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Diaz, Delmy

Chemistry

2003

M.S.

The discovery of cell-penetrating peptides and their efficient transduction of cargoes into the cells have opened new possibilities for medical treatments. Some drugs, genes, antisense oligonucleotides and proteins can, when covalently attached to a CPP, translocate the cell membrane into the interior of the cell and therefore be effective to treat different diseases. The major purpose of this work is to report the design, synthesis and evaluation of a number of novel cell penetrating peptides (CPP) that have been developed in our group during the past few years. A series of six fluorescein-labeled cell penetrating peptides that possess one ornithine and eight arginines were synthesized. The ornithine was guanylated with a modified guanidine head group that contains an alkyl group (methyl through hexyl). Biological testing of the cell introductory peptides was performed to prove their ability to cross the cell membrane.
To conduct the quantification of cellular uptake of the modified peptides, confocal microscopy and flow cytometry were used in rat C6 glial cells. The results have shown that the fluorescently labeled peptides cross the cell membrane with an efficiency comparable to Arg9 or better. Arg9 is the CPP that has shown better results in the cellular uptake until the present work. The design and synthesis of cell-penetrating properties of fluorescently labeled oligoguanidines were also explored. However, these attempts were interrupted to focus on the synthesis of the modified peptides. The synthesis and evaluation of proline-rich peptides capable of entering cells is also reported in this work. The main characteristics of these peptides are that they contain five prolines, and two arginines that offer a cationic character to the peptide, a fact that is a crucial feature to improved cellular uptake.