Synthesis and evaluation of technetium-99m-labeled bioreductive pharmacophores conjugated with amino acids and peptides for tumor imaging

Development of molecular imaging agents to target tumor has become a major trend in nuclear medicine. With the aim to develop new potential ^99mTc-radiopharmaceuticals for targeting tumor, we have synthesized 5-nitroimidazolyl amino acids and RGD-coupled 2-nitroimidazoles. Technetium-99m radiolabeling with high radiochemical purity (>90%) was achieved for all the compounds. The radiolabeled complexes exhibited substantial in vitro stability in saline, serum, and histidine solution (10^-2 m). Cell binding studies in EAC and B16F10 cell lines also revealed rapid and comparatively high cellular internalization. Among all the compounds studied, the binding of ^99mTc(CO)₃-5 to B16F10 cells was moderately inhibited by the competitive peptide c[RGDfV], suggesting specificity of the radioligand toward α_vβ₃ receptor. However, no significant displacement of bound radioligand was observed when the binding of the ^99mTc-labeled complexes to above cells was challenged with excess competitive peptide. Fluorescent microscopy study provided direct evidence of intracellular localization of 5(6)-carboxyfluorescein-labeled 2-nitroimidazolyl-RGD-peptide in α_vβ₃-positive B16F10 mouse melanoma cell line. The ligands caused only 8-13% of hemolysis toward rat erythrocytes at concentrations as high as 100 μm. Imaging and biodistribution studies were performed in Swiss albino mice bearing induced tumor. ^99mTc-1 and ^99mTc(CO)₃-5 demonstrated a very favorable in vivo profile. Selective uptake and retention in tumor with encouraging tumor/muscle and tumor/blood ratio and significant cellular uptake of fluorescence-labeled-2-nitroimidazolyl RGD indicate the great potentiality of the pharmacophore for further evaluation as potential molecular imaging agent in cancer diagnosis.