pp. 383–88, (2019).
2- Bagher Farhood, Behzad Raei, Reza Malekzadeh, Maryam Shirvani, Masoud Najafi, and Tohid Mortezazadeh, "A review of incidence and mortality of colorectal, lung, liver, thyroid, and bladder cancers in Iran and compared to other countries." Contemporary Oncology/Współczesna Onkologia, Vol. 23 (No. 1), pp. 7–15, (2019).
3- Sivapar V Mathan, Mohit Rajput, and Rana P Singh, "Chemotherapy and radiation therapy for cancer." in Understanding cancer: Elsevier, (2022), pp. 217–36.
4- Dennis EJGJ Dolmans, Dai Fukumura, and Rakesh K Jain, "Photodynamic therapy for cancer." Nature reviews cancer, Vol. 3 (No. 5), pp. 380–87, (2003).
5- A Ferreira Dos Santos, DR Queiroz de Almeida, Leticia Ferreira Terra, Maurício S Baptista, and Leticia Labriola, "Photodynamic therapy in cancer treatment-an update review." J. cancer metastasis Treat, Vol. 5 (No. 25), p. 10.20517, (2019).
6- Omid Talaee et al., "Characterization of a HER-2 targeted silica-gold nanoshells for breast cancer radio-photothermal therapy." Applied Radiation and Isotopes, p. 112454, (2026).
7- Tae Eun Kim and Ji-Eun Chang, "Recent studies in photodynamic therapy for cancer treatment: from basic research to clinical trials." Pharmaceutics, Vol. 15 (No. 9), p. 2257, (2023).
8- Stefan W Ryter et al., "Mechanisms of cell death in oxidative stress." Antioxidants & redox signaling, Vol. 9 (No. 1), pp. 49–89, (2007).
9- Srivalleesha Mallidi, Sriram Anbil, Anne-Laure Bulin, Girgis Obaid, Megumi Ichikawa, and Tayyaba Hasan, "Beyond the barriers of light penetration: strategies, perspectives and possibilities for photodynamic therapy." Theranostics, Vol. 6 (No. 13), p. 2458, (2016).
10- Mohammad Ali Mozayeni, Farzaneh Vatandoost, Mohammad Asnaashari, Mehdi Shokri, Saranaz Azari-Marhabi, and Negin Asnaashari, "Comparing the efficacy of toluidine blue, methylene blue and curcumin in photodynamic therapy against Enterococcus faecalis." Journal of lasers in medical sciences, Vol. 11 (No. Suppl 1), p. S49, (2020).
11- Benedicto de Campos Vidal and Maria Luiza S Mello, "Toluidine blue staining for cell and tissue biology applications." Acta histochemica, Vol. 121 (No. 2), pp. 101–12, (2019).
12- Lama Misba, Shatavari Kulshrestha, and Asad U Khan, "Antibiofilm action of a toluidine blue O-silver nanoparticle conjugate on Streptococcus mutans: a mechanism of type I photodynamic therapy." Biofouling, Vol. 32 (No. 3), pp. 313–28, (2016).
13- Bahman Alipour et al., "Advances in nano-scale metal-based contrast agents for computed tomography: a systematic review." Radiation Physics and Chemistry, Vol. 226p. 112195, (2025).
14- Yasin Ayyami, Marjan Ghorbani, Masoumeh Dastgir, Reza Malekzadeh, and Tohid Mortezazadeh, "Chitosan-modified manganese oxide-conjugated methotrexate nanoparticles delivering 5-aminolevulinic acid as a dual-modal T1–T2* MRI contrast agent in U87MG cell detection." Magnetic resonance materials in physics, biology and medicine, Vol. 37 (No. 5), pp. 909–24, (2024).
15- Masoumeh Dastgir et al., "Synergistic radiosensitization of breast cancer using bismuth oxide nanoparticles coated with chitosan and β-cyclodextrin loaded with curcumin and 5-aminolevulinic acid." Cancer Nanotechnology, Vol. 17 (No. 1), p. 8, (2026).
16- Reza Malekzadeh et al., "Dose enhancement by gold, iron oxide, bismuth oxide, and platinum nanoparticles in radiotherapy: a comprehensive meta-analysis." Radiological Physics and Technology, Vol. 19 (No. 2), pp. 479–98, (2026).
17- Haniye Ahrabi et al., "Dual-Purpose 5-Aminolevulinic Acid-Loaded Iron Oxide Nanoparticles: Enhancing Radiotherapy and MRI in Glioblastoma." Journal of Drug Delivery Science and Technology, p. 107853, (2025).
18- Yasin Ayyami et al., "Characterization and application of targeted MnO2/CS@ ALA-MTX nano-radiosensitizers for boosting X-ray radiotherapy of brain tumors." Colloids and Surfaces A: Physicochemical and Engineering Aspects, Vol. 692p. 133975, (2024).
19- Maedeh Yektamanesh, Yasin Ayyami, Marjan Ghorbani, Masoumeh Dastgir, Reza Malekzadeh, and Tohid Mortezazadeh, "Characterization of multifunctional β-cyclodextrin-coated Bi2O3 nanoparticles conjugated with curcumin for CT imaging-guided synergetic chemo-radiotherapy in breast cancer." International Journal of Pharmaceutics, Vol. 659p. 124264, (2024).
20- Reza Malekzadeh et al., "Nanoarchitecture-based photothermal ablation of cancer: a systematic review." Environmental research, Vol. 236p. 116526, (2023).
