G-CSF-lentivirus administration in rats provided sustained elevated neutrophil counts and subsequent EPO-lentivirus administration increased hematocrits

J Gene Med. 2007 Jul;9(7):571-8. doi: 10.1002/jgm.1050.

Abstract

Background: Towards gene therapy treatment of patients with neutropenia, characterized by neutrophil counts < 500 cells/microl, we investigated the ability of lentivirus vectors to provide sustained granulocyte colony-stimulating factor (G-CSF) delivery and to permit transgene expression from a second virus administration in a preclinical rat model.

Methods: Rats were injected intramuscularly (IM) with 24 x 10(6) and 9 x 10(6) infectious units (IU) of a VSV-G-pseudotyped self-inactivating (SIN) lentivirus encoding rat G-CSF cDNA and containing cPPT and PRE elements. To determine the effectiveness of a second virus administration treated rats and a naïve rat received erythropoietin (EPO)-lentivirus IM. Rats were monitored for neutrophil and red blood cell production. Lentivirus antibodies were assayed by virus-neutralizing assay and ELISA.

Results: High and low dose virus administration increased neutrophil counts to 5660 +/- 930 cells/microl (mean +/- SD) and 4010 +/- 850 cells/microl, respectively, that were sustained for > 17 months and were significantly higher than controls counts of 1890 +/- 570 cells/microl (p< or =0.0002). Rats treated with EPO-virus produced significantly increased hematocrits (HCT) (63.1 +/- 4.3% vs. 46.0 +/- 3.2%, p < 0.0001) without effect on G-CSF-virus-mediated neutrophil production. Antivirus antibodies were not detectable at serum dilutions > or =1:10 by virus neutralization or ELISA. Lymphocytes and platelets were not significantly different between control and treated animals (p > 0.1). Only genomic DNA from muscle at injection sites was positive for provirus suggesting lack of virus spread.

Conclusions: G-CSF-lentivirus administered IM provided elevated, sustained neutrophil counts that were unchanged by subsequent EPO-lentivirus administration. Significantly increased hematocrits were obtained following EPO-lentivirus delivery. These data support the treatment of patients with severe chronic neutropenia by dosed lentivirus delivery IM.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Antibodies, Viral / blood
  • Blood Cell Count
  • DNA / metabolism
  • Erythropoietin / genetics*
  • Erythropoietin / metabolism*
  • Genetic Therapy*
  • Genome / genetics
  • Granulocyte Colony-Stimulating Factor / genetics*
  • Granulocyte Colony-Stimulating Factor / metabolism*
  • HeLa Cells
  • Hematocrit
  • Humans
  • Lentivirus / genetics*
  • Male
  • Neutrophils / cytology*
  • Proviruses / metabolism
  • Rats
  • Rats, Inbred F344

Substances

  • Antibodies, Viral
  • Erythropoietin
  • Granulocyte Colony-Stimulating Factor
  • DNA