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Novel stable analogues of the neurotensin C-terminal hexapeptide containing unnatural amino acids

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Abstract

Neurotensin (NT) (pGlu–Leu–Tyr–Glu–Asn–Lys–Pro–Arg–Arg–Pro–Tyr–Ile–Leu) exerts a dual function as a neurotransmitter/neuromodulator in the central nervous system and as a hormone/cellular mediator in periphery. This dual function of NT establishes a connection between brain and peripheral tissues that renders this peptide a central player in energy homeostasis. Many biological actions of NT are mediated through its interaction with three types of NT receptors (NTS receptors). Despite its role in energy homeostasis, NT has a short half-life that hampers further determination of the biological actions of this peptide and its receptors in brain and periphery. The short half-life of NT is due to the proteolytic degradation of its C-terminal side by several endopeptidases. Therefore, it is important to synthesize NT analogues with resistant bonds against metabolic deactivation. Based on these findings, we herein report the synthesis of ten linear, two cyclic and two dimeric analogues of NT with modifications in its structure that improve their metabolic stability, while retaining the ability to bind to NTS receptors. Modifications at position 11 (introduction of d-Tyrosine (OEthyl) [d-Tyr(Et)] or d-1-naphtylalanine [d-1-Nal] were combined with introduction of a l-Lysine or a d-Arginine at positions 8 or 9, and 1-[2-(aminophenyl)-2-oxoethyl]-1H-pyrrole-2-carboxylic acid (AOPC) at positions 7 or 8, resulting in compounds NT4-NT21. AOPC is an unnatural amino acid with promise in applications as a building block for the synthesis of peptidomimetic compounds. To biologically evaluate these analogues, we determined their plasma stability and their binding affinities to type 1 NT receptor (NTS1), endogenously expressed in HT-29 cells, Among the fourteen NT analogues, compounds, NT5, NT6, and NT8, which have d-Tyr(Et) at position 11, bound to NTS1 in a dose–response manner and with relatively high affinity but still lower than that of the natural peptide. Despite their lower binding affinities compared to NT, the NT5, NT6, and NT8 exhibited a remarkably higher stability, as a result of their chemistry, which provides protection from enzymatic activity. These results will set the basis for the rational design of novel NT molecules with improved pharmacological properties and enhanced enzymatic stability.

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Acknowledgements

Funding for this work was provided by UPAT’s “C. Carathéodory” Research Grant (to V. Magafa_2010_D.156) of the University of Patras and by an UoC’s ELKE grant (to G. Liapakis).

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  1. Vassiliki Magafa and Minos-Timotheos Matsoukas contributed equally to this work.

Authors and Affiliations

  1. Department of Pharmacy, University of Patras, 26504, Rion, Greece

    Vassiliki Magafa, Minos-Timotheos Matsoukas, Revekka Exarchakou & Georgios A. Spyroulias

  2. Department of Pharmacology, Faculty of Medicine, University of Crete, 71003, Heraklion, Crete, Greece

    Vlasios Karageorgos, Michail Pardalos & George Liapakis

  3. Department of Clinical Chemistry, School of Medicine, University of Crete, 71003, Heraklion, Crete, Greece

    Eirini Dermitzaki & Andrew N. Margioris

  4. Department of Chemistry, University of Ioannina, 45110, Ioannina, Greece

    Evgenios Κ. Stylos, George Varvounis & Andreas G. Tzakos

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  1. Vassiliki Magafa
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  2. Minos-Timotheos Matsoukas
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Correspondence to Vassiliki Magafa or George Liapakis.

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Magafa, V., Matsoukas, MT., Karageorgos, V. et al. Novel stable analogues of the neurotensin C-terminal hexapeptide containing unnatural amino acids. Amino Acids 51, 1009–1022 (2019). https://doi.org/10.1007/s00726-019-02741-2

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