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Published on: July 30, 2018
Adenovirus-mediated Bak gene transfer induces apoptosis in mesothelioma cell lines
1Department of Thoracic and Cardiovascular Surgery, and the Department of Molecular Pathology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas 77030, USA.
Objective:
Conventional treatment for mesothelioma is largely ineffective. We therefore evaluated the novel approach of adenoviral gene transfer of the proapoptotic Bcl-2 family member Bak in mesothelioma cancer cell lines, which are sensitive and resistant to adenoviral p53.
Methods:
Binary adenoviral Bak (Ad/GT-Bak and Ad/GV16) and LacZ (Ad/GT-LacZ and Ad/GV16) vectors were used for transduction of the mesothelioma cell lines I-45 (p53 resistant) and REN (p53 sensitive). Protein levels were determined by Western blotting. Apoptosis was assessed by morphologic changes, caspase-3 cleavage, and fluorescence-activated cell sorter analysis of subdiploid populations. Cell viability was determined with the XTT assay. Statistical analysis was performed with analysis of variance and the Student t test.
Results:
High levels of Bak gene transfer were seen after coadministration of Ad/GT-Bak and Ad/GV16 in both mesothelioma cell lines. Apoptosis was induced 24 hours after Bak but not LacZ gene transfer ([Bak: I-45, 36%; REN, 25%] vs [LacZ: I-45, 1%; REN, 3%], P <.05]) in p53-sensitive (REN) and p53-resistant (I-45) cell lines. Cellular viability was significantly decreased 48 to 72 hours after Bak gene transfer compared with control vector in both cell lines (72 hours: Bak I-45, 1.4% +/- 1.0%, and Bak REN, 4.7% +/- 1%, vs Lac-Z I-45, 83% +/- 3%, and Lac-Z REN, 100% +/- 1%; P <.05).
Conclusions:
Adenovirus-mediated overexpression of the Bak gene induces apoptosis and decreased cellular viability in p53-sensitive and p53-resistant mesothelioma cells. These data suggest that the gene transfer of proapoptotic Bcl-2 family members may represent a novel gene therapy strategy to treat mesothelioma.
Insights
Adenoviral gene transfer of the Bak gene effectively induced apoptosis and reduced cell viability in both p53-sensitive and p53-resistant mesothelioma cells, offering a potential new gene therapy approach.
Area of Science:
- Oncology
- Gene Therapy
- Molecular Biology
Background:
- Mesothelioma treatment remains a significant challenge due to the ineffectiveness of conventional therapies.
- The proapoptotic Bcl-2 family member Bak presents a potential therapeutic target for cancer treatment.
Purpose of the Study:
- To evaluate the efficacy of adenoviral gene transfer of Bak in mesothelioma cell lines.
- To determine if Bak gene transfer can induce apoptosis and reduce cell viability in both p53-sensitive and p53-resistant mesothelioma cells.
Main Methods:
- Utilized binary adenoviral vectors (Ad/GT-Bak and Ad/GV16) for Bak gene transfer into mesothelioma cell lines I-45 (p53 resistant) and REN (p53 sensitive).
- Assessed protein levels via Western blotting, apoptosis through morphological changes, caspase-3 cleavage, and subdiploid populations (FACS).
- Cell viability was measured using the XTT assay, with statistical analysis by ANOVA and Student's t-test.
Main Results:
- High Bak gene expression was achieved following coadministration of Ad/GT-Bak and Ad/GV16 in both cell lines.
- Bak gene transfer significantly induced apoptosis (I-45: 36%, REN: 25%) compared to LacZ control (P <.05).
- Cellular viability was markedly reduced 48-72 hours post-Bak gene transfer (e.g., 72 hours: Bak I-45 1.4%, Bak REN 4.7%) versus controls (LacZ I-45 83%, LacZ REN 100%; P <.05).
Conclusions:
- Adenovirus-mediated Bak gene overexpression effectively triggers apoptosis and diminishes cell viability in mesothelioma cells, irrespective of p53 status.
- Gene transfer of proapoptotic Bcl-2 family members, like Bak, shows promise as a novel gene therapy strategy for mesothelioma treatment.
