1044. Radioprotective Gene Therapy through Retroviral Expression of Manganese Superoxide Dismutase

Abstract

Overexpression of manganese superoxide dismutase (MnSOD, also known as SOD2) has been hypothesized to provide haemopoietic cells with radioprotection through the scavenging of oxygen radicals induced by ionizing radiation. Here we demonstrate, using both biological and physical assays, that overexpression of SOD2 substantially protects both murine bone marrow and K562 cells from ionising radiation injury. For this study the human SOD2 cDNA was cloned within a retroviral vector (SF|[beta]|91), optimised for initiation of transcription in primitive haemopoietic cells, along with eGFP as a marker. Retroviral producer lines were established and used to infect murine bone marrow and the human erythroleukaemic cell line, K562. Cells were characterised by Western and Southern blotting and shown to be polyclonal by LAM PCR. SOD enzymatic activity was assessed using a water-soluble tetrazolium salt microplate assay, whilst survival was determined by assessing colony forming ability after irradiation of cells at various doses. The levels of DNA strand breaks were assessed using a COMET assay.