Long-term expression of erythropoietin in the systemic circulation of mice after intramuscular injection of a plasmid DNA vector

Abstract

Erythropoietin (Epo)-responsive anemia is a common and debilitating complication of chronic renal failure and human immunodeficiency virus infection. Current therapy for this condition involves repeated intravenous or subcutaneous injections of recombinant of recombinant Epo. In this report, we describe the development of a novel muscle-based gene transfer approach that produces long-term expression of physiologically significant levels of Epo in the systemic circulation of mice. We have constructed a plasmid expression vector, pVRmEpo, that contains the murine Epo cDNA under the transcriptional control of the cytomegalovirus immediate early (CMV-IE) promoter, the CMV-IE 5' untranslated region, and intron A. A single intramuscular (i.m.) injection of as little as 10 μg of this plasmid into immunocompetent adult mice produced physiologically significant elevations is serum Epo levels and increased hematocrits form preinjection levels of 48 ± 0.4% to levels of 64 ± 3.3% 45 days after injection. Hematocrits in these animals remained elevated at greater than 60% for at least 90 days after a single i.m. injection of 10 μg of pVRmEpo. We observed a dose-response relationship between the amount of plasmid DNA injected and subsequent elevations in hematocrits. Mice injected once with 300 μg of pVRmEpo displays 5-fold increased serum Epo levels and elevated hematocrits of 79 ± 3.3% at 45 days after injection. The i.m. injected of plasmid DNA represents a viable nonviral gene transfer method for the treatment of acquired and inherited serum protein deficiencies.