Stimulation of erythropoiesis by in vivo gene therapy: physiologic consequences of transfer of the human erythropoietin gene to experimental animals using an adenovirus vector.
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 10(9) 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 intraperitoneal 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.
Setoguchi
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Y
Pulmonary Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD.
Danel
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C
Crystal
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RG
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D004920:Erythropoiesis; D004921:Erythropoietin / Q000008:administration & dosage*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D000860:Hypoxia; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D016205:Sigmodontinae
D004921:Erythropoietin