The development and characterization of a human mesothelioma in vitro 3D model to investigate immunotoxin therapy
| Autor: | Xinran Xiang, Kunio Nagashima, Yen Phung, V. Courtney Broaddus, Mitchell Ho, Jingli Zhang, Mingqian Feng, David J. FitzGerald, Raffit Hassan |
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| Přispěvatelé: | Egles, Christophe |
| Rok vydání: | 2011 |
| Předmět: |
Mesothelioma
Pathology Cell Culture Techniques lcsh:Medicine Immunotoxin Monoclonal Tumor Cells Cultured Fluorescence microscope Tumor Microenvironment lcsh:Science Cancer Microscopy Multidisciplinary Cultured biology Chemistry Immunotoxins Antibodies Monoclonal Cell Biology/Extra-Cellular Matrix Cadherins Tumor Cells Antibody Research Article medicine.medical_specialty General Science & Technology Oncology/Oncology Agents Electron Antibodies Rare Diseases In vivo Spheroids Cellular medicine Humans Oncology/Lung Cancer Tumor microenvironment lcsh:R medicine.disease In vitro Microscopy Electron Cell Biology/Cell Adhesion Kinetics Orphan Drug Cell culture Cancer research biology.protein Cell Biology/Morphogenesis and Cell Biology lcsh:Q Cellular Spheroids |
| Zdroj: | PloS one, vol 6, iss 1 PLoS ONE, Vol 6, Iss 1, p e14640 (2011) PLoS ONE |
| Popis: | Background Tumor microenvironments present significant barriers to penetration by antibodies and immunoconjugates. Tumor microenvironments, however, are difficult to study in vitro. Cells cultured as monolayers exhibit less resistance to therapy than those grown in vivo and an alternative research model more representative of the in vivo tumor is more desirable. SS1P is an immunotoxin composed of the Fv portion of a mesothelin-specific antibody fused to a bacterial toxin that is presently undergoing clinical trials in mesothelioma. Methodology/Principal Findings Here, we examined how the tumor microenvironment affects the penetration and killing activity of SS1P in a new three-dimensional (3D) spheroid model cultured in vitro using the human mesothelioma cell line (NCI-H226) and two primary cell lines isolated from the ascites of malignant mesothelioma patients. Mesothelioma cells grown as monolayers or as spheroids expressed comparable levels of mesothelin; however, spheroids were at least 100 times less affected by SS1P. To understand this disparity in cytotoxicity, we made fluorescence-labeled SS1P molecules and used confocal microscopy to examine the time course of SS1P penetration within spheroids. The penetration was limited after 4 hours. Interestingly, we found a significant increase in the number of tight junctions in the core area of spheroids by electron microscopy. Expression of E-Cadherin, a protein involved in the assembly and sealing of tight junctions and highly expressed in malignant mesothelioma, was found significantly increased in spheroids as compared to monolayers. Moreover, we found that siRNA silencing and antibody inhibition targeting E-Cadherin could enhance SS1P immunotoxin therapy in vitro. Conclusion/Significance This work is one of the first to investigate immunotoxins in 3D tumor spheroids in vitro. This initial description of an in vitro tumor model may offer a simple and more representative model of in vivo tumors and will allow for further investigations of the microenvironmental effects on drug penetration and tumor cell killing. We believe that the methods developed here may apply to the studies of other tumor-targeting antibodies and immunoconjugates in vitro. |
| Databáze: | OpenAIRE |
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