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Suppression of human glioma growth by adenovirus-mediated Rb gene transfer
J Fueyo1, C Gomez-Manzano, W K Yung
1Department of Neuro-Oncology, The University of Texas M.D. Anderson Cancer Center, Houston 77030, USA.
Objective:
This study was conducted to obtain evidence that restoration of the retinoblastoma protein function may have therapeutic application for gliomas.
Background:
The development of glioblastoma multiforme involves progressive inactivation of several tumor suppressor genes. Abnormalities of the retinoblastoma tumor suppressor gene are found in the majority of cancers, including at least 30% of malignant gliomas. No final evidence has been produced about the role of Rb in suppressing glioma growth.
Methods:
To address this question, the Ad5CMV-Rb adenovirus carrying a 3.2-kb cDNA of the Rb gene was constructed. Expression of the exogenous protein was assessed by immunoblot and immunohistochemistry analyses. Growth curve assays were used to evaluate the effect of the Rb protein on glioma cell growth. Flow-cytometry analyses were used to analyze the phenotype of the cell cycle after the transfer of Rb. Human glioma xenografts implanted subcutaneously in nude mice were used for the tumorigenicity assay.
Results:
After the transfer of Rb, 80% of the treated cells expressed high levels of the retinoblastoma protein for at least 7 days. Within 5 days of treatment, the cells lost the neoplastic morphology and showed marked growth suppression. The majority of the Rb-expressing cells were arrested in the G1 phase of the cell cycle. In addition, the restoration of the retinoblastoma activity rendered the human glioma cells unable to form tumors in nude mice.
Conclusions:
These findings provide direct evidence that inactivation of the retinoblastoma protein is a critical event in gliomas, and suggest that the restoration of wild-type retinoblastoma activity in these tumors may have therapeutic utility.
Insights
Restoring retinoblastoma (Rb) protein function suppressed glioma growth and tumor formation in mice. This suggests Rb restoration may be a promising therapeutic strategy for gliomas.
Area of Science:
- Oncology
- Molecular Biology
- Gene Therapy
Background:
- Glioblastoma multiforme development involves inactivation of tumor suppressor genes.
- Retinoblastoma (Rb) tumor suppressor gene abnormalities occur in at least 30% of malignant gliomas.
- The precise role of Rb in suppressing glioma growth remains under investigation.
Purpose of the Study:
- To investigate the therapeutic potential of restoring retinoblastoma protein function in gliomas.
- To provide evidence for the role of Rb in glioma suppression.
Main Methods:
- Constructed Ad5CMV-Rb adenovirus for exogenous Rb gene delivery.
- Assessed Rb protein expression using immunoblot and immunohistochemistry.
- Evaluated glioma cell growth suppression and cell cycle effects (G1 arrest) via growth assays and flow cytometry.
- Tested tumorigenicity of Rb-treated human glioma cells in nude mice xenografts.
Main Results:
- Transferred Rb protein was highly expressed in 80% of treated glioma cells for over 7 days.
- Treated cells exhibited loss of neoplastic morphology and significant growth suppression within 5 days.
- Rb-expressing cells predominantly arrested in the G1 phase of the cell cycle.
- Restored Rb activity prevented human glioma cells from forming tumors in vivo.
Conclusions:
- Inactivation of the retinoblastoma protein is a critical event in glioma development.
- Restoring wild-type retinoblastoma activity demonstrates therapeutic utility for gliomas.