Hypoxia-response plasmid vector producing bcl-2 shRNA enhances the apoptotic cell death of mouse rectum carcinoma.
Hypoxia-induced gene expression frequently occurs in malignant solid tumors because they often have hypoxic areas in which circulation is compromised due to structurally disorganized blood vessels. Hypoxia-response elements (HREs) are responsible for activating gene transcription in response to hypoxia. In this study, we constructed a hypoxia-response plasmid vector producing short hairpin RNA (shRNA) against B-cell leukemia/lymphoma-2 (bcl-2), an anti-apoptotic factor. The hypoxia-response promoter was made by inserting tandem repeats of HREs upstream of cytomegalovirus (CMV) promoter (HRE-CMV). HRE-CMV shbcl-2 vector consisted of bcl-2 shRNA under the control of HRE-CMV promoter. In hypoxic mouse rectum carcinoma cells (colon-26), the production of bcl-2 shRNA driven by HRE-CMV promoter was approximately 2-fold greater than that driven by CMV promoter. A single intratumoral (i.t.) injection of 40 microg HRE-CMV shbcl-2 to colon-26 tumor-bearing mice caused apoptotic cell death, and repetitive treatment with HRE-CMV shbcl-2 (40 microg/mouse, i.t.) also significantly suppressed the growth of colon-26 tumor cells implanted in mice. Apoptotic and anti-tumor effects were not observed in tumor-bearing mice treated with CMV shbcl-2. These results reveal the ability of HRE-CMV shbcl-2 vector to suppress the expression of bcl-2 in hypoxic tumor cells and suggest the usefulness of our constructed hypoxia-response plasmid vector to treat malignant tumors. [Supplementary Figures: available only at http://dx.doi.org/10.1254/jphs.10054FP].
Fujioka
Takashi
T
Department of Pharmaceutics, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
Matsunaga
Naoya
N
Okazaki
Hiroyuki
H
Koyanagi
Satoru
S
Ohdo
Shigehiro
S
D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D015153:Blotting, Western; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D003587:Cytomegalovirus / Q000235:genetics; D005822:Genetic Vectors; D051379:Mice; D010957:Plasmids; D011401:Promoter Regions, Genetic; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics*; D012004:Rectal Neoplasms / Q000473:pathology*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering