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Published on: September 19, 2013
Radiation-enhanced endostatin gene expression and effects of combination treatment
Xian Luo1, Melba L Andres, Tatyana M Timiryasova
1Department of Radiation Medicine, Chan Shun Pavilion, Room A-1010, 11175 Campus Street, Loma Linda University Medical Center, Loma Linda, CA 92354, USA.
Abstract:
Targeting cells that support tumor growth by administering potent angiogenesis inhibitors is currently an area of intense interest. In the present study, a unique plasmid vector for the mouse endostatin gene, pXLG-mEndo, was constructed and evaluated with and without radiation using the Lewis lung carcinoma (LLC) cell line. The physical properties of the expressed endostatin protein were validated by PCR, gel electrophoresis, and Western blot. Enzyme-linked immunosorbent and immunocytochemical analyses for the therapeutic gene demonstrated that transfected LLC cells secreted the protein into the medium. Exposure of the cells to 2 gray (Gy) gamma-rays reduced the time to reach the maximum expression level of the endostatin gene and also increased the amount of secreted endostatin protein (P<0.001). Biological activity of the endostatin was demonstrated by the inhibition of tube formation by human umbilical vein endothelial cells (HUVEC). Based on (3)H-thymidine incorporation, endostatin expression significantly depressed DNA synthesis in HUVEC and LLC cells compared to controls transfected with parental vector or no vector (P<0.005). In addition, radiation increased the efficiency of endostatin-mediated inhibition of both cell types over a 3-day period post-exposure (P<0.05 or less). Intratumoral injection of 100 small mu g pXLG-mEndo combined with 10 Gy radiation significantly delayed LLC tumor growth, especially when each modality was delivered twice (P<0.05 or less compared to all other groups). No toxicity was observed. These findings are very promising and suggest that endostatin therapy with a plasmid vector, such as pXLG-mEndo, may enhance the efficacy of radiotherapy for lung cancer.
Insights
This study shows that combining radiation with endostatin gene therapy using the pXLG-mEndo plasmid effectively inhibits Lewis lung carcinoma growth and enhances radiotherapy efficacy without toxicity.
Area of Science:
- Molecular biology
- Cancer research
- Gene therapy
Background:
- Angiogenesis inhibitors are crucial for targeting tumor growth.
- Endostatin is a potent anti-angiogenic protein with therapeutic potential.
Purpose of the Study:
- To construct and evaluate a plasmid vector (pXLG-mEndo) for the mouse endostatin gene.
- To assess the combined effects of endostatin gene therapy and radiation on Lewis lung carcinoma (LLC).
Main Methods:
- Plasmid construction and validation (PCR, gel electrophoresis, Western blot).
- Transfection of LLC cells and assessment of endostatin secretion (ELISA, immunocytochemistry).
- Evaluation of endostatin's biological activity on endothelial cells (HUVEC) and LLC cells (DNA synthesis assay).
- In vivo studies involving intratumoral injection of pXLG-mEndo and radiation therapy in LLC tumor models.
Main Results:
- pXLG-mEndo successfully expressed and secreted functional endostatin protein.
- Gamma radiation enhanced endostatin expression levels and secretion.
- Endostatin inhibited endothelial cell tube formation and DNA synthesis in HUVEC and LLC cells.
- Combined endostatin gene therapy and radiation significantly delayed LLC tumor growth with no observed toxicity.
Conclusions:
- The pXLG-mEndo plasmid is a viable tool for endostatin gene therapy.
- Combining endostatin therapy with radiotherapy shows synergistic potential for lung cancer treatment.
- This approach offers a promising strategy for enhancing cancer therapy efficacy.
