Document Type
Article
Publication Date
8-18-2024
Abstract
Delivering nucleic acid therapeutics across cell membranes is a significant challenge. Cell-penetrating peptides (CPPs) containing arginine (R), tryptophan (W), and histidine (H) show promise for siRNA delivery. To improve siRNA delivery and silence a model STAT3 gene, we hypothesized that oleyl acylation to CPPs, specifically (WRH)n, would enhance STAT3 silencing efficiency in breast and ovarian cancer cells. Using Fmoc/tBu solid-phase peptide chemistry, we synthesized, purified, and characterized the oleyl-conjugated (WRH)n (n = 1–4) peptides. The peptide/siRNA complexes were non-cytotoxic at N/P 40 (~20 μM) against MDA-MB-231, MCF-7, SK-OV-3, and HEK-293 cells after 72 h incubation. All peptide/siRNA complexes showed serum stability at N/P ≥ 40. The synthesized conjugates, with a diameter of <100 nm, formed nano-complexes with siRNA and exhibited a stable range of zeta potential values (13–18 mV at N/P = 40). Confocal microscopy and flow cytometry analysis provided qualitative and quantitative evidence of a successful cellular internalization of siRNA. The peptides oleyl-(WRH)3 and oleyl-(WRH)4 showed ~60% and ~75% cellular uptake of siRNA, respectively, in both MDA-MB-231 and SK-OV-3 cells. Western blot analysis of oleyl-(WRH)4 demonstrated effective silencing of the STAT-3 gene, with ~75% silencing in MDA-MB-231 cells and ~45% in SK-OV-3 cells.
Recommended Citation
Rai, M.S.; Sajid, M.I.; Moreno, J.; Parang, K.; Tiwari, R.K. Design, Synthesis, and Evaluation of Oleyl-WRH Peptides for siRNA Delivery. Pharmaceuticals 2024, 17, 1083. https://doi.org/10.3390/ph17081083
Supplementary Materials consisting of crude MALDI-TOF mass spectrometry spectra of synthesized peptides (Figures S1–S8) and purified analytical HPLC chromatograms (Figures S9–S12).
Copyright
The authors
Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 License.
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Amino Acids, Peptides, and Proteins Commons, Cancer Biology Commons, Cell Biology Commons, Medicinal and Pharmaceutical Chemistry Commons, Nucleic Acids, Nucleotides, and Nucleosides Commons, Oncology Commons, Therapeutics Commons
Comments
This article was originally published in Pharmaceuticals, volume 17, in 2024. https://doi.org/10.3390/ph17081083
A thesis with the same title is available here.