Malignant gliomas have a dismal prognosis and significant efforts are being made to develop more effective treatments. Sonodynamic therapy (SDT) is an emerging modality for cancer treatment which combines ultrasound with sonosensitizers to produce a localized cytotoxic effect. The aim of this study is to demonstrate the efficacy of SDT with fluorescein (FL) and low-intensity focused ultrasound in inhibiting the growth of ectopic gliomas implanted in the rat’s subcutaneous tissue.
In vivo cytotoxicity of FL-SDT was evaluated in C6 rat glioma cells which were inoculated subcutaneously. Tumor specific extracellular FL extravasation and accumulation was assessed with IVIS imaging in rats receiving systemic FL. Effects of FL-SDT with focused low-intensity ultrasound on tumor growth, and histological features of the rat’s tumors were investigated. Treatment related apoptosis and necrosis were analyzed using hematoxylin & eosin, and apoptosis-specific staining.
IVIS imaging revealed a high degree of FL accumulation within the tumor, with a nearly threefold increase in tumoral epifluorescence signal over background. SDT significantly inhibited outgrowth of ectopic C6 gliomas across all three FUS exposure conditions. TUNEL and active caspase-3 staining did not reveal conclusive trends across control and SDT condition for apoptosis.
Our results suggest that SDT with FL and low-intensity FUS is effective in inhibiting the growth of ectopic malignant gliomas in rats. The selective FL extravasation and accumulation in the tumor areas where the blood–brain barrier is damaged suggests the tumor-specificity of the treatment. The possibility to use this treatment in intracranial models and in human gliomas will have to be explored in further studies.
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Reactive oxygen species
TdT-mediated dUTP nick end labelling
Magnetic resonance imaging
High intensity focused ultrasound
Suehiro S, Ohnishi T, Yamashita D, Kohno S, Inoue A, Nishikawa M, Ohue S, Tanaka J, Kunieda T (2018) Enhancement of antitumor activity by using 5-ALA-mediated sonodynamic therapy to induce apoptosis in malignant gliomas: significance of high-intensity focused ultrasound on 5-ALA-SDT in a mouse glioma model. J Neurosurg 129(6):1416–1428. https://doi.org/10.3171/2017.6.JNS162398
McDannold N, Clement GT, Black P, Jolesz F, Hynynen K (2010) Transcranial magnetic resonance imaging- guided focused ultrasound surgery of brain tumors: initial findings in 3 patients. Neurosurgery 66(2):323–332. https://doi.org/10.1227/01.NEU.0000360379.95800.2Fdiscussion 332
McHale AP, Callan JF, Nomikou N, Fowley C, Callan B (2016) Sonodynamic therapy: concept, mechanism and application to cancer treatment. Therapeutic ultrasound. Springer, Berlin, pp 429–450
Rosenthal I, Sostaric JZ, Riesz P (2004) Sonodynamic therapy–a review of the synergistic effects of drugs and ultrasound. Ultrason Sonochem 11(6):349–363. https://doi.org/10.1016/j.ultsonch.2004.03.004
Costley D, Mc Ewan C, Fowley C, McHale AP, Atchison J, Nomikou N, Callan JF (2015) Treating cancer with sonodynamic therapy: a review. Int J Hyperthermia 31(2):107–117. https://doi.org/10.3109/02656736.2014.992484
Endo S, Kudo N, Yamaguchi S, Sumiyoshi K, Motegi H, Kobayashi H, Terasaka S, Houkin K (2015) Porphyrin derivatives-mediated sonodynamic therapy for malignant gliomas in vitro. Ultrasound Med Biol 41(9):2458–2465. https://doi.org/10.1016/j.ultrasmedbio.2015.05.007
