Retrovirus-mediated expression of an artificial β-endorphin precursor in primary fibroblasts

Andreas S. Beutler, Michaela S. Banck, Flemming W. Bach, Fred H. Gage, Frank Porreca, Edward J. Bilsky, Tony L. Yaksh

Research output: Contribution to journalArticlepeer-review

25 Scopus citations

Abstract

Peptides are of potential interest in the field of gene therapy but require modification by genetic engineering to facilitate their secretion. Amino terminal addition of a signal peptide is not always sufficient to achieve this goal, as found in this study for β-endorphin. To overcome this problem, addition of the pre-pro-sequence of mouse nerve growth factor to β- endorphin was tested. Retrovirus-mediated expression of a hybrid construct of the pre-pro-sequence of nerve growth factor and human β-endorphin in primary fibroblasts resulted in the secretion of β-endorphin immunoreactivity at a rate of 620 pg/h/106 cells. Analysis of the secreted β-endorphin immunoreactivity with reverse-phase HPLC, immunoassays using three different antibodies, and an assay for the specific displacement of [3H][D-Ala2,N- MePhe4,Gly-ol5]enkephalin from μ-opioid receptors suggests that the pre- pro-sequence is cleaved off from the pre-pro-sequence/β-endorphin construct prior to secretion, resulting in bona fide β-endorphin. Transplantation of β-endorphin-secreting cells into brain or spinal cord may provide a gene therapy approach for the treatment of chronic, opioid-sensitive pain states.

Original languageEnglish (US)
Pages (from-to)475-481
Number of pages7
JournalJournal of neurochemistry
Volume64
Issue number2
StatePublished - Feb 1995

Keywords

  • Furin
  • Gene therapy
  • Nerve growth factor
  • Opioid peptides
  • Pain
  • Pre-pro-sequence

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience
  • Biochemistry

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