Conjugation of a cell-penetrating peptide to parathyroid hormone affects its structure, potency, and transepithelial permeation
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Conjugation of a cell-penetrating peptide to parathyroid hormone affects its structure, potency, and transepithelial permeation. / Kristensen, Mie; de Groot, Anne Marit; Berthelsen, Jens; Franzyk, Henrik; Sijts, Alice; Nielsen, Hanne Mørck.
In: Bioconjugate Chemistry, Vol. 26, 18.03.2015, p. 477-488.Research output: Contribution to journal › Journal article › Research › peer-review
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T1 - Conjugation of a cell-penetrating peptide to parathyroid hormone affects its structure, potency, and transepithelial permeation
AU - Kristensen, Mie
AU - de Groot, Anne Marit
AU - Berthelsen, Jens
AU - Franzyk, Henrik
AU - Sijts, Alice
AU - Nielsen, Hanne Mørck
PY - 2015/3/18
Y1 - 2015/3/18
N2 - Delivery of therapeutic peptides and proteins by the use of cell-penetrating peptides (CPPs) as carriers has been suggested as a feasible strategy. The aim of the present study was to investigate the effect of conjugating a series of well-known CPPs to the biologically active part of parathyroid hormone, i.e. PTH(1-34), and to evaluate the effect with regards to secondary structure, potency in Saos-2 cells, immunogenicity, safety as well as the transepithelial permeation across monolayers by using the Caco-2 cell culture model. Further, co-administration of CPP and PTH(1-34) as an alternative to covalent conjugation was compared with regards to the transepithelial permeation. CPP-conjugated PTH(1-34) fusion peptides were successfully expressed in Escherichia coli and purified from inclusion bodies. No clear correlation between the degree of secondary structure of the CPP-conjugated PTH(1-34) fusion peptides and their potency was found, albeit a general decrease in permeation was observed for both N- and C-terminally CPP-conjugated PTH(1-34) as compared to native PTH(1-34). However, attachment of CPP to the N-terminus significantly increased permeation across Caco-2 cell monolayers as compared to the corresponding C-terminally CPP-conjugated PTH(1-34). In addition, the nonaarginine sequence proved to be the only CPP capable of increasing permeation when conjugated to PTH(1-34) as compared to co-administration of CPP and PTH(1-34). This enhancement effect was, however, associated with an unacceptable low level of cell viability. In conclusion, covalent conjugation of CPPs to PTH(1-34) influenced both the secondary structure, potency, and transepithelial permeation efficiency of the resulting conjugate, and hence this approach appears not to be favorable as compared to co-administration when optimizing CPP-mediated permeation of PTH(1-34) across an intestinal epithelium.
AB - Delivery of therapeutic peptides and proteins by the use of cell-penetrating peptides (CPPs) as carriers has been suggested as a feasible strategy. The aim of the present study was to investigate the effect of conjugating a series of well-known CPPs to the biologically active part of parathyroid hormone, i.e. PTH(1-34), and to evaluate the effect with regards to secondary structure, potency in Saos-2 cells, immunogenicity, safety as well as the transepithelial permeation across monolayers by using the Caco-2 cell culture model. Further, co-administration of CPP and PTH(1-34) as an alternative to covalent conjugation was compared with regards to the transepithelial permeation. CPP-conjugated PTH(1-34) fusion peptides were successfully expressed in Escherichia coli and purified from inclusion bodies. No clear correlation between the degree of secondary structure of the CPP-conjugated PTH(1-34) fusion peptides and their potency was found, albeit a general decrease in permeation was observed for both N- and C-terminally CPP-conjugated PTH(1-34) as compared to native PTH(1-34). However, attachment of CPP to the N-terminus significantly increased permeation across Caco-2 cell monolayers as compared to the corresponding C-terminally CPP-conjugated PTH(1-34). In addition, the nonaarginine sequence proved to be the only CPP capable of increasing permeation when conjugated to PTH(1-34) as compared to co-administration of CPP and PTH(1-34). This enhancement effect was, however, associated with an unacceptable low level of cell viability. In conclusion, covalent conjugation of CPPs to PTH(1-34) influenced both the secondary structure, potency, and transepithelial permeation efficiency of the resulting conjugate, and hence this approach appears not to be favorable as compared to co-administration when optimizing CPP-mediated permeation of PTH(1-34) across an intestinal epithelium.
U2 - 10.1021/bc5005763
DO - 10.1021/bc5005763
M3 - Journal article
C2 - 25611217
VL - 26
SP - 477
EP - 488
JO - Bioconjugate Chemistry
JF - Bioconjugate Chemistry
SN - 1043-1802
ER -
ID: 130643656