Generation of Carrier Peptides for the Delivery of Nucleic Acid Drugs in Primary Cells

Abstract

Now that the human genome has been decoded, the demand for novel therapeutic concepts, such as gene and stem cell therapy, is higher than ever before. Although new and better pharmaceutical agents are available, their efficient delivery to the intracellular site of action is still a serious challenge. A possible solution to this problem is the use of cell-penetrating peptides as delivery vectors, including derivatives of human calcitonin (hCT). The aim of this study was to synthesise novel branched hCT-derived peptides for the noncovalent delivery of nucleic acids. The uptake of the resulting oligocationic peptides into various cell lines as well as primary cells was monitored by fluorescence microscopy. To determine the appropriate peptide-plasmid charge ratios for efficient cell transfection, electromobility shift assays were carried out. Finally, flow cytometric and fluorescence microscopic studies of gene expression highlighted two novel hCT-derived peptides as highly effective in the delivery of noncovalently complexed plasmid DNA. Thus, the absence of cytotoxicity paired with highly efficient cell internalisation and transfection rates, in primary cells as well, make both peptides powerful candidates as drug delivery vectors, especially for plasmid DNA, for both in vivo and ex vivo therapeutic applications.

Cell-penetrating peptides as a promising tool for delivery of various molecules into the cells

“…The fourth class of CPPs are peptides derived from hydrophobic sequences of proteins which naturally interact with plasma membrane, e.g. integrin b3-fragment [16], Hepatitis B virus translocation motif [17] or calcitonin fragment (Table 1) [18].…”

Specific labeling with potent radiolabels alters the uptake of cell‐penetrating peptides

“…11 All peptides were generated by automated multiple solidphase peptide synthesis (SPPS) (Syro, MultiSynTech, Bochum, Germany) using Fmoc/ t Bu strategy and synthesized as C-terminal amides by using the rink amide resin (30 mg, resin loading 0.45 mmol g À1 ). Labeling of the N-terminus with the NBD probe or the metal complex was achieved as described in Refs.…”

“…10 Since then, very promising branched structures have been developed and have served for the delivery of plasmid DNA as well as nanoparticles. 11,12 The side sequence of these branched peptides contains positive charged amino acids for complexation of negatively charged DNA or RNA molecules. However, one crucial prerequisite for a successful radionuclide labeling is that the nature of the active compound is not affected by the label.…”

Specific Targeting of Hypoxic Tumor Tissue with Nitroimidazole–Peptide Conjugates

“…[8,9] Using this method, we recently reported the successful and efficient delivery of nanocrystals, [10] oligonucleotides, [11] gallium complexes [12] and bioactive metal complexes [13][14][15][16] into cancer, as well as primary cells. However, while CPPs provide a tool through which cell-impermeable compounds can reach intracellular targets, one of their major drawbacks is their lack of selectivity towards different cell lines.…”