Electrochemotherapy with Bleomycin Enhances Radiosensitivity of Uveal Melanomas: First In Vitro Results in 3D Cultures of Primary Uveal Melanoma Cell Lines
Abstract
:Simple Summary
Abstract
1. Introduction
2. Results
2.1. Characterization of 3D Tumor Spheroids of UM Cell Lines and UPM Cells
2.2. Impact of Combination Treatment on 3D Tumor Spheroid Growth
2.3. Proliferation and Necrosis of 3D Tumor Spheroids in Response to Combination Treatment
2.4. Viability of 3D Tumor Spheroids in Response to Combination Treatment
2.5. Cytotoxic Long-Term Effects on Tumor Spheroid Cells after Combination Treatment
3. Discussion
4. Materials and Methods
4.1. Culture of Uveal Melanoma Cells Lines
4.2. Cell Viability Assay
4.3. Tumor Spheroids
4.4. Determination of Spheroid Growth
4.5. Treatment of Spheroids
4.6. Immunohistochemistry of Tumor Spheroids
4.7. Spheroid Viability Assay
4.8. Spheroid Cell Survival Assay
4.9. Statistical Analysis
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Acknowledgments
Conflicts of Interest
References
- Kaliki, S.; Shields, C.L. Uveal melanoma: Relatively rare but deadly cancer. Eye 2017, 31, 241–257. [Google Scholar] [CrossRef] [Green Version]
- Shields, C.L.; Furuta, M.; Thangappan, A.; Nagori, S.; Mashayekhi, A.; Lally, D.R.; Kelly, C.C.; Rudich, D.S.; Nagori, A.V.; Wakade, O.A.; et al. Metastasis of uveal melanoma millimeter-by-millimeter in 8033 consecutive eyes. Arch. Ophthalmol. 2009, 127, 989–998. [Google Scholar] [CrossRef] [PubMed]
- Dogrusöz, M.; Jager, M.J.; Damato, B. Uveal Melanoma Treatment and Prognostication. Asia Pac. J. Ophthalmol. 2017, 6, 186–196. [Google Scholar] [CrossRef]
- Coupland, S.E.; Lake, S.L.; Zeschnigk, M.; Damato, B.E. Molecular pathology of uveal melanoma. Eye 2013, 27, 230–242. [Google Scholar] [CrossRef]
- Kaliki, S.; Shields, C.L.; Shields, J.A. Uveal melanoma: Estimating prognosis. Indian J. Ophthalmol. 2015, 63, 93–102. [Google Scholar] [CrossRef]
- Doherty, R.E.; Alfawaz, M.; Francis, J.; Lijka-Jones, B.; Sisley, K. Genetics of Uveal Melanoma. In Noncutaneous Melanoma; Scott, J.F., Gerstenblith, M.R., Eds.; Codon Publications: Brisbane (AU), Australia, 2018. [Google Scholar]
- Helgadottir, H.; Höiom, V. The genetics of uveal melanoma: Current insights. Appl. Clin. Genet. 2016, 9, 147–155. [Google Scholar] [CrossRef] [Green Version]
- Mallone, F.; Sacchetti, M.; Lambiase, A.; Moramarco, A. Molecular Insights and Emerging Strategies for Treatment of Metastatic Uveal Melanoma. Cancers 2020, 12, 2761. [Google Scholar] [CrossRef]
- Larkin, J.O.; Collins, C.G.; Aarons, S.; Tangney, M.; Whelan, M.; O’Reily, S.; Breathnach, O.; Soden, D.M.; O’Sullivan, G.C. Electrochemotherapy: Aspects of preclinical development and early clinical experience. Ann. Surg. 2007, 245, 469–479. [Google Scholar] [CrossRef] [PubMed]
- Edhemovic, I.; Gadzijev, E.M.; Brecelj, E.; Miklavcic, D.; Kos, B.; Zupanic, A.; Mali, B.; Jarm, T.; Pavliha, D.; Marcan, M.; et al. Electrochemotherapy: A new technological approach in treatment of metastases in the liver. Technol. Cancer Res. Treat. 2011, 10, 475–485. [Google Scholar] [CrossRef] [Green Version]
- Esmaeili, N.; Friebe, M. Electrochemotherapy: A Review of Current Status, Alternative IGP Approaches, and Future Perspectives. J. Healthc. Eng. 2019, 2019, 2784516. [Google Scholar] [CrossRef] [PubMed]
