RETROVIRUS-MEDIATED EXPRESSION OF AN ARTIFICIAL BETA-ENDORPHIN PRECURSOR IN PRIMARY FIBROBLASTS

Peptides are of potential interest in the field of gene therapy but re quire modification by genetic engineering to facilitate their secretio n. Amino terminal addition of a signal peptide is not always sufficien t to achieve this goal, as found in this study for beta-endorphin. To overcome this problem, addition of the pre-pro-sequence of mouse nerve growth factor to beta-endorphin was tested. Retrovirus-mediated expre ssion of a hybrid construct of the pre-pro-sequence of nerve growth fa ctor and human beta-endorphin in primary fibroblasts resulted in the s ecretion of beta-endorphin immunoreactivity at a rate of 620 pg/h/10(- 6) cells. Analysis of the secreted beta-endorphin immunoreactivity wit h reverse-phase HPLC, immunoassays using three different antibodies, a nd an assay for the specific displacement of [H-3][D-Ala(2),N-MePhe(4) ,Gly-ol(5)]enkephalin from mu-opioid receptors suggests that the pre-p ro-sequence is cleaved off from the pre-pro-sequence/beta-endorphin co nstruct prior to secretion, resulting in bona fide beta-endorphin. Tra nsplantation of beta-endorphin-secreting cells into brain or spinal co rd may provide a gene therapy approach for the treatment of chronic, o pioid-sensitive pain states.