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- W3129603604 abstract "The spontaneous membrane translocating peptides (TPs) TP1, TP2 and TP3 were previously identified by high throughput screening. Dye-labelled TP1, 2 and 3 readily translocate across cell plasma membranes without forming complexes or pores, and without causing cytosolic leakage. Here, we test the ability of the TPs to deliver phalloidin across the plasma membranes of living cells. Phalloidin is a membrane-impermeable bicyclic heptapeptide that binds to F-actin and prevents cell movement. Compared to dye molecules, phalloidin is a much more challenging cargo to translocate. Success in delivering phalloidin would show that the TPs and their variants have the potential to act as a novel transportation platform for many cyclic peptide drugs. In this study, TP1, 2 and 3 were synthesized by solid phase peptide synthesis (SPPS) with a C-terminal cysteine. Phalloidin, containing a position 7 lysine, was attached to the TPs via SMCC, a heterobifunctional crosslinker. Reverse phase HPLC and MALDI-TOF mass spectrometry were used to validate SPPS. TP1, 2 and 3-SMCC-phalloidin (TPn-Ph) were individually applied for four hours to HT1080 fibrosarcoma cells in 96-well plates following serum starvation. Cells were then scratched with a 10 μL multichannel pipette and imaged every hour for 12 hours to assess wound closure, which requires cell migration. Cell migration was quantified through ImageJ by tracing gap area over time. TP1-Ph, TP2-Ph and TP3-Ph all inhibited cell movement substantially at 25 and 12 μM relative to positive (serum free media) and negative (cytochalasin D) controls. These results have direct implications for developing live cell actin staining. Overall, this work demonstrates that TP1, 2 and 3 can deliver membrane impermeable cyclic peptides to the cytosol, and contributes a platform for the future design, development and delivery of cyclic peptide drugs." @default.
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- W3129603604 date "2021-02-01" @default.
- W3129603604 modified "2023-09-25" @default.
- W3129603604 title "Delivering the Cyclic Peptide Phalloidin to the Cell Cytosol by Membrane Translocating Carrier Peptides" @default.
- W3129603604 doi "https://doi.org/10.1016/j.bpj.2020.11.1056" @default.
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