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. 2018 Jun 29;23(7):1590.
doi: 10.3390/molecules23071590.

Design, Synthesis, and Evaluation of Homochiral Peptides Containing Arginine and Histidine as Molecular Transporters

Naglaa Salem El-Sayed  1   2 , Taryn Miyake  3 , Amir N Shirazi  4 , Shang Eun Park  5 , Jimmy Clark  6 , Stephani Buchholz  7 , Keykavous Parang  8 , Rakesh Tiwari  9
Affiliations

Design, Synthesis, and Evaluation of Homochiral Peptides Containing Arginine and Histidine as Molecular Transporters

Naglaa Salem El-Sayed et al. Molecules. .

Abstract

Linear (HR)n and cyclic [HR]n peptides (n = 4,5) containing alternate arginine and histidine residues were synthesized. The peptides showed 0⁻15% cytotoxicity at 5⁻100 µM in human ovarian adenocarcinoma (SK-OV-3) cells while they exhibited 0⁻12% toxicity in human leukemia cancer cell line (CCRF-CEM). Among all peptides, cyclic [HR]₄ peptide was able to improve the delivery of a cell impermeable fluorescence-labeled phosphopeptide by two-fold. Fatty acids of different alkyl chain length were attached at the N-terminal of the linear peptide (HR)₄ to improve the molecular transporter property. Addition of fatty acyl chains was expected to help with the permeation of the peptides through the cell membrane. Thus, we synthesized seven fatty acyl derivatives of the linear (HR)₄ peptide. The peptides were synthesized using Fmoc/tBu solid phase peptide chemistry, purified by reverse-phase high-performance liquid chromatography (RP-HPLC) and characterized by matrix-assisted laser desorption/ionization (MALDI) spectrometry. The fatty acyl peptides containing C₈, C12, C14, and C18 alkyl chain did not show cytotoxicity on SK-OV-3 or CCRF-CEM cell lines up to 50 µM concentration; however, at higher concentration (100 µM), they showed mild cytotoxicity. For example, C16-(HR)₄ was also found to reduce the proliferation of SK-OV-3 cells by 11% at 50 µM and C20-(HR)₄ showed mild toxicity at 10 µM, reducing the proliferation of SK-OV-3 cells by 21%. Increase in the length of alkyl chain showed cytotoxicity to the cell lines. C20-(HR)₄ peptide showed better efficiency in translocation of F′-GpYEEI to SK-OV-3 than the phosphopeptide alone. Further investigation of C20-(HR)₄ peptide efficacy showed that the peptide could deliver doxorubicin and epirubicin into SK-OV-3 and also improved the drug antiproliferative ability. These studies provided insights into understanding the structural requirements for optimal cellular delivery of the fatty acyl-(HR)₄ peptide conjugates.

Keywords: anticancer drugs; cytotoxicity; fatty acyl peptides; histidine; molecular transporters; phosphopeptides.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Scheme 1
Scheme 1
Synthesis of l(HR)4, l(HR)5, c[HR]4, and c[HR]5.
Scheme 2
Scheme 2
Synthesis of fatty acyl conjugates of linear (HR)4 peptides.
Figure 1
Figure 1
Chemical structures of C20-(HR)2, C20-(HR)3, and C20-(HR)5.
Figure 2
Figure 2
Cytotoxicity study for l(HR)4, l(HR)5, c[HR]4, and c[HR]5 at 5, 10, 25, 50, and 100 µM in SK-OV-3 and CCRF-CEM cell lines after 24 h incubation.
Figure 3
Figure 3
Cytotoxicity study for the fatty acyl-(HR)4 derivatives at concentrations of 10, 50 and 100 µM after 72 h incubation with CCRF-CEM and SK-OV-3 cells.
Figure 4
Figure 4
Cellular uptake study of F′-GpYEEI phosphopeptide at 5 µM by c[HR]4, c[HR]5, l(HR)4, and l(HR)5 at 50 µM peptide concentration in SK-OV-3 cancer cell lines.
Figure 5
Figure 5
Cellular uptake study of F′-GpYEEI (5 µM) by fatty acyl-(HR)4 derivatives at 50 µM in SK-OV-3 cancer cell lines.
Figure 6
Figure 6
Cellular uptake studies F′-GpYEEI (5 µM) in the presence of C20-(HR)4 in SK-OV-3 cell lines at a concentration of 5, 10 and 50 µM of C20-(HR)4.
Figure 7
Figure 7
Comparative study for the cellular uptake of F′-GpYEEI (5 µM) by l(HR)4, c[HR]4, and C20-(HR)4 at 50 µM in SK-OV-3 cell line.
Figure 8
Figure 8
The cellular uptake of F′-GpYEEI at 5 µM concentration by C20-(HR)n peptides at 50 µM peptide concentration in CCRF-CEM cell line.
Figure 9
Figure 9
Anti-proliferative assay results for C20-(HR)4 with different anticancer drugs.
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