Wang S, Hu Z, Wang X, Gu C, Gao Z, Cao W, Zheng J (2014) 5-Aminolevulinic acid-mediated sonodynamic therapy reverses macrophage and dendritic cell passivity in murine melanoma xenografts. Ultrasound Med Biol 40(9):2125–2133. https://doi.org/10.1016/j.ultrasmedbio.2014.05.007
Gao Z, Zheng J, Yang B, Wang Z, Fan H, Lv Y, Li H, Jia L, Cao W (2013) Sonodynamic therapy inhibits angiogenesis and tumor growth in a xenograft mouse model. Cancer Lett 335(1):93–99. https://doi.org/10.1016/j.canlet.2013.02.006
Kujawska T, Secomski W, Bilmin K, Nowicki A, Grieb P (2014) Impact of thermal effects induced by ultrasound on viability of rat C6 glioma cells. Ultrasonics 54(5):1366–1372. https://doi.org/10.1016/j.ultras.2014.02.002
Rengeng L, Qianyu Z, Yuehong L, Zhongzhong P, Libo L (2017) Sonodynamic therapy, a treatment developing from photodynamic therapy. Photodiagnosis Photodyn Ther 19:159–166. https://doi.org/10.1016/j.pdpdt.2017.06.003
Hersh DS, Kim AJ, Winkles JA, Eisenberg HM, Woodworth GF, Frenkel V (2016) Emerging applications of therapeutic ultrasound in neuro-oncology: moving beyond tumor ablation. Neurosurgery 79(5):643–654. https://doi.org/10.1227/NEU.0000000000001399
Yoshida M, Kobayashi H, Terasaka S, Endo S, Yamaguchi S, Motegi H, Itay R, Suzuki S, Brokman O, Shapira Y, Moriyama K, Kawase Y, Akahane T, Kato Y, Kamada H, Houkin K (2019) Sonodynamic therapy for malignant Glioma Using 220-kHz Transcranial Magnetic Resonance Imaging-Guided focused ultrasound and 5-aminolevulinic acid. Ultrasound Med Biol 45(2):526–538. https://doi.org/10.1016/j.ultrasmedbio.2018.10.016
Wang X, Jia Y, Wang P, Liu Q, Zheng H (2017) Current status and future perspectives of sonodynamic therapy in glioma treatment. Ultrason Sonochem 37:592–599. https://doi.org/10.1016/j.ultsonch.2017.02.020
Tserkovsky DA, Alexandrova EN, Chalau VN, Istomin YP (2012) Effects of combined sonodynamic and photodynamic therapies with photolon on a glioma C6 tumor model. Exp Oncol 34(4):332–335
Sun Y, Wang H, Wang P, Zhang K, Geng X, Liu Q, Wang X (2019) Tumor targeting DVDMS-nanoliposomes for an enhanced sonodynamic therapy of gliomas. Biomater Sci 7(3):985–994. https://doi.org/10.1039/c8bm01187g
Pi Z, Huang Y, Shen Y, Zeng X, Hu Y, Chen T, Li C, Yu H, Chen S, Chen X (2019) Sonodynamic therapy on intracranial glioblastoma xenografts using sinoporphyrin sodium delivered by ultrasound with microbubbles. Ann Biomed Eng 47(2):549–562. https://doi.org/10.1007/s10439-018-02141-9
Ohmura T, Fukushima T, Shibaguchi H, Yoshizawa S, Inoue T, Kuroki M, Sasaki K, Umemura S (2011) Sonodynamic therapy with 5-aminolevulinic acid and focused ultrasound for deep-seated intracranial glioma in rat. Anticancer Res 31(7):2527–2533
Nonaka M, Yamamoto M, Yoshino S, Umemura S, Sasaki K, Fukushima T (2009) Sonodynamic therapy consisting of focused ultrasound and a photosensitizer causes a selective antitumor effect in a rat intracranial glioma model. Anticancer Res 29(3):943–950
Li E, Sun Y, Lv G, Li Y, Zhang Z, Hu Z, Cao W (2019) Sinoporphyrin sodium based sonodynamic therapy induces anti-tumor effects in hepatocellular carcinoma and activates p53/caspase 3 axis. Int J Biochem Cell Biol. https://doi.org/10.1016/j.biocel.2019.01.009
Ju D, Yamaguchi F, Zhan G, Higuchi T, Asakura T, Morita A, Orimo H, Hu S (2016) Hyperthermotherapy enhances antitumor effect of 5-aminolevulinic acid-mediated sonodynamic therapy with activation of caspase-dependent apoptotic pathway in human glioma. Tumour Biol 37(8):10415–10426. https://doi.org/10.1007/s13277-016-4931-3
