Production of Recombinant Human Growth Hormone Conjugated with a Transcytotic Peptide in Pichia pastoris for Effective Oral Protein Delivery

Abstract

Among the possible delivery routes, the oral administration of a protein is simple and achieves high patient compliance without pain. However, the low bioavailability of a protein drug in the intestine due to the physical barriers of the intestinal epithelia is the most critical problem that needs to be solved. To overcome the low bioavailability of a protein drug in the intestine, we aimed to construct a recombinant Pichia pastoris expressing a human growth hormone (hGH) fusion protein conjugated with a transcytotic peptide (TP) that was screened through peroral phage display to target goblet cells in the intestinal epithelia. The TP-conjugated hGH was successfully produced in P. pastoris in a secreted form at concentrations of up to 0.79 g/l. The function of the TP-conjugated hGH was validated by in vitro and in vivo assays. The transcytotic function of the TP through the intestinal epithelia was verified only in the C terminus conjugated hGH, which demonstrated the induction of IGF-1 in a HepG2 cell culture assay, a higher translocation of recombinant hGH into the ileal villi after oral administration in rats and both IGF-1 induction and higher body weight gain in rats after oral administration. The present study introduces the possibility for the development of an effective oral protein delivery system in the pharmaceutical and animal industries through the introduction of an effective TP into hGH.

Faculty Opinions recommendation of Intestinal macromolecular transport supporting adaptive immunity.

“…39 Interestingly, the endocytic properties of intestinal goblet cells to take up luminal substances has been noted for decades, [40][41][42][43] and intriguingly this property of goblet cells is being leveraged for oral drug delivery. [44][45][46][47][48][49] Observations support that LP-APCs acquiring luminal substances via GAPs are effective at inducing antigen-specific T-cell responses. When goblet cells and GAPs are absent or when GAPs are inhibited, LP-APCs cannot acquire luminal substances in a manner capable of stimulating antigen-specific T-cell responses in ex vivo assays.…”

Oral Administration of Recombinant Saccharomyces boulardii Expressing Ovalbumin-CPE Fusion Protein Induces Antibody Response in Mice

“…In this study, we have used c-CPE for tight junction targeting and augmentation of intestinal absorption of foreign antigens. However, this process can be further improved by addition of other ligands such as transcytotic peptides (TP) ( Kang et al, 2008 ; Lee et al, 2015 ), which have been recommended for intestinal absorption of large molecules such as therapeutic peptides.…”

Intestinal Macromolecular Transport Supporting Adaptive Immunity

“…Recently, it was identified that goblet cells can take up luminal high-molecular-weight substances and transfer them to LP-APCs in a process termed goblet cell–associated antigen passages (GAPs) 39 . Interestingly, the endocytic properties of intestinal goblet cells to take up luminal substances has been noted for decades,40, 41, 42, 43 and intriguingly this property of goblet cells is being leveraged for oral drug delivery 44, 45, 46, 47, 48, 49. Observations support that LP-APCs acquiring luminal substances via GAPs are effective at inducing antigen-specific T-cell responses.…”