- Rezaee, Z.; Yadollahpour, A.; Rashidi, S.; Kunwar, P.S. Radiosensitizing Effect of Electrochemotherapy: A Systematic Review of Protocols and Efficiency. Curr. Drug Targets 2017, 18, 1893–1903. [Google Scholar] [CrossRef]
- Mir, L.M.; Gehl, J.; Sersa, G.; Collins, C.G.; Garbay, J.R.; Billard, V.; Geertsen, P.F.; Rudolf, Z.; O’Sullivan, G.C.; Marty, M. Standard operating procedures of the electrochemotherapy: Instructions for the use of bleomycin or cisplatin administered either systemically or locally and electric pulses delivered by Cliniporator by means of invasive or non-invasive electrodes. Eur. J. Cancer Suppl. 2006, 4, 14–25. [Google Scholar] [CrossRef]
- Mir, L.M. Therapeutic perspectives of in vivo cell electropermeabilization. Bioelectrochemistry 2001, 53, 1–10. [Google Scholar] [CrossRef]
- Mir, L.M.; Orlowski, S. The basis of electrochemotherapy. Methods Mol. Med. 2000, 37, 99–117. [Google Scholar] [CrossRef] [PubMed]
- Fiorentzis, M.; Kalirai, H.; Katopodis, P.; Seitz, B.; Viestenz, A.; Coupland, S.E. Electrochemotherapy with bleomycin and cisplatin enhances cytotoxicity in primary and metastatic uveal melanoma cell lines in vitro. Neoplasma 2018, 65, 210–215. [Google Scholar] [CrossRef] [Green Version]
- Fiorentzis, M.; Viestenz, A.; Siebolts, U.; Seitz, B.; Coupland, S.E.; Heinzelmann, J. The Potential Use of Electrochemotherapy in the Treatment of Uveal Melanoma: In Vitro Results in 3D Tumor Cultures and In Vivo Results in a Chick Embryo Model. Cancers 2019, 11, 1344. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Kranjc, S.; Cemazar, M.; Grosel, A.; Scancar, J.; Sersa, G. Electroporation of LPB sarcoma cells in vitro and tumors in vivo increases the radiosensitizing effect of cisplatin. Anticancer Res. 2003, 23, 275–281. [Google Scholar]
- Kranjc, S.; Cemazar, M.; Grosel, A.; Sentjurc, M.; Sersa, G. Radiosensitising effect of electrochemotherapy with bleomycin in LPB sarcoma cells and tumors in mice. BMC Cancer 2005, 5, 115. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Kranjc, S.; Tevz, G.; Kamensek, U.; Vidic, S.; Cemazar, M.; Sersa, G. Radiosensitizing effect of electrochemotherapy in a fractionated radiation regimen in radiosensitive murine sarcoma and radioresistant adenocarcinoma tumor model. Radiat. Res. 2009, 172, 677–685. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Jiang, G.L.; Ang, K.K.; Thames, H.D.; Wong, C.S.; Wendt, C.D. Response of plateau-phase C3H 10T1/2 cells to radiation and concurrent administration of bleomycin. Radiat. Res. 1989, 120, 306–312. [Google Scholar] [CrossRef]
- Teicher, B.A.; Herman, T.S.; Holden, S.A. Combined modality therapy with bleomycin, hyperthermia, and radiation. Cancer Res. 1988, 48, 6291–6297. [Google Scholar] [PubMed]
- Skarlatos, I.; Kyrgias, G.; Mosa, E.; Provatopoulou, X.; Spyrou, M.; Theodorou, K.; Lepouras, A.; Gounaris, A.; Koukourakis, M. Electrochemotherapy in cancer patients: First clinical trial in Greece. In Vivo 2011, 25, 265–274. [Google Scholar] [PubMed]
- De Waard-Siebinga, I.; Blom, D.J.; Griffioen, M.; Schrier, P.I.; Hoogendoorn, E.; Beverstock, G.; Danen, E.H.; Jager, M.J. Establishment and characterization of an uveal-melanoma cell line. Int. J. Cancer 1995, 62, 155–161. [Google Scholar] [CrossRef]