Jeong EJ, Seo SJ, Ahn YJ, Choi KH, Kim KH, Kim JK (2012) Sonodynamically induced antitumor effects of 5-aminolevulinic acid and fractionated ultrasound irradiation in an orthotopic rat glioma model. Ultrasound Med Biol 38(12):2143–2150. https://doi.org/10.1016/j.ultrasmedbio.2012.07.026
Dai S, Xu C, Tian Y, Cheng W, Li B (2014) In vitro stimulation of calcium overload and apoptosis by sonodynamic therapy combined with hematoporphyrin monomethyl ether in C6 glioma cells. Oncol Lett 8(4):1675–1681. https://doi.org/10.3892/ol.2014.2419
Acerbi F, Broggi M, Schebesch K-M, Höhne J, Cavallo C, De Laurentis C, Eoli M, Anghileri E, Servida M, Boffano CJCCR (2018) Fluorescein-guided surgery for resection of high-grade gliomas: a multicentric prospective phase II study (FLUOGLIO). World Neurosurg 24(1):52–61
Folaron M, Strawbridge R, Samkoe KS, Filan C, Roberts DW, Davis SC (2018) Elucidating the kinetics of sodium fluorescein for fluorescence-guided surgery of glioma. J Neurosurg. https://doi.org/10.3171/2018.4.JNS172644
Zou M, Zhang L, Wang J, Wang Q, Gao J, Fan P (2013) Investigation on interaction and sonodynamic damage of fluorescein derivants to bovine serum albumin (BSA) under ultrasonic irradiation. Spectrochim Acta A 110:364–376. https://doi.org/10.1016/j.saa.2013.03.073
Acerbi F, Broggi M, Schebesch KM, Hohne J, Cavallo C, De Laurentis C, Eoli M, Anghileri E, Servida M, Boffano C, Pollo B, Schiariti M, Visintini S, Montomoli C, Bosio L, La Corte E, Broggi G, Brawanski A, Ferroli P (2018) Fluorescein-guided surgery for resection of high-grade gliomas: a multicentric prospective phase II study (FLUOGLIO). Clin Cancer Res 24(1):52–61. https://doi.org/10.1158/1078-0432.CCR-17-1184
Barth RF, Kaur B (2009) Rat brain tumor models in experimental neuro-oncology: the C6, 9L, T9, RG2, F98, BT4C, RT-2 and CNS-1 gliomas. J Neurooncol 94(3):299–312. https://doi.org/10.1007/s11060-009-9875-7
Gieryng A, Pszczolkowska D, Bocian K, Dabrowski M, Rajan WD, Kloss M, Mieczkowski J, Kaminska B (2017) Immune microenvironment of experimental rat C6 gliomas resembles human glioblastomas. Sci Rep 7(1):17556. https://doi.org/10.1038/s41598-017-17752-w
Jacobs VL, Valdes PA, Hickey WF, De Leo JA (2011) Current review of in vivo GBM rodent models: emphasis on the CNS-1 tumour model. ASN Neuro 3(3):e00063. https://doi.org/10.1042/AN20110014
Song D, Yue W, Li Z, Li J, Zhao J, Zhang N (2014) Study of the mechanism of sonodynamic therapy in a rat glioma model. Onco Targets Ther 7:1801–1810. https://doi.org/10.2147/OTT.S52426
Liu H, Zhou M, Sheng Z, Chen Y, Yeh CK, Chen W, Liu J, Liu X, Yan F, Zheng H (2018) Theranostic nanosensitizers for highly efficient MR/fluorescence imaging-guided sonodynamic therapy of gliomas. J Cell Mol Med 22(11):5394–5405. https://doi.org/10.1111/jcmm.13811
Yamaguchi F, Asakura T, Takahashi H, Kitamura T, Teramoto A (2013) Low frequency ultrasonication induced antitumor effect in 5-aminolevulinic acid treated malignant glioma. J Cancer Ther 4(1):170
Umemura S, Yumita N, Nishigaki R, Umemura K (1990) Mechanism of cell damage by ultrasound in combination with hematoporphyrin. Jpn J Cancer Res 81(9):962–966. https://doi.org/10.1111/j.1349-7006.1990.tb02674.x
Yumita N, Nishigaki R, Umemura K, Umemura S (1989) Increase in the generation of superoxide radicals and in the inhibitory effect on Yoshida sarcoma of anthracycline antitumor agents by ultrasound. Nihon Gan Chiryo Gakkai Shi 24(1):63–68
Wu SK, Santos MA, Marcus SL, Hynynen K (2019) MR-guided focused ultrasound facilitates sonodynamic therapy with 5-aminolevulinic acid in a rat Glioma model. Sci Rep 9(1):10465. https://doi.org/10.1038/s41598-019-46832-2
