Novel dimeric DOTA-coupled peptidic Y1-receptor antagonists for targeting of neuropeptide Y receptor-expressing cancers

ABSTRACT

Understanding GPCR signaling in the brain- the path to CNS drug discovery.

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Contribution à l étude des relations structure-activité de l urotensine II humaine (UIIh) et de l urotensin II-related peptide (URP) : études pharmacologiques ex vivo et in vitro

L urotensine II (UII) et l UII-related peptide (URP) sont des peptides cycliques vasoconstricteurs, ligands endogènes du GPR14. Dans ce travail, nous avons montré que l immunoréactivité de type UII localisée dans les motoneurones spinaux humains correspond à un peptide mature de 11 aa, H-Glu-Thr-Pro-Asp-Cys-Phe-Trp-Lys-Tyr-Cys-Val-OH. Le blocage de l extrémité N-terminale et l amidation C-terminale n altèrent pas l activité contractile de l UIIh sur des anneaux d aorte de rat. Les Alascan et D-scan de l UIIh-NH2 ainsi que la synthèse de peptides tronqués révèlent que l octapeptide C-terminal UIIh(4-11) est le fragment minimal isoactif. La présence d une chaîne latérale fonctionnalisée sur le résidu N-terminal de l UIIh(4-11) n est pas requise pour l activité du peptide. En revanche, l intégrité et l orientation du pont disulfure sont des paramètres essentiels pour l activité biologique de l UIIh. La iodation de la Tyr6 de l UIIh(4-11) potentialise l effet vasoconstricteur du peptide. Des modifications en position 3 et 5, [Cha3]UIIh(4-11) et [4-amino-Phe5]UIIh(4-11), conduisent à des molécules aux propriétés antagonistes. Les résidus intracycliques de l URP, qui présentent une conformation unique caractérisée par un coude -inversé, sont directement impliqués dans la liaison et l activation du récepteur. Enfin, les analogues [D-Trp4]URP, [D-Tyr6]URP et [Orn5]URP se comportent comme des antagonistes du GPR14. L ensemble de ces travaux démontre que 1) l UIIh(4-11) est la séquence minimale active de l UIIh sur le GPR14, 2) la iodation du résidu Tyr6 de l UIIh(4-11) conduit à un agoniste 5 fois plus puissant que le peptide natif, 3) le remplacement des résidus Phe3 et Lys5 respectivement par un résidu cyclohexyl-Ala ou 4-amino-Phe, de même que l inversion de configuration du C du Trp4 ou de la Tyr6 de l URP constituent des voies d accès à des antagonistes du GPR14 qui pourraient contribuer à la conception de nouvelles molécules anti-hypertensives à visée thérapeutique.Urotensin II (UII) and UII-related peptide are the endogenous ligands of a G-protein coupled receptor named GPR14. In this work, we have shown, in human, that UII-immunoreactivity is located in human spinal motoneurons and that the UII precursor is processed to generate a mature peptide of 11 residues, H-Glu-Thr-Pro-Asp-Cys-Phe-Trp-Lys-Tyr-Cys-Val-OH. N-blocked and C-terminal amidated analogues exhibited the same vasocontractile activity as hUII. Alascan or D-scan of hUII-NH2 and the synthesis of truncated fragments indicated that the C-terminal octapeptide retains full biological activity. A functionalized side-chain is not required on the first residue of hUII(4-11) to maintain full activity. Conversely, the disulphide bridge and its orientation play a crucial role in the biological activity of hUII. Monoiodination of the Tyr6 residue of hUII(4-11) enhances the contractile potency of the peptide. Modification at position 3 and 5 generated compounds, [Cha3]hUII(4-11) and [4-amino-Phe5]hUII(4-11), able to inhibit the hUII-evoked contraction on rat aortic rings. Intracyclic residues of URP, which exhibit a single conformation characterized by an inverse -turn, play a crucial role in the biological activity of the peptide. Finally, [D-Trp4]URP, [Orn5]URP and [D-Tyr6]URP behave as antagonists of GPR14. In conclusion, structure-activity relationship studies have shown that 1) the C-terminal octapeptide hUII(4-11) is the minimal core sequence, 2) monoiodination of the Tyr6 residue of hUII(4-11) yielded to an agonist that was 5 times more potent than the native peptide, and 3) substitution of the Phe3 or the Lys5 residues with 4-amino-Phe or cyclohexyl-Ala, respectively, as well as substitution of the Trp4 and the Tyr6 residues of URP by their D-isomer open new ways for the synthesis of GPR14 antagonists which may contribute to the design of novel ligands of GPR14 with therapeutic value as anti-hypertensive drugs.ROUEN-BU Sciences (764512102) / SudocROUEN-BU Sciences Madrillet (765752101) / SudocSudocFranceF

