Carbaboranylation of Truncated C‐Terminal Neuropeptide Y Analogue Leads to Full hY₁ Receptor Agonism

Abstract

The human Y receptor is overexpressed in breast tumour cells and is, therefore, a valuable target for site-selective drug delivery. The well-established hY R-selective ligand [Phe7,Pro34]NPY has been used to couple to drugs but its length of 36 amino acids also implies complex synthesis and high production costs. Therefore, shorter ligands are desirable. However, truncated versions of neuropeptide Y (NPY) usually result in reduced binding, antagonists, or only partial G-protein-biased agonists. Herein, we report on a nonamer peptide derived from the C terminus of NPY that is modified by a carbaborane, which is able to activate hY R fully in terms of G-protein activation but also arrestin recruitment and internalisation. We provide evidence that this unique behaviour is due to the bulky nature of the carbaborane cluster, which might address a specific second binding pocket in hY R that is crucial for arrestin recruitment. Thus, this study helps in deciphering ligand-induced onset of different pathways in hY R mediated signalling.

Identification of a Suitable Peptidic Molecular Platform for the Development of NPY(Y1)R‐Specific Imaging Agents

“…Over the last two decades, nearly 80 truncated peptides analogues of NPY were developed, exhibiting a linear, cyclic or dimeric peptide backbone in order to identify derivatives with high NPY(Y 1 )R targeting ability. [12,14,15,17,18,19,20,21,29,30] Among these, several showed low affinities, NPY(Y 1 ) receptor subtype selectivities or stabilities. Of those remaining, we chose to directly compare five different peptide analogues regarding their metabolic stability (Figure 1), representing the most promising agents in terms of in vitro properties (NPY(Y 1 )R affinity, Table 1, and receptor subtype selectivity) within their respective group of linear, cyclic and dimeric truncated NPY analogues: i) linear [Pro 30 ,Lys(DOTA) 31 DOTA = (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrayl)tetraacetic acid), serving as reference compound with known low metabolic stability but high NPY(Y 1 )R affinity and receptor subtype selectivity, [18] ii) [Pro 30 ,Lys(DOTA) 31 ,Bip 32 ,Leu 34 ]NPY [28][29][30][31][32][33][34][35][36] (2), a variant of 1 comprising an artificial Bip (biphenylalanine) amino acid in position 32 instead of tyrosine, [12] and Lys(DOTA) in position 31 for radiolabeling, [15] iii) [Lys(lauroyl) 27 ,Pro 30 ,Lys (DOTA) 31 ,Bip 32 ,Leu 34 ]NPY [27][28][29][30][31][32][33][34]…”

“…[12,18] Alternatively, the chelator can, for example, be conjugated to a side-chain functionality of an additionally introduced amino acid at the N terminus. [19] However, as it was shown for BVD 15 that the DOTA conjugation least impedes peptide-receptor interaction when introduced in position 31 by using Lys(N ɛ -DOTA), [18] we applied this molecular design, resulting in peptide precursors 2, 3 and 4. Derivatives 1 and 5 were already described as DOTA-modified agents.…”

“…[12] This low stability was attributed to the reduced number of amino acids resulting in a poorly defined secondary structure of these short peptides entailing a faster proteolytic degradation. [16] Nevertheless, many truncated NPY analogues -several of them exhibiting very good to excellent in vitro properties such as high NPY(Y 1 )R affinity and receptor subtype selectivity [12,14,15,17,18,19,20,21] -were developed with the aim of finding a peptide sequence applicable as NPY(Y 1 )R-specific tumor-targeting agent, for instance for molecular imaging. However, respective radiolabeled analogues applied for in vivo tumor visualization using highly sensitive in vivo PET imaging were seldomly reported and showed suboptimal pharmacokinetics [15,22] caused by the stability issues discussed above.…”

“…Thus, we synthesized and directly compared 68 Ga-radiolabeled analogues of the most promising described truncated NPY analogues [12,15,17,18,19,20] with regard to their metabolic stability. This comparison was based on in vitro stability determinations in human serum as well as human liver microsomal stability assays, both providing a good, quantifiable measure on the intrinsic proteolytic degradation of the peptidic ligands.…”

New Method for Synthesis of Substituted N-Borylated Dipeptides Based on Acetonitrile Derivative of the closo-Dodecaborate Anion

“…The effectiveness of drugs for 10 B-NCT depends on the selectivity of the accumulation of boron compounds in tumor cells, which requires the introduction of vector groups into the cluster cage. Amino acids and peptides are important transport groups due to their ability to bind selectively to various receptors, mainly expressed by tumor cells [7][8][9][10][11].…”