Cavallo C, De Laurentis C, Vetrano IG, Falco J, Broggi M, Schiariti M, Ferroli P, Acerbi F (2018) The utilization of fluorescein in brain tumor surgery: a systematic review. J Neurosurg Sci 62(6):690–703. https://doi.org/10.23736/S0390-5616.18.04480-6
Han J, Jose J, Mei E, Burgess K (2007) Chemiluminescent energy-transfer cassettes based on fluorescein and nile red. Angew Chem Int Ed Engl 46(10):1684–1687. https://doi.org/10.1002/anie.200603307
Hynynen K, Vykhodtseva NI, Chung AH, Sorrentino V, Colucci V, Jolesz FA (1997) Thermal effects of focused ultrasound on the brain: determination with MR imaging. Radiology 204(1):247–253. https://doi.org/10.1148/radiology.204.1.9205255
Ram Z, Cohen ZR, Harnof S, Tal S, Faibel M, Nass D, Maier SE, Hadani M, Mardor Y (2006) Magnetic resonance imaging-guided, high-intensity focused ultrasound for brain tumor therapy. Neurosurgery 59(5):949–955. https://doi.org/10.1227/01.NEU.0000254439.02736.D8discussion 955–946
Coluccia D, Fandino J, Schwyzer L, O’Gorman R, Remonda L, Anon J, Martin E, Werner B (2014) First noninvasive thermal ablation of a brain tumor with MR-guided focusedultrasound. J Ther Ultrasound 2(1):17
Eames MD, Farnum M, Khaled M, Elias WJ, Hananel A, Snell JW, Kassell NF, Aubry JF (2015) Head phantoms for transcranial focused ultrasound. Med Phys 42(4):1518–1527. https://doi.org/10.1118/1.4907959
Gao HJ, Zhang WM, Wang XH, Zheng RN (2010) Adriamycin enhances the sonodynamic effect of chlorin e6 against the proliferation of human breast cancer MDA-MB-231 cells in vitro. Nan Fang Yi Ke Da Xue Xue Bao 30(10):2291–2294
McEwan C, Kamila S, Owen J, Nesbitt H, Callan B, Borden M, Nomikou N, Hamoudi RA, Taylor MA, Stride E, McHale AP, Callan JF (2016) Combined sonodynamic and antimetabolite therapy for the improved treatment of pancreatic cancer using oxygen loaded microbubbles as a delivery vehicle. Biomaterials 80:20–32. https://doi.org/10.1016/j.biomaterials.2015.11.033
Miyoshi N, Kundu SK, Tuziuti T, Yasui K, Shimada I, Ito Y (2016) Combination of sonodynamic and photodynamic therapy against cancer would be effective through using a regulated size of nanoparticles. Nanosci Nanoeng 4(1):1–11. https://doi.org/10.13189/nn.2016.040101
Kamath AA, Friedman DD, Akbari SHA, Kim AH, Tao Y, Luo J, Leuthardt EC (2019) Glioblastoma treated with magnetic resonance imaging-guided laser interstitial thermal therapy: safety, efficacy, and outcomes. Neurosurgery 84(4):836–843. https://doi.org/10.1093/neuros/nyy375
Sheehan J (2012) Stereotactic radiosurgery for glioblastoma–time to revisit this approach. World Neurosurg 78(6):592–593. https://doi.org/10.1016/j.wneu.2012.05.023
Falco J, Cavallo C, Vetrano IG, de Laurentis C, Siozos L, Schiariti M, Broggi M, Ferroli P, Acerbi F (2019) Fluorescein application in cranial and spinal tumors enhancing at preoperative MRI and operated with a dedicated filter on the surgical microscope: preliminary results in 279 patients enrolled in the FLUOCERTUM prospective study. Front Surg 6:49. https://doi.org/10.3389/fsurg.2019.00049
This work was conducted at University of Virginia through a grant from the Focused Ultrasound Foundation.
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Prada, F., Sheybani, N., Franzini, A. et al. Fluorescein-mediated sonodynamic therapy in a rat glioma model. J Neurooncol 148, 445–454 (2020). https://doi.org/10.1007/s11060-020-03536-2
- Sonodynamic therapy
- Focused ultrasound
- Brain tumor