 |
3R-Project 115-09
Engineering of a human brain tumour model to replace animal experimentation
Olivier Preynat-Seauve1 and Thierry Virolle2
1University of Geneva, CH-1211 Geneva, Switzerland
2INSERM/University of Nice, F-06107 Nice, France
olivier.preynat-seauve@unige.ch, virolle@unice.fr
Keywords: human; brain; stem cells; tumour; tumourigenesis; reduction; replacement
Duration: 3 years End of the Project: 2012
Background and Aim
Gliomas account for more than 70% of all brain tumours, and of these, glioblastoma is the most frequent and malignant histological type. Fewer than 3% of glioblastoma patients are still alive at 5 years after diagnosis. Recently, there has been important progress in our understanding of the molecular pathogenesis of malignant gliomas, and especially the importance of cancer stem cells. Models are central to allow the transition from scientific concepts to the understanding of the reality of a tumour in a person. They may be used for therapeutic screens, in preclinical trials, or to study the basic biology of brain tumours. Injection of glioblastoma cells either subcutaneously or directly into the animal brain, usually using immunodeficient strains such as Nude or SCID, or transgenic mouse models spontaneously developing brain tumours, allow in-vivo modelling with similar histopathology, etiology, and biology. These mouse models are widely used but, unfortunately, these available model systems are very aggressive for animals (infections, painful, severe motor and behavioural symptoms). In addition, such animal models do not reflect the real interaction between tumoural cells and the host that occurs in the human tissue. Indeed, human tumour cells are generally injected into an animal brain. Thus, new models that (i) replace animal experiments and (ii) provide a more relevant tumour/host interaction are needed.
Using the technology of embryonic stem cells (ESC), we propose to create an integrated and fully in-vitro model of a human glioblastoma developing within a human brain tissue. To achieve this goal, we will combine two technologies already available in our laboratory: (i) ENT technology: We recently developed in our laboratory the technology of Engineered Neural Tissues (ENTs). ENTs are reconstituted brain-like tissue pieces derived in-vitro from human embryonic stem cells (Preynat-Seauve, O. et al., 2008.). (ii) EGT technology: in parallel, using the same technology as ENTs, we have developed EGTs (Engineered Glial Tumors). EGTs are a model of reconstituted glioblastomas in three dimension derived in-vitro from patient brain tumour samples (Preynat-Seauve, O. et al., 2008.). We propose to combine ENT and EGT technologies in order to develop a fully in-vitro model of a human glioblastoma growing within a human brain-like tissue. This method will provide a powerful human model for the study of glioblastomas replacing animal models.
Method and Results
in progress (present status)
The basis of this project is to combine the two technologies in order to associate a human brain-like tissue with a human glioblastoma-like tumor. The first step will be the production of ENTs. The second step will be the physical association between ENTs and glioblastoma cells. In all cases, gliomaspheres are labelled by reporter protein to distinguish them from the host tissues. Dr Thierry Virolle and Dr Laurent Turchi (INSERM U898, University of Nice, France) ) in collaboration with Dr Hervé Chneiweiss (INSERM U894, Paris, France) have produced recently gliomaspheres expressing the red fluorescent Tomato Fluorescent Protein (TFP). Several gliomaspheres derived from different patient samples will be processed and tested for their ability to develop within ENTs: The ENT/EGT association will be characterized by histology and molecular biology for its resemblance to a glioblastoma developing in patients and its tumorigenicity.
Conclusions and Relevance for 3R
The development of a fully in-vitro human brain tumour cells within a reconstituted human brain-like tissue is expected to reduce the number of experimentations performed on animals. It will also reduce the impact on animals used in animal experimentation since the existing brain tumour animal models are very invasive for the animals, the latter suffering from severe deficits.
References
Olivier Preynat-Seauve 1*, David M. Suter 1, Diderik Tirefort 1, Laurant Turchi 2, Thierry Virolle 2, Herve Chneiweiss 3, Michelangelo Foti 4, Johannes- Alexander Lobrinus 5, Luc Stoppini 6, Ania Feki 7, Michel Dubois-Dauphin 1, Karl Heinz Krause 1* (2009) Development of Human Nervous Tissue upon Differentiation of Embryonic Stem Cells in Three Dimensional Culture. Stem Cells 27, 509-520.
DOI: 10.1634/stemcells.2008-0600
*Correspondence to Olivier Preynat-Seauve and Heinz Krause
1 Department of Pathology and Immunology, Faculty of Medicine, University of Geneva and Department of Genetic and Laboratory Medicine, Geneva Hospital, Switzerland
INSERM U898, Faculty of medicine, Nice, France
3 INSERM U894, Faculty of medicine Paris Descartes, Paris, France
4 Department of Cell Physiology and Metabolism, Faculty of Medicine, University of Geneva, Switzerland
5 Neuropathology Unit, Department of Genetic and Laboratory Medicine, Geneva Hospital, Switzerland
6 Geneva Institute of Technology, HES-SO//University of Applied Sciences Western Switzerland
7 Stem cell research Laboratory, Department of Gynecology and Obstetrics, Geneva Hospital, Switzerland
Figures
Figure 1: Macroscopic view of a human ENT derived in vitro from embryonic stem cells.
Figure 2: Coloration with fluorescent probes of an engineered neural tissue section. Blue staining indicate cell nuclei, showing the tissular organization. Red staining indicates germinal layers of neural stem cells generating the mature neuronal tissue (in green).
Coloration with fluorescent probes of an engineered neural tissue section. Blue staining indicate cell nuclei, showing the tissular organization. Red staining indicates germinal layers of neural stem cells generating the mature neuronal tissue (in green)
| Dernières modifications: 07.03.2010 |  |