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Paracellular permeation-enhancing effect of AT1002 C-terminal amidation in nasal delivery.

Song KH, Kim SB, Shim CK, Chung SJ, Kim DD, Rhee SK, Choi GJ, Kim CH, Kim K - Drug Des Devel Ther (2015)

Bottom Line: Pep1 was nontoxic to cell monolayers and showed a relatively low decrease in peak area compared to AT1002.In addition, administration of mannitol with Pep1 resulted in significant increases in the area under the plasma concentration-time curve and peak plasma concentration at 3.63-fold and 2.68-fold, respectively, compared to mannitol alone.Thus, Pep1 increased the stability or possibly reduced the instability of AT1002, which resulted in an increased permeation-enhancing effect of AT1002.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Engineering, Soonchunhyang University, Asan, Republic of Korea.

ABSTRACT

Background: The identification of permeation enhancers has gained interest in the development of drug delivery systems. A six-mer peptide, H-FCIGRL-OH (AT1002), is a tight junction modulator with promising permeation-enhancing activity. AT1002 enhances the transport of molecular weight markers or agents with low bioavailability with no cytotoxicity. However, AT1002 is not stable in neutral pH or after incubation under physiological conditions, which is necessary to fully uncover its permeation-enhancing effect. Thus, we increased the stability or mitigated the instability of AT1002 by modifying its terminal amino acids and evaluated its subsequent biological activity.

Methods: C-terminal-amidated (FCIGRL-NH2, Pep1) and N-terminal-acetylated (Ac-FCIGRL, Pep2) peptides were analyzed by liquid chromatography-mass spectrometry. We further assessed cytotoxicity on cell monolayers, as well as the permeation-enhancing activity following nasal administration of the paracellular marker mannitol.

Results: Pep1 was nontoxic to cell monolayers and showed a relatively low decrease in peak area compared to AT1002. In addition, administration of mannitol with Pep1 resulted in significant increases in the area under the plasma concentration-time curve and peak plasma concentration at 3.63-fold and 2.68-fold, respectively, compared to mannitol alone. In contrast, no increase in mannitol concentration was shown with mannitol/AT1002 or mannitol/Pep2 compared to the control. Thus, Pep1 increased the stability or possibly reduced the instability of AT1002, which resulted in an increased permeation-enhancing effect of AT1002.

Conclusion: These results suggest the potential usefulness of C-terminal-amidated AT1002 in enhancing nasal drug delivery, which may lead to the development of a practical drug delivery technology for drugs with low bioavailability.

No MeSH data available.


Related in: MedlinePlus

Full-scan spectra and SIM chromatograms at each m/z of FCIGRL-NH2 (Pep1), Ac-FCIGRL (Pep2), Ac-FCIGRL-NH2 (Pep3), and FCIGRL (AT1002).Abbreviations: SIM, selected-ion monitoring.
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f1-dddt-9-1815: Full-scan spectra and SIM chromatograms at each m/z of FCIGRL-NH2 (Pep1), Ac-FCIGRL (Pep2), Ac-FCIGRL-NH2 (Pep3), and FCIGRL (AT1002).Abbreviations: SIM, selected-ion monitoring.

Mentions: Figure 1 shows that each single-charged ion with a protonated molecule [M + H]+ appeared at m/z 708.4, 707.4, 750.4, and 749.4 in the full-scan spectra under positive ESI, which corresponded precisely to the molecular mass of AT1002 and the terminally modified AT1002 peptides (Pep1, Pep2, Pep3). A peptide including one cysteine can be easily oxidized to form dimerized disulfide bonds, which can result in peptide instability due to aggregation or polymerization.18,22,23 However, we did not observe dimerization of AT1002, Pep1, Pep2, or Pep3 in each full-scan spectra due to the instability of AT1002, as shown in our previous report,13 and due to its degradation in solution. We considered that peptide dimerization was not due to the hydrophobic characteristics of phenylalanine and isoleucine flanking each cysteine in AT1002, Pep1, Pep2, and Pep3.


