Evaluation of an 111In-Radiolabeled Peptide as a Targeting and Imaging Agent for ErbB-2 Receptor–Expressing Breast Carcinomas

Abstract

Purpose: The cellular targeting and tumor imaging properties of a novel ErbB-2-avid peptide, discovered from bacteriophage display, were evaluated in human breast carcinoma cells and in breast carcinoma^xenografted mice. Experimental Design: The affinity of the ErbB-2 targeting peptide KCCYSL and its alanine substituted counterparts for the extracellular domain (ECD) of purified recombinant ErbB-2 (ErbB-2-ECD) was assessed by fluorescence titration. Binding of the KCCYSL peptide to breast and prostate carcinoma cells was analyzed by confocal microscopy. A DOTA(GSG)-KCCYSL peptide conjugate was radiolabeled with 111 In, and stability, target binding, and internalization were analyzed in vitro. In vivo biodistribution and single-photon emission computed tomography imaging studies were done with the radiolabeled peptide in MDA-MB-435 human breast tumorb earing severe combined immunodeficient mice. Results: KCCYSL peptide exhibited high affinity (295 F 56 nmol/L) to ErbB-2-ECD. Substitution of alanine for lysine, tryptophan, and cysteine reduced the peptide affinity f 1-to 2.4-fold, whereas replacing leucine completely abolished binding. Both biotin- KCCYSL and 111 In-DOTA(GSG)-KCCYSL were capable of binding ErbB-2^expressing human breast carcinoma cells in vitro. Approximately 11% of the total bound radioactivity was internalized in the carcinoma cells. Competitive binding studies indicated that the radiolabeled peptide exhibited an IC 50 value of 42.5 F 2.76 nmol/L for the breast carcinoma cells.

111In-Labeled Galectin-3–Targeting Peptide as a SPECT Agent for Imaging Breast Tumors

“…Few other bacteriophage-selected peptides that bind tumor-associated antigens have been reported to function as efficacious tumor imaging agents in vivo. Recently, we successfully used an ErbB-2 receptor-targeting peptide (KCCYSL), derived from a bacteriophage display library, for SPECT/CT of human xenografted breast tumors in mice (18). Here, we demonstrated that 111 In-DOTA(GSG)-G3-C12 can be used to image galectin-3-expressing MDA-MB-435 human breast cancer xenografts in SCID mice.…”

“…The 111 In-labeling of DOTA(GSG)-ANTPCGPYTHDCPVKR (G3-C12) or DOTA(GSG)-PPPKGDVHTNRTYACC (scrambled version of G3-C12) was performed essentially as previously described (18). Briefly, the peptides were radiolabeled with 111 In by incubating 25 mg of peptide in 5 mL of 111 InCl 3 (185 MBq/500 mL) (Mallinckrodt) and 80 mL of ammonium acetate, pH 5.5, at 85°C for 60 min, after which 10 mL of 2 mM ethylenediamine tetraacetic acid were added to complex unreacted 111 In.…”

Phage display and molecular imaging: expanding fields of vision in living subjects

“…After production by solid phase synthesis, electrospray ionization mass spectrometry analysis found that the peptide exists as a mixture of oxidized and reduced monomer in solution without measurable amounts of a dimeric form. The binding affinity to recombinant ErbB2-ECD, as determined by fluorescence titration experiments in solution, was reported to be K D ~300 nM (Kumar et al 2007). A DOTA chelating group was added to the N-terminus of the peptide through a Gly-Ser-Gly linker and labeled with 111 In.…”

Diffuse optical tomography of the breast: preliminary findings of a new prototype and comparison with magnetic resonance imaging

“…A molecular imaging approach using certain fluorescent probes may vastly improve signal-to-noise ratios by signal amplification [27]. Target-specific fluorescent probes for optical imaging have already been used successfully in animal experiments [28][29][30].…”