Erythropoietin (Epo), a 30.4-kD glycoprotein, is the principal regulator of erythropoiesis. To evaluate the concept that in vivo gene transfer might be used as an alternative to recombinant human Epo (rhEpo) in applications requiring a 1- to 3-week stimulation of erythropoiesis, the replication- deficient recombinant adenovirus AdMLP.Epo was constructed by deleting the majority of E1 from adenovirus type 5, and replacing E1 with an expression cassette containing the adenovirus type 5 major late promoter (MLP) and the human Epo gene, including the 3' cis-acting hypoxia response element. In vitro studies showed that infection of the human hepatocyte cell line Hep3B with AdMLP.Epo resulted in a 15-fold increase in Epo production in 24 hours that was enhanced to 116-fold in the presence of a hypoxic stimulus. One- time in vivo administration of AdMLP.Epo (7 x 109 plaque-forming units/kg) to the peritoneum of cotton rats caused a marked increase in red blood cell production, with a 2.6-fold increase in bone marrow erythroid precursors by day 4, and sevenfold increase in reticulocyte count by day 7. The hematocrit increased gradually, with a maximum of 64% ± 4% at day 14 (compared with an untreated baseline of 46% ± 2%), and a level of 55% ± 1% at day 24. Furthermore, one-time subcutaneous administration of AdMLP.Epo caused an increase in hematocrit that peaked at 14 days (57% ± 2%) and was still elevated at day 42. Hematocrit level in animals receiving subcutaneous administration of AdMLP.Epo sustained a long-term increase compared with animals receiving intra-peritoneal administration. In the context of these observations, gene therapy with a single administration of an adenovirus vector containing the human EPO gene may provide a means of significantly augmenting the circulating red blood cell mass over the 1- to 3-week period necessary for many clinical applications.
|Number of pages||8|
|Publication status||Published - 1 Nov 1994|
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