Structure-function studies of vector peptides: Increasing the bioavailability of bioactive compounds.

摘要

Cell-permeable “vector” peptides (vps) are lysine- and arginine-rich import sequences that carry unrelated molecular cargoes across cellular membranes without relying on classical endocytosis. Vector peptides have already been used to facilitate the intracellular delivery of other small peptides, oligonucleotides, and pharmacological cargoes. Furthermore, a recent report suggests the potential for vector peptides to translocate across the blood-brain barrier (BBB).; The purpose of this project was to advance vector peptide technology. Our goal was to improve the efficient delivery of intact, bioactive cargo compounds across cellular membranes. We used Antennapedia (Antp)-related, the HIV-1 transactivator of transcription (TAT)-derived, and arginine (Arg)-7 vector peptides and various model cargo peptides to test the hypothesis that efficient cellular penetration by vector peptides can be improved by pursuing novel coupling strategies and vector modifications. We tested the hypothesis that such novel coupling and modification strategies enhanced both the bioavailability and biological potency of intact cargoes. Our experiments revealed that vectorized peptides consistently retained their biological activity. Our results also indicated appreciable cellular penetration of impermeable peptides, such as a hexapeptide HIV-1 integrase inhibitor that exhibited unexpected cytotoxicity. Importantly, we found that internalization efficiency is complicated by the fact that cationic vector peptides bind tightly to inert surfaces. A corollary to this finding, our results indicated that micromolar concentrations of vectorized cargo are necessary to achieve appreciable internalization. We further clarified a hierarchy of efficient cellular penetration by different vector peptide families, where [Arg-7]-vp proved significantly more superior than Antp- and TAT-vps. In addition, we wanted to test the capability for vector peptides to transcytose cellular barriers. Utilizing a co-culture BBB model, our preliminary results suggested that the [Pro50] Antp-vp could carry intact bioactive cargoes across a cellular barrier.; This project helped demonstrate the feasibility of cationic vector peptides to internalize several different cargo peptides. These results also suggested that the unique structural properties that allow vps to penetrate cells might be responsible for their unexpectedly non-specific attachment to inert surfaces, such as glass. Therefore, findings contained herein may prove useful in expanding the utility of vector peptide technology in a broad range of biomedical applications.