Orthogonally protected artificial amino acid as tripod ligand for automated peptide synthesis and labeling with [(99m)Tc(OH(2))(3)(CO)(3)](+)

Bioconjug Chem. 2013 Jan 16;24(1):26-35. doi: 10.1021/bc3003327. Epub 2012 Dec 26.

Abstract

1,2-Diamino-propionic acid (Dap) is a very strong chelator for the [(99m)Tc(CO)(3)](+) core, yielding small and hydrophilic complexes. We prepared the lysine based Dap derivative l-Lys(Dap) in which the ε-NH(2) group was replaced by the tripod through conjugation to its α-carbon. The synthetic strategy produced an orthogonally protected bifunctional chelator (BFC). The -NH(2) group of the α-amino acid portion is Fmoc- and the -NH(2) of Dap are Boc-protected. Fmoc-l-Lys(Dap(Boc)) was either conjugated to the N- and C-terminus of bombesin BBN(7-14) or integrated into the sequence using solid-phase peptide synthesis (SPPS). We also replaced the native lysine in a cyclic RGD peptide with l-Lys(Dap). For all peptides, quantitative labeling with the [(99m)Tc(CO)(3)](+) core at a 10 μM concentration in PBS buffer (pH = 7.4) was achieved. For comparison, the rhenium homologues were prepared from [Re(OH(2))(3)(CO)(3)](+) and Lys(Dap)-BBN(7-14) or cyclo-(RGDyK(Dap)), respectively. Determination of integrin receptor binding showed low to medium nanomolar affinities for various receptor subtypes. The IC(50) of cyclo-(RGDyK(Dap[Re(CO)(3)])) for α(v)β(3) is 7.1 nM as compared to 3.1 nM for nonligated RGD derivative. Biodistribution studies in M21 melanoma bearing nude mice showed reasonable α(v)β(3)-integrin specific tumor uptake. Altogether, orthogonally protected l-Lys(Dap) represents a highly versatile building block for integration in any peptide sequence. Lys(Dap)-precursors allow high-yield (99m)Tc-labeling with [(99m)Tc(OH(2))(3)(CO)(3)](+), forming small and hydrophilic complexes, which in turn leads to peptide radiopharmaceuticals with excellent in vivo characteristics.

MeSH terms

  • Animals
  • Bombesin / analogs & derivatives*
  • Bombesin / chemical synthesis
  • Bombesin / chemistry
  • Cell Line, Tumor
  • Chelating Agents / chemical synthesis
  • Chelating Agents / chemistry
  • Humans
  • Integrin alphaVbeta3 / metabolism
  • Lysine / analogs & derivatives*
  • Lysine / chemical synthesis
  • Melanoma / diagnosis*
  • Melanoma / metabolism
  • Mice
  • Mice, Nude
  • Oligopeptides / chemical synthesis
  • Oligopeptides / chemistry*
  • Organotechnetium Compounds / chemical synthesis
  • Organotechnetium Compounds / chemistry*
  • Peptide Fragments / chemical synthesis
  • Peptide Fragments / chemistry*
  • Propionates / chemical synthesis
  • Propionates / chemistry*
  • Solid-Phase Synthesis Techniques / methods*

Substances

  • Chelating Agents
  • Integrin alphaVbeta3
  • Oligopeptides
  • Organotechnetium Compounds
  • Peptide Fragments
  • Propionates
  • bombesin (7-14)
  • arginyl-glycyl-aspartic acid
  • Lysine
  • Bombesin