Radioprotection in vitro and in vivo by minicircle plasmid carrying the human manganese superoxide dismutase transgene.
Manganese superoxide dismutase plasmid liposomes (MnSOD-PL) confer organ-specific in vivo ionizing irradiation protection. To prepare for potential intravenous clinical trials of systemic MnSOD-PL for radioprotection in humans, plasmid and bacterial sequences were removed and a new minicircle construct was tested. Minicircle MnSOD was purified and then cotransfected into 32D cl 3 murine interleukin-3-dependent hematopoietic progenitor cells along with another plasmid carrying the neo gene. Cells were selected in G418 (50 microg/ml) and cloned by limiting dilution. Biochemical analysis of minicircle MnSOD-transfected cells showed an MnSOD biochemical activity level of 5.8 +/- 0.5 U/mg compared with 2.7 +/- 0.1 U/mg for control 32D cl 3 cells (p = 0.0039). 32D-mc-MnSOD cells were as radioresistant as full-length MnSOD-PL transgene-expressing 2C6 cells, relative to 32D cl 3 parent cells, with an increased shoulder on the radiation survival curve (n = 4.8 +/- 0.2 and n = 4.6 +/- 0.2, respectively, compared with 1.5 +/- 0.5 for 32D cl 3 cells; p = 0.007). C57BL/6NHsd mice received intraoral mc-MnSOD-PL, mc-DsRed-PL control, full-length MnSOD-PL, or blank-PL and then were irradiated 24 hr later with 31 Gy to the esophagus. Mice receiving mc-MnSOD-PL showed increased survival compared with control mice or mice treated with mc-DsRed-PL (p = 0.0003 and 0.039, respectively), and comparable to full-length MnSOD-PL. Intravenous, systemic administration of mc-MnSOD-PL protected mice from total body irradiation (9.75 Gy). Therefore, minicircle DNA containing the human MnSOD transgene confers undiminished radioprotection in vitro and in vivo.
Zhang
Xichen
X
Department of Radiation Oncology, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213, USA.
Epperly
Michael W
MW
Kay
Mark A
MA
Chen
Zhi-Ying
ZY
Dixon
Tracy
T
Franicola
Darcy
D
Greenberger
Benjamin A
BA
Komanduri
Paavani
P
Greenberger
Joel S
JS
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004307:Dose-Response Relationship, Radiation; D004947:Esophagus / Q000528:radiation effects; D005260:Female; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000008:administration & dosage; D006801:Humans; D008081:Liposomes / Q000008:administration & dosage*; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D010957:Plasmids / Q000008:administration & dosage* / Q000235:genetics; D011833:Radiation Injuries, Experimental / Q000517:prevention & control*; D011836:Radiation Tolerance / Q000235:genetics; D011839:Radiation, Ionizing; D011837:Radiation-Protective Agents / Q000627:therapeutic use*; D013482:Superoxide Dismutase / Q000235:genetics*; D014162:Transfection; D019076:Transgenes; D014916:Whole-Body Irradiation
D008081:Liposomes; D011837:Radiation-Protective Agents; D013482:Superoxide Dismutase