- Jager, M.J.; Magner, J.A.; Ksander, B.R.; Dubovy, S.R. Uveal Melanoma Cell Lines: Where do they come from? (An American Ophthalmological Society Thesis). Trans. Am. Ophthalmol. Soc. 2016, 114, T5. [Google Scholar]
- Griewank, K.G.; Yu, X.; Khalili, J.; Sozen, M.M.; Stempke-Hale, K.; Bernatchez, C.; Wardell, S.; Bastian, B.C.; Woodman, S.E. Genetic and molecular characterization of uveal melanoma cell lines. Pigment Cell Melanoma Res. 2012, 25, 182–187. [Google Scholar] [CrossRef] [Green Version]
- Amirouchene-Angelozzi, N.; Nemati, F.; Gentien, D.; Nicolas, A.; Dumont, A.; Carita, G.; Camonis, J.; Desjardins, L.; Cassoux, N.; Piperno-Neumann, S.; et al. Establishment of novel cell lines recapitulating the genetic landscape of uveal melanoma and preclinical validation of mTOR as a therapeutic target. Mol. Oncol. 2014, 8, 1508–1520. [Google Scholar] [CrossRef] [PubMed]
- Van den Aardweg, G.J.; Kiliç, E.; de Klein, A.; Luyten, G.P. Dose fractionation effects in primary and metastatic human uveal melanoma cell lines. Invest. Ophthalmol. Vis. Sci. 2003, 44, 4660–4664. [Google Scholar] [CrossRef]
- Nareyeck, G.; Zeschnigk, M.; Bornfeld, N.; Anastassiou, G. Novel cell lines derived by long-term culture of primary uveal melanomas. Ophthalmologica 2009, 223, 196–201. [Google Scholar] [CrossRef]
- Djeridane, M.; Oudard, S.; Escoffre-Barbe, M.; Lacotte-Thierry, L.; Desablens, B.; Briére, J.; Dib, M.; Cassasus, P.; Ghandour, C.; Lamy, T.; et al. Treatment of patients with advanced or bulky Hodgkin disease with a 12-week doxorubicin, bleomycin, vinblastine, and dacarbazine-like chemotherapy regimen followed by extended-field, full-dose radiotherapy: Long-term results of the Groupe Ouest et Est des Leucémies et Autres Maladies de Sang H90-A/B Multicenter Randomized Trial. Cancer 2002, 95, 2169–2179. [Google Scholar] [CrossRef]
- Lelieveld, P.; Scoles, M.A.; Brown, J.M.; Kallman, R.F. The effect of treatment in fractionated schedules with the combination of X-irradiation and six cytotoxic drugs on the RIF-1 tumor and normal mouse skin. Int. J. Radiat. Oncol. Biol. Phys. 1985, 11, 111–121. [Google Scholar] [CrossRef]
- Molin, J.; Søgaard, P.E.; Overgaard, J. Experimental studies on the radiation-modifying effect of bleomycin in malignant and normal mouse tissue in vivo. Cancer Treat. Rep. 1981, 65, 583–589. [Google Scholar]
- Smid, L.; Budihna, M.; Zakotnik, B.; Soba, E.; Strojan, P.; Fajdiga, I.; Zargi, M.; Oblak, I.; Dremelj, M.; LeSnicar, H. Postoperative concomitant irradiation and chemotherapy with mitomycin C and bleomycin for advanced head-and-neck carcinoma. Int. J. Radiat. Oncol. Biol. Phys. 2003, 56, 1055–1062. [Google Scholar] [CrossRef]
- Fu, K.K.; Phillips, T.L.; Silverberg, I.J.; Jacobs, C.; Goffinet, D.R.; Chun, C.; Friedman, M.A.; Kohler, M.; McWhirter, K.; Carter, S.K. Combined radiotherapy and chemotherapy with bleomycin and methotrexate for advanced inoperable head and neck cancer: Update of a Northern California Oncology Group randomized trial. J. Clin. Oncol. 1987, 5, 1410–1418. [Google Scholar] [CrossRef]
- Suntharalingam, M.; Haas, M.L.; Van Echo, D.A.; Haddad, R.; Jacobs, M.C.; Levy, S.; Gray, W.C.; Ord, R.A.; Conley, B.A. Predictors of response and survival after concurrent chemotherapy and radiation for locally advanced squamous cell carcinomas of the head and neck. Cancer 2001, 91, 548–554. [Google Scholar] [CrossRef]
