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Author
Lucana Meneses, Celia
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Abstract
Peptides are among drugs based on small molecules and therapeutic biomolecules, allowing them to combine the best of these two worlds. Like proteins, peptides can have high affinity and selectivity and, like small molecules, have low immunogenicity and production costs, and are synthetically accessible. The main obstacle to the widespread application of peptides is the low resistance to proteases of conventional linear peptides made from proteinogenic amino acids. However, various chemical modification strategies make it possible to overcome this limitation.
In this thesis, we focused on applying cyclization to different peptides that target the transferrin receptor, with the aim of increasing their proteolytic resistance and affinity.In the first project, we aimed to develop handfuls (stapled) bicyclic peptides with a scaffold. chemical, in order to improve the transport of therapies through the blood-brain barrier, which represents the greatest challenge in the development of drugs directed to the central nervous system. We have designed a small library of cyclic peptides. Subsequently, we produce them by solid phase synthesis, solution cyclization, purification by preparative reverse phase HPLC, and characterization by UPLC-UV and MALDI-TOF. We found that cyclization improves serum half-life from 2 hours for the original peptide, up to 7 hours in some cyclic versions. Most notably, the cellular association was improved with all cyclic peptides,increasing the apparent Kd up to 30 times compared to the linear version. This increase in cellular association was independent of the stabilization of the secondary structure evaluated by circular dichroism.
In the second project, our goal was to generate cyclic peptides as independent therapeutic agents, to protect ischemic neural cells. Several peptides were rationally designed based on the alpha helix-turn-beta sheet structure of transferrin, which is involved in the interaction of this protein with its receptor. Handful and bicyclic peptides were synthesized, purified and characterized. Circular dichroism confirmed the alpha helix structure of the peptides and the relative stabilization provided by the handful. The results of a competition assay with transferrin showed that none of the candidates could compete with the endogenous protein in the studied cell line; therefore, other lines with higher levels of this receptor will be studied.
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