Recombinant granulocyte colony-stimulating factor-transferrin fusion protein as an oral myelopoietic agent.
An expression construct harboring granulocyte colony-stimulating factor (G-CSF)-transferrin (Tf) fusion protein (G-CSF-Tf) was engineered by fusing human cDNAs encoding G-CSF and Tf to explore the feasibility of using Tf as a carrier moiety for oral delivery of therapeutic proteins. The recombinant protein, G-CSF-Tf, was harvested from protein-free, conditioned medium of transfected HEK293 cells. The in vitro studies demonstrated that the purified G-CSF-Tf fusion protein possesses the activity of both Tf receptor (TfR) binding in Caco-2 cells and G-CSF-dependent stimulation of NFS-60 cell proliferation. Subcutaneous administration of G-CSF-Tf fusion protein to BDF1 mice demonstrated a pharmacological effect comparable to the commercial G-CSF on the increase of absolute neutrophil counts (ANC). However, the fusion protein elicited a significant increase in ANC upon oral administration to BDF1 mice, whereas G-CSF had no effect. This study also showed that orally administered G-CSF-Tf elicits a sustained myelopoietic effect up to 3 days, whereas the s.c. administered G-CSF or G-CSF-Tf lasts only 1 day. Furthermore, coadministration of free Tf abolished the increase of ANC by orally delivered G-CSF-Tf, suggesting that the recombinant protein is absorbed via a TfR-mediated process in the gastrointestinal tract. Taken together, we conclude that the Tf-based recombinant fusion protein technology represents a promising approach for future development of orally effective peptide and protein drugs.
Bai
Yun
Y
Department of Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90033, USA.
Ann
David K
DK
Shen
Wei-Chiang
WC
D000284:Administration, Oral; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects*; D018076:DNA, Complementary / Q000235:genetics; D041981:Gastrointestinal Tract / Q000378:metabolism; D016179:Granulocyte Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D008297:Male; D051379:Mice; D038042:Myelopoiesis / Q000187:drug effects*; D009504:Neutrophils; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D011994:Recombinant Proteins; D013997:Time Factors; D014168:Transferrin / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*
D018076:DNA, Complementary; D011994:Recombinant Proteins; D014168:Transferrin; D016179:Granulocyte Colony-Stimulating Factor