Incorporation of ortho-Carbaboranyl-N_ε-Modified l-Lysine into Neuropeptide Y Receptor Y₁- and Y₂-Selective Analogues

Abstract

Nontoxic ortho-carbaborane is one of the most promising structure for boron neutron capture therapy (BNCT). For directed uptake of ortho-carbaborane by tumor cells, receptor-subtype selective neuropeptide Y (NPY) and its derivatives were modified with ortho-carbaborane. The derivative [F(7), P(34)]-NPY has been shown to be a breast cancer selective ligand that binds to the Y(1)-receptor subtype, whereas [Ahx(5-24)]-NPY selectively addresses Y(2)-receptor subtypes that are found in neuroblastoma cells. ortho-Carbaboranyl propionic acid was synthesized and linked to the ε-amino group of N(α)-Fmoc protected L-lysine. The characterization of the compounds was performed by NMR, IR, and MS studies. The carbaborane-modified amino acid was incorporated into NPY, [F(7), P(34)]-NPY, and [Ahx(5-24)]-NPY by an optimized solid phase peptide synthesis using Fmoc protection. Binding studies and IP accumulation assays confirmed nanomolar affinity and activity of the modified analogues despite of the large carbaborane cluster. Internalization studies revealed excellent and receptor subtype specific uptake of the conjugates into respective cells.

Quantitative impedimetric NPY-receptor activation monitoring and signal pathway profiling in living cells

“…The statistical analysis of three to four independent experiments revealed an averaged EC 50 value of 1.73 nM for NPY and 2.30 nM for FP-NPY determined by impedimetric monitoring in comparison to 4.14 nM for NPY determined by IP accumulation assay. These values are in line with previously reported EC 50 values in a COS-7 IP-accumulation assay (Ahrens et al, 2011). This underlines the sensitivity and comparability of our impedance spectroscopy based NPY-activation monitoring to the established radio ligand based IP-accumulation assay.…”

“…The peptide ligand pNPY (abbreviated as NPY), the Y1-receptor preferring [F7,P34]-NPY (abbreviated as FP-NPY) (Soll et al, 2001), the Y2-receptor preferring [Ahx5-24]-NPY (abbreviated as Ahx-NPY) (Rist et al, 1996) and the Y5-receptor preferring [Ala31, Aib32]-NPY (abbreviated as AAib-NPY) (Cabrele et al, 2000) were synthesized by solid phase peptide synthesis as described previously (Ahrens et al, 2011). For selective Y1R inhibition BIBP3226 (SantaCruz Biotechnology, USA) was used (Rudolf et al, 1994).…”

Automated solid-phase peptide synthesis to obtain therapeutic peptides

“…With respect to the great potential of NPY ligands as pharmaceuticals for various indications such as cancer [133–134] and obesity [99], automated SPPS also offers a versatile repertoire. As an example, a method to produce radiolabeled Y 1 -receptor preferring agonists of NPY has been described, which could selectively target breast cancer cells and allowed specific tumor diagnosis.…”

Charge‐Compensated Metallacarborane Building Blocks for Conjugation with Peptides

“…In quantitative studies, the content of boron in the cancerc ells was found to exceed the significant amount of 10 9 boron atoms per cell. [10] Besides NPY,s everal other peptides have been proposed ast umor-targeting carriers:e xamples include somatostatin, epidermal growth factor,n eurotensin, substance P, gastrin-releasing peptide, insulin-like growth factor, [11] amelanocyte-stimulating hormone, cholecystokinin, vasoactive intestinal peptide, and bombesin. [12] In view of this large number of peptides,i ta ppears surprising that the cobalt bis(dicarbollide) anion 1 has never been incorporated into any tumor-selective peptided espite obvious possible advantages.…”

“…[14] To avoid theses ide effects, we designed an ew chargecompensated (zwitterionic) key intermediate of 1,w hich allows furtherm odifications with moieties of biochemical relevance, such as monosaccharides. The resulting buildingb locks were successfully coupled to the tumor-selective peptide [ F 7 ,P 34 ]-NPY [10] with retention of excellent Y 1 receptor activity.…”