Normal-tissue radioprotection by overexpression of the copper-zinc and manganese superoxide dismutase genes.
OBJECTIVE
Protection of normal tissue against radiation-induced damage may increase the therapeutic ratio of radiotherapy. A promising strategy for testing this approach is gene therapy-mediated overexpression of the copper-zinc (CuZnSOD) or manganese superoxide dismutase (MnSOD) using recombinant adeno-associated viral (rAAV2) vectors. The purpose of this study was to test the modulating effects of the SOD genes on human primary lung fibroblasts (HPLF) after irradiation.


METHODS
HPLF were transduced with rAAV2 vectors containing cDNA for the CuZnSOD, MnSOD or a control gene. The cells were irradiated (1-6 Gy), and gene transfer efficiency, apoptosis, protein expression/activity, and radiosensitivity measured by the colony-forming assay determined.


RESULTS
After transduction, 90.0% +/- 6.4% of the cells expressed the transgene. A significant fivefold overexpression of both SOD was confirmed by an SOD activity assay (control: 21.1 +/- 12.6, CuZnSOD: 95.1 +/- 17.1, MnSOD: 108.5 +/- 36.0 U SOD/mg protein) and immunohistochemistry. CuZnSOD and MnSOD overexpression resulted in a significant radioprotection of HPLF compared to controls (surviving fraction [SF] ratio SOD/control > 1): CuZnSOD: 1.18-fold (95% confidence interval [CI]: 1.06-1.32; p = 0.005), MnSOD: 1.23-fold (95% CI: 1.07-1.43; p = 0.01).


CONCLUSIONS
Overexpression of CuZnSOD and MnSOD in HPLF mediated an increase in clonogenic survival after irradiation compared to controls. In previous works, a lack of radioprotection in SOD-overexpressing tumor cells was observed. Therefore, the present results suggest that rAAV2 vectors are promising tools for the delivery of radioprotective genes in normal tissue.
Veldwijk
Marlon R
MR
Department of Radiation Oncology, University Medical Center Mannheim, University of Heidelberg, Mannheim, Germany.
Herskind
Carsten
C
Sellner
Leopold
L
Radujkovic
Aleksandar
A
Laufs
Stephanie
S
Fruehauf
Stefan
S
Zeller
W Jens
WJ
Wenz
Frederik
F
D017209:Apoptosis / Q000235:genetics / Q000528:radiation effects; D002460:Cell Line; D002470:Cell Survival / Q000235:genetics* / Q000528:radiation effects*; D003114:Colony-Forming Units Assay; D000229:Dependovirus; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D018014:Gene Transfer Techniques; D006801:Humans; D008856:Microscopy, Fluorescence; D011836:Radiation Tolerance / Q000235:genetics*; D013482:Superoxide Dismutase / Q000235:genetics*
D013482:Superoxide Dismutase