Paracellular permeation-enhancing effect of AT1002 C-terminal amidation in nasal delivery.

Song KH, Kim SB, Shim CK, Chung SJ, Kim DD, Rhee SK, Choi GJ, Kim CH, Kim K - Drug Des Devel Ther (2015)

Full-scan spectra and SIM chromatograms at each m/z of FCIGRL-NH2 (Pep1), Ac-FCIGRL (Pep2), Ac-FCIGRL-NH2 (Pep3), and FCIGRL (AT1002).Abbreviations: SIM, selected-ion monitoring.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4383222&req=5

f1-dddt-9-1815: Full-scan spectra and SIM chromatograms at each m/z of FCIGRL-NH2 (Pep1), Ac-FCIGRL (Pep2), Ac-FCIGRL-NH2 (Pep3), and FCIGRL (AT1002).Abbreviations: SIM, selected-ion monitoring.
Mentions: Figure 1 shows that each single-charged ion with a protonated molecule [M + H]+ appeared at m/z 708.4, 707.4, 750.4, and 749.4 in the full-scan spectra under positive ESI, which corresponded precisely to the molecular mass of AT1002 and the terminally modified AT1002 peptides (Pep1, Pep2, Pep3). A peptide including one cysteine can be easily oxidized to form dimerized disulfide bonds, which can result in peptide instability due to aggregation or polymerization.18,22,23 However, we did not observe dimerization of AT1002, Pep1, Pep2, or Pep3 in each full-scan spectra due to the instability of AT1002, as shown in our previous report,13 and due to its degradation in solution. We considered that peptide dimerization was not due to the hydrophobic characteristics of phenylalanine and isoleucine flanking each cysteine in AT1002, Pep1, Pep2, and Pep3.

Bottom Line: Pep1 was nontoxic to cell monolayers and showed a relatively low decrease in peak area compared to AT1002.In addition, administration of mannitol with Pep1 resulted in significant increases in the area under the plasma concentration-time curve and peak plasma concentration at 3.63-fold and 2.68-fold, respectively, compared to mannitol alone.Thus, Pep1 increased the stability or possibly reduced the instability of AT1002, which resulted in an increased permeation-enhancing effect of AT1002.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Engineering, Soonchunhyang University, Asan, Republic of Korea.

ABSTRACT

Background: The identification of permeation enhancers has gained interest in the development of drug delivery systems. A six-mer peptide, H-FCIGRL-OH (AT1002), is a tight junction modulator with promising permeation-enhancing activity. AT1002 enhances the transport of molecular weight markers or agents with low bioavailability with no cytotoxicity. However, AT1002 is not stable in neutral pH or after incubation under physiological conditions, which is necessary to fully uncover its permeation-enhancing effect. Thus, we increased the stability or mitigated the instability of AT1002 by modifying its terminal amino acids and evaluated its subsequent biological activity.

Methods: C-terminal-amidated (FCIGRL-NH2, Pep1) and N-terminal-acetylated (Ac-FCIGRL, Pep2) peptides were analyzed by liquid chromatography-mass spectrometry. We further assessed cytotoxicity on cell monolayers, as well as the permeation-enhancing activity following nasal administration of the paracellular marker mannitol.

Results: Pep1 was nontoxic to cell monolayers and showed a relatively low decrease in peak area compared to AT1002. In addition, administration of mannitol with Pep1 resulted in significant increases in the area under the plasma concentration-time curve and peak plasma concentration at 3.63-fold and 2.68-fold, respectively, compared to mannitol alone. In contrast, no increase in mannitol concentration was shown with mannitol/AT1002 or mannitol/Pep2 compared to the control. Thus, Pep1 increased the stability or possibly reduced the instability of AT1002, which resulted in an increased permeation-enhancing effect of AT1002.

Conclusion: These results suggest the potential usefulness of C-terminal-amidated AT1002 in enhancing nasal drug delivery, which may lead to the development of a practical drug delivery technology for drugs with low bioavailability.

No MeSH data available.


Related in: MedlinePlus