Stimulation of erythropoiesis by in vivo gene therapy

Physiologic consequences of transfer of the human erythropoietin gene to experimental animals using an adenovirus vector

Yasuhiro Setoguchi, Claire Danel, Ronald Crystal

Research output: Contribution to journalArticle

72 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)2946-2953
Number of pages8
JournalBlood
Volume84
Issue number9
Publication statusPublished - 1 Nov 1994
Externally publishedYes

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Gene therapy
Erythropoiesis
Erythropoietin
Adenoviridae
Genetic Therapy
Animals
Genes
Hematocrit
Cells
Blood
Erythrocytes
Sigmodontinae
Erythrocyte Volume
Gene transfer
Reticulocyte Count
Peritoneum
Response Elements
Cotton
Rats
Hepatocytes

ASJC Scopus subject areas

  • Hematology

Cite this

Stimulation of erythropoiesis by in vivo gene therapy : Physiologic consequences of transfer of the human erythropoietin gene to experimental animals using an adenovirus vector. / Setoguchi, Yasuhiro; Danel, Claire; Crystal, Ronald.

In: Blood, Vol. 84, No. 9, 01.11.1994, p. 2946-2953.

Research output: Contribution to journalArticle

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abstract = "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.",
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