Abstract
Superoxide dismutase (SOD) fusion of TAT was proved to be radioprotective in our previous work. On that basis, a bifunctional recombinant protein which was the fusion of glutathione S-transferase (GST), SOD, and TAT was constructed and named GST-TAT-SOD. Herein we report the investigation of the cytotoxicity, cell-penetrating activity, and in vitro radioprotective effect of GST-TAT-SOD compared with wild SOD, single-function recombinant protein SOD-TAT, and amifostine. We demonstrated that wild SOD had little radioprotective effect on irradiated L-02 and Hep G2 cells while amifostine was protective to both cell lines. SOD-TAT or GST-TAT-SOD pretreatment 3 h prior to radiation protects irradiated normal liver cells against radiation damage by eliminating intracellular excrescent superoxide, reducing cellular MDA level, enhancing cellular antioxidant ability and colony formation ability, and reducing apoptosis rate. Compared with SOD-TAT, GST-TAT-SOD was proved to have better protective effect on irradiated normal liver cells and minimal effect on irradiated hepatoma cells. Besides, GST-TAT-SOD was safe for normal cells and effectively transduced into different organs in mice, including the brain. The characteristics of this protein suggest that it may be a potential radioprotective agent in cancer therapy better than amifostine. Fusion of two antioxidant enzymes and cell-penetrating peptides is potentially valuable in the development of radioprotective agent.
Highlights
As a component of therapy for a wide range of malignant conditions, radiotherapy is estimated to be used by half of all cancer patients during the course of their treatment for cancer
Amifostine is the only clinical radioprotector approved by the Food and Drug Administration (FDA) for head and neck cancer patients [3]
E. coli strains with recombinant plasmid of glutathione S-transferase (GST)-TAT-Superoxide dismutase (SOD) containing GST, TAT-PTD, and human Cu/Zn-SOD and recombinant protein SODTAT were obtained from Institute of Biotechnology, Fuzhou University (Fujian, China)
Summary
As a component of therapy for a wide range of malignant conditions, radiotherapy is estimated to be used by half of all cancer patients during the course of their treatment for cancer. Superoxide radicals produced by ionizing radiation are highly reactive and potentially damaging to cells. The hypothesis that SOD is radioprotective has been supported by many studies through transgenic experiments [10,11,12,13,14,15], there were many limitations on its protecting against radiation-induced chronic injury in humans. Some of them have been proved to be radioprotective in various radiation injury models [16] Their reaction efficiency of scavenging superoxide anion is still inferior to wild SOD. Their mechanism, selectivity, and toxicity of mimics may vary compared with natural enzyme [16]. Current study investigated the selective radioprotective effects of this cell permeable bifunctional antioxidant enzyme compared with SOD-TAT and amifostine
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