Reduction of radiation-induced apoptosis by specific expression of Bcl-2 in normal cells

Abstract

Radiation-induced apoptosis is thought to underlie the toxicity of radiation to normal tissues as well as to cancer cells. We hypothesized that specific ectopic overexpression of the antiapoptotic molecule Bcl-2 in normal cells would inhibit radiationinduced apoptosis and thereby reduce radiation-induced toxicity in normal cells. To express Bcl-2 specifically in normal cells (which have wild-type (wt) p53) but not in cancer cells (which often have mutated p53), we constructed a Bcl-2 expression plasmid (PG 13 -Bcl-2) with a minimal promoter regulated by multiple wt p53 DNA-binding sites and found that the presence of wt p53 protein strongly upregulated Bcl-2 expression through this plasmid. Transfection of NIH 3T3 fibroblasts, which express wt p53, with PG 13 -Bcl-2 increased cell survival and reduced apoptosis; however, transfection of MDA-MB-231 breast cancer cells, which have mutated p53, did not affect survival and apoptosis of those cells. These results indicate that irradiation of normal cells rapidly upregulates the expression of wt p53, which binds to the p53 binding sequence of the PG 13 -Bcl-2 plasmid and increases the transcriptional activity of Bcl-2. Ectopic expression of Bcl-2 reduced radiation-induced apoptosis only in normal cells (not in cancer cells). Bcl-2 expression was detected in the lung from mice injected via a tail vein with LPD-PG 13 -Bcl-2 or LPD-CMV-Bcl-2, but did not in the lung from mice treated with DOTAP or LPD-PG 13 -mock. This novel approach to inhibiting radiation-induced apoptosis in normal cells may allow such cells to be protected from radiation-induced toxicity. Further preclinical in vivo studies are needed.

B cell translocation gene 1 contributes to antisense Bcl-2-mediated apoptosis in breast cancer cells

“…The MDA-231 breast cancer cell line, stable control, and Bcl-2 transfectants derived from MDA-231 (17), and MCF-7 breast cancer cells were used for our study. Parental MDA-231 and control Neo MDA-231-derived stable transfectant cells express relatively low levels of endogenous Bcl-2, whereas MCF-7 cells overexpress Bcl-2 (Fig.…”

“…Bcl-2-overexpressing MDA-231 clones (referred to as clone 4 and clone 5 cells) and the Neo control stable transfectant were created as previously described (17). Briefly, the pCl-neo/ Bcl-2 (CMV-Bcl-2) plasmid encoding the full-length Bcl-2 coding sequence or the backbone vector pCl-neo was transfected into MDA-231 cells with subsequent neomycin (G418) antibiotic selection and screening of clones by immunoblot.…”