21- Salah M Tawfik, Shavkatjon Azizov, Mohamed R Elmasry, Mirkomil Sharipov, and Yong-Ill Lee, "Recent advances in nanomicelles delivery systems." Nanomaterials, Vol. 11 (No. 1), p. 70, (2020).
22- Hoang M Trinh, Mary Joseph, Kishore Cholkar, Ranjana Mitra, and Ashim K Mitra, "Nanomicelles in diagnosis and drug delivery." in Emerging nanotechnologies for diagnostics, drug delivery and medical devices: Elsevier, (2017), pp. 45–58.
23- Haniye Ahrabi, Reza Malekzadeh, and Ghazale Geraily, "Application of 5-aminolevulinic acid in cancer radio-dynamic therapy: A systematic review." Radiation Physics and Chemistry, Vol. 236p. 112910, (2025).
24- Hua Guo, Huimin Wang, Hong Deng, Yiyi Zhang, Xue Yang, and Weiqi Zhang, "Facile preparation of toluidine blue-loaded DNA nanogels for anticancer photodynamic therapy." Frontiers in Bioengineering and Biotechnology, Vol. 11p. 1180448, (2023).
25- KM Gangotri, RC Meena, and Rajni Meena, "Use of micelles in photogalvanic cells for solar energy conversion and storage: cetyl trimethyl ammonium bromide-glucose-toluidine blue system." Journal of Photochemistry and Photobiology A: Chemistry, Vol. 123 (No. 1-3), pp. 93–97, (1999).
26- Reza Malekzadeh, Behnaz Babaye Abdollahi, Marjan Ghorbani, Jalil Pirayesh Islamian, and Tohid Mortezazadeh, "Trastuzumab conjugated PEG–Fe3O4@ Au nanoparticle as an MRI biocompatible nano-contrast agent." International Journal of Polymeric Materials and Polymeric Biomaterials, Vol. 72 (No. 10), pp. 759–70, (2023).
27- Ana-Maria Vlasceanu, Daniela L Baconi, Bianca Galateanu, Miriana Stan, and Cristian Balalau, "Comparative cytotoxicity study of nicotine and cotinine on MRC-5 cell line." Journal of Mind and Medical Sciences, Vol. 5 (No. 1), p. 17, (2018).
28- Alwan Al-Qushawi et al., "Preparation and characterization of three tilmicosin-loaded lipid nanoparticles: physicochemical properties and in-vitro antibacterial activities." Iranian Journal of Pharmaceutical Research: IJPR, Vol. 15 (No. 4), p. 663, (2016).
29- Xiaoxiao Gu, Haoyang Yuan, Cailing Li, Lixian Xu, Shuang Li, and Dinghua Yu, "Toluidine blue O photosensitizer combined with caffeic acid improves antibacterial performance by increasing the permeability of cell membrane." Colloids and Surfaces B: Biointerfaces, Vol. 233p. 113657, (2024).
30- Parinaz Mehnati, Reza Malekzadeh, and Sooteh Mohammad Yousefi, "New Bismuth composite shield for radiation protection of breast during coronary CT angiography." (2019).
31- Parinaz Mehnati, Reza Malekzadeh, Mohammad Yousefi Sooteh, and Soheila Refahi, "Assessment of the efficiency of new bismuth composite shields in radiation dose decline to breast during chest CT." The Egyptian Journal of Radiology and Nuclear Medicine, Vol. 49 (No. 4), pp. 1187–89, (2018).
32- Nikan Asadpour, Reza Malekzadeh, Saeed Rajabpour, Soheila Refahi, Parinaz Mehnati, and Ahmad Shanei, "Shielding performance of multi-metal nanoparticle composites for diagnostic radiology: an MCNPX and Geant4 study." Radiological Physics and Technology, Vol. 16 (No. 1), pp. 57–68, (2023).
33- Behnaz Babaye Abdollahi et al., "Main approaches to enhance radiosensitization in cancer cells by nanoparticles: A systematic review." Advanced pharmaceutical bulletin, Vol. 11 (No. 2), p. 212, (2020).
34- Hedieh Mansouri et al., "A biocompatible theranostic nanoplatform based on magnetic gadolinium-chelated polycyclodextrin: in vitro and in vivo studies." Carbohydrate polymers, Vol. 254p. 117262, (2021).
35- Junjie Liu, Jinjin Shi, Weimin Nie, Sijie Wang, Genhua Liu, and Kaiyong Cai, "Recent progress in the development of multifunctional nanoplatform for precise tumor phototherapy." Advanced healthcare materials, Vol. 10 (No. 1), p. 2001207, (2021).
36- Lama Misba, Hayder Abdulrahman, and Asad U Khan, "Photodynamic efficacy of toluidine blue O against mono species and dual species bacterial biofilm." Photodiagnosis and Photodynamic Therapy, Vol. 26pp. 383–88, (2019).
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| Keywords: Photodynamic therapy Toluidine blue Nanomicelles Lung cancer Selective Cytotoxicity. | ||
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How to Cite
1.
Khalili Najafabad B, Hoseini M, Rashnoodi A, Rouhi S, Najafisarvestani S, Seraj Z, Sadraei G. Toluidine Blue-Loaded Nanomicelles for Photodynamic Therapy of Lung Cancer: An In-vitro Study. Frontiers Biomed Technol. 2026;.
