Efficient Intracellular Delivery of Cell-Impermeable Cargo Molecules by Peptides Containing Tryptophan and Histidine

Abstract

We have previously evaluated and reported numerous classes of linear and cyclic peptides containing hydrophobic and hydrophilic segments for intracellular delivery of multiple molecular cargos. Herein, a combination of histidine and tryptophan amino acids were designed and evaluated for their efficiency in intracellular delivery of cell-impermeable phosphopeptides and the anti-HIV drug, emtricitabine. Two new decapeptides, with linear and cyclic natures, both containing alternate tryptophan and histidine residues, were synthesized using Fmoc/tBu solid-phase chemistry. The peptides were characterized and purified by using matrix-assisted laser desorption/ionization (MALDI) spectroscopy and high-performance liquid chromatography (HPLC), respectively. These peptides did not show significant toxicity up to 100 µM in ovarian cancer (SK-OV-3) and leukemia cancer (CCRF-CEM) cells. Furthermore, the cellular uptake of a fluorescence (F’)-labeled cell-impermeable phosphopeptide (F’-GpYEEI) was enhanced in the presence of linear (WH)5 and cyclic [WH]5 by 2- and 8-fold, respectively, compared to the uptake of the phosphopeptide alone. The cellular uptake was not significantly changed in the presence of endocytosis inhibitors. Furthermore, the intracellular uptake of the fluorescently-labeled anti-HIV drug, emtricitabine (F’-FTC), by linear (WH)5 and cyclic [WH]5 in SK-OV-3 cancer cell lines was found to be enhanced by 3.5- and 9-fold, respectively, compared to that of the drug alone. Fluorescent uptake experiments confirmed the localization of F’-GpYEEI-loaded cyclic [WH]5 intracellularly in the SK-OV-3 cancer cell line after 3 h of incubation. Thus, these data demonstrated that [WH]5 containing tryptophan and histidine enhanced the cellular uptake of F’-GpYEEI and emtricitabine.

Cyclic Peptide-Gadolinium Nanoparticles for Enhanced Intracellular Delivery

“…In recent years, synthetically engineered cyclic peptides have been utilized as drug delivery systems for transporting various types of molecular cargos such as anticancer drugs and siRNAs [16]. The cyclic nature of peptides has been discovered to be critically important to enhance their intracellular transporting efficiency [17][18][19]. We introduced a class of homochiral L-cyclic peptides with enhanced potency in nuclear targeting delivery of anti-HIV drugs and biomolecules [20].…”

Considerations on the Rational Design of Covalently Conjugated Cell-Penetrating Peptides (CPPs) for Intracellular Delivery of Proteins: A Guide to CPP Selection Using Glucarpidase as the Model Cargo Molecule

“…After excluding cyclic peptides and CPPs that had un-natural residues or amino acids with D-conformations, 1155 unique CPPs remained. Although some of the excluded CPPs—CPPs with unnatural or D-conformation residues and cyclic CPPs—have shown promising results in experimental studies [81,82,83], current computational web servers are unable to analyze these sequences. Mature CPG2 amino acid sequence from Pseudomonas sp.…”

A Global Review on Short Peptides: Frontiers and Perspectives

“…We have reported the application of cyclic CPPs containing alternatively positively charged arginine and hydrophobic tryptophan residues, [WR] 4 and [WR] 5 , as drug delivery tools and nuclear targeting tools in 2011 [ 209 ]. Several other monocyclic CPPs were engineered based on this template and were shown to be efficient molecular transporters for enhancing the efficacy of existing chemotherapeutic, antiviral, and antibacterial agents [ 210 , 211 , 212 , 213 , 214 , 215 , 216 , 217 , 218 , 219 ]. The monocyclic CPPs containing tryptophan and arginine residues were also used to conjugate with potential therapeutic agents.…”

“…Adding to the previous work, we also demonstrated that several monocyclic peptides containing cysteine and arginine residues, such as cyclic [CR] 4 , significantly enhanced the cellular uptake of a cell-impermeable phosphopeptide (F’)-Gly-(pTyr)-Glu-Glu-Ile (F’-GpYEEI) in the presence of endocytic inhibitors and sodium azide in the lymphoblastic leukemia cell line (CCRF-CEM) [ 218 ]. Furthermore, tryptophan and histidine-containing cyclic decapeptides [WH] 5 demonstrated an increase of the intracellular delivery of a cell-impermeable phosphopeptide and an anti-HIV drug, emtricitabine [ 217 ]. In another effort, monocyclic [HR] 4 peptides were used as a molecular transporter and were able to double the permeability of F’-GpYEEI in human ovarian adenocarcinoma cells (SK-OV-3) cells [ 214 ].…”