- Chen, J.; Ghorai, M.K.; Kenney, G.; Stubbe, J. Mechanistic studies on bleomycin-mediated DNA damage: Multiple binding modes can result in double-stranded DNA cleavage. Nucleic Acids Res. 2008, 36, 3781–3790. [Google Scholar] [CrossRef] [Green Version]
- Povirk, L.F. DNA damage and mutagenesis by radiomimetic DNA-cleaving agents: Bleomycin, neocarzinostatin and other enediynes. Mutat. Res. 1996, 355, 71–89. [Google Scholar] [CrossRef]
- Sersa, G.; Cemazar, M.; Rudolf, Z.; Fras, A.P. Adenocarcinoma skin metastases treated by electrochemotherapy with cisplatin combined with radiation. Radiol. Oncol. 1999, 33, 291–296. [Google Scholar]
- Leith, J.T.; Lee, E.S.; Leite, D.V.; Glicksman, A.S. Enhanced X ray sensitivity of human colon tumor cells by combination of N-methylformamide with chemotherapeutic agents. Int. J. Radiat. Oncol. Biol. Phys. 1986, 12, 1423–1427. [Google Scholar] [CrossRef]
- Begg, A.C.; Deurloo, M.J.; Kop, W.; Bartelink, H. Improvement of combined modality therapy with cisplatin and radiation using intratumoral drug administration in murine tumors. Radiother. Oncol. 1994, 31, 129–137. [Google Scholar] [CrossRef]
Cell Line | Genetics | Morphology/Doubling Time | References |
---|---|---|---|
UM92.1 | GNAQ Q209L, Disomy-3, WT BAP1, EIF1AX | Epithelioid/38–58 h | [24,25,26,27] |
Mel270 | GNAQ Q209P, Disomy-3, WT BAP1 | Spindle/43 h | [25,26,28] |
UPMD2 | GNA11 Q209L, Isodisomy-3, WT BAP1 | Epithelioid/150 h | [16,29] |
UPMM3 | GNAQ Q209P, Monosomy-3, Mutant BAP1 | Spindle and epithelioid/100 h | [29] |
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. |
© 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Fiorentzis, M.; Sokolenko, E.A.; Bechrakis, N.E.; Ting, S.; Schmid, K.W.; Sak, A.; Stuschke, M.; Seitz, B.; Berchner-Pfannschmidt, U. Electrochemotherapy with Bleomycin Enhances Radiosensitivity of Uveal Melanomas: First In Vitro Results in 3D Cultures of Primary Uveal Melanoma Cell Lines. Cancers 2021, 13, 3086. https://doi.org/10.3390/cancers13123086
Fiorentzis M, Sokolenko EA, Bechrakis NE, Ting S, Schmid KW, Sak A, Stuschke M, Seitz B, Berchner-Pfannschmidt U. Electrochemotherapy with Bleomycin Enhances Radiosensitivity of Uveal Melanomas: First In Vitro Results in 3D Cultures of Primary Uveal Melanoma Cell Lines. Cancers. 2021; 13(12):3086. https://doi.org/10.3390/cancers13123086
Chicago/Turabian StyleFiorentzis, Miltiadis, Ekaterina A. Sokolenko, Nikolaos E. Bechrakis, Saskia Ting, Kurt W. Schmid, Ali Sak, Martin Stuschke, Berthold Seitz, and Utta Berchner-Pfannschmidt. 2021. "Electrochemotherapy with Bleomycin Enhances Radiosensitivity of Uveal Melanomas: First In Vitro Results in 3D Cultures of Primary Uveal Melanoma Cell Lines" Cancers 13, no. 12: 3086. https://doi.org/10.3390/cancers13123086
APA StyleFiorentzis, M., Sokolenko, E. A., Bechrakis, N. E., Ting, S., Schmid, K. W., Sak, A., Stuschke, M., Seitz, B., & Berchner-Pfannschmidt, U. (2021). Electrochemotherapy with Bleomycin Enhances Radiosensitivity of Uveal Melanomas: First In Vitro Results in 3D Cultures of Primary Uveal Melanoma Cell Lines. Cancers, 13(12), 3086. https://doi.org/10.3390/cancers13123086