Discovery of New Allosteric Modulators of the Urotensinergic System through Substitution of the Urotensin II-Related Peptide (URP) Phenylalanine Residue

International audienceUrotensin II (UII) and urotensin II-related peptide (URP) are functionally selective, suggesting that these two hormones might play distinct physiological role through different interactions with their cognate receptor UT. Hypothesizing that the Phe3 residue of URP, which is also present in UII, is a key-element of its specific UT activation, we evaluated the impact of its replacement by non-natural amino acids in URP. Each compound was evaluated for its ability to bind UT, induce rat aortic ring contraction, and activate Gq, G12, and β-arrestin 1 signaling pathways. Such modifications impaired contractile efficacy, reflected by a reduced aptitude to activate G12 in URP but not in the truncated but equipotent UII4-11. Moreover, we have identified two structurally different UT modulators: [d-Phe(pI)3]URP and [Bip3]URP, which exert a probe-dependent action against UII and URP. These compounds should help us understand the specific roles of these hormones as well as guide further therapeutic development

Synthesis of P-chiral enephosphonic acid derivatives

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Identification of an unusual cleavage site for the prolyl endopeptidase: investigation of the breakdown of the octadecaneuropeptide ODN. Peptides 2004, Proceedings of the 3rd International and 28th European Peptide Symposium, pp 890-891

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Receptor-independent cellular uptake of pituitary adenylate cyclase-activating polypeptide.

International audiencePituitary adenylate cyclase-activating polypeptide (PACAP), a hypophysiotropic neurohormone, participates in the regulation of pleiotropic functions. The recent discovery of intracellular PACAP receptors in the brain and the testis as well as the physico-chemical characteristics of PACAP, i.e. extended α-helix containing basic residues, prompted us to evaluate the propensity of PACAP to cross the plasma membrane in a receptor-independent manner. Using confocal microscopy and flow cytometry, we demonstrated the ability of FITC-conjugated PACAP to efficiently penetrate into the internal cell compartment by direct translocation and endocytosis through clathrin-coated pits and macropinocytosis. Our study also revealed that, once inside the cells, PACAP38 is not entirely degraded by intracellular enzymes and that a significant amount of intact PACAP38 is also able to exit cells. Moreover, using binding assay on rat nuclear fractions from various tissues, PACAP nuclear receptors were identified. We also found that PACAP stimulates calcium release in rat testis nuclei. Interestingly, PACAP27 and PACAP38 but not VIP were able to upregulate de novo DNA synthesis in testis nuclei and that this effect was abolished by PACAP(6-38). These results support the presence of PAC1 receptors at the nuclear membrane and raise questions about their role in the biological activity of the peptide. These findings contribute to the characterization of PACAP as an intracrine factor and suggest that these intracellular PAC1 binding sites, probably associated with specific biological activities, should be taken into account during the development of PACAP-based drugs

Carbon-Supported PtNi Nanocrystals for Alkaline Oxygen Reduction and Evolution Reactions: Electrochemical Activity and Durability upon Accelerated Stress Tests

International audiencePtNi is amongst the most active electrocatalyst for the oxygen reduction reaction, but its stability in operation is uncertain. Intuitively, alkaline environments lead to milder degradations than acidic ones, although carbon-supported Pt-group metal nanoparticles are particularly degraded even in dilute alkaline electrolytes. To date, PtNi catalysts durability has not been characterized for alkaline oxygen reduction and evolution reactions (ORR and OER). Herein, carbon-supported shape controlled PtNi catalysts were compared in terms of activity and durability during alkaline ORR and OER. The PtNi catalysts are shape-controlled Pt-rich alloy, Ni-rich alloy, and Pt core/Ni shell (Pt@Ni) synthesized on Vulcan XC72R carbon. Their morphology and composition were evaluated by identical-location transmission electron microscopy, X-ray photoelectron spectroscopy and X-ray diffraction pre and post accelerated stress test. Compared to Pt/C and Ni/C benchmark catalysts, the core-shell and Ni-rich alloy catalysts gave high and stable OER activities. After accelerated stress test, the catalysts show two features which are believed to play a major role in the durability: a Ni-enrichment at the nanoparticles' surface and an improved attachment of the catalyst to the carbon support