title: Preclinical characterization and validation of a dual-labeled trastuzumab-based imaging agent for diagnosing breast cancer.
abstract: OBJECTIVE
  The combination of both nuclear and fluorescent reporters provides unique opportunities for noninvasive nuclear imaging with subsequent fluorescence image-guided resection and pathology. Our objective was to synthesize and optimize a dual-labeled trastuzumab-based imaging agent that can be used to validate an optical imaging agent with potential use in identifying tumor metastases in human epidermal growth factor receptor 2 (HER2) positive breast cancer patients.
  
  
  METHODS
  [(111)In]-DTPA-trastuzumab-IRDye 800 was synthesized by a three-step procedure. Purity, stability, immunoreactivity, internalization and biodistribution were explored in HER2+ SKBR-3 cells. Biodistribution of [(111)In]-DTPA-trastuzumab-IRDye 800 was performed in a SKBR-3 xenograft model.
  
  
  RESULTS
  [(111)In]-DTPA-trastuzumab-IRDye 800 demonstrated high purity by both chemical and fluorometric determinations. Both flow cytometry and the Lindmo assay demonstrated a high binding affinity of [(111)In]-DTPA-trastuzumab-IRDye 800 to HER2-overexpressing cells. The dual-labeled conjugate was stable in PBS, but not in serum after 24 h at 37 °C. Larger molecules (>150 kD) were seen after a 24 h-incubation in human serum. Biodistribution studies revealed tumor-specific accumulation of [(111)In]-DTPA-trastuzumab-IRDye 800 in SKBR-3 tumors, and tumor uptakes at 24 and 48 h were (12.42±1.72)% and (9.96±1.05)%, respectively, following intravenous administration. The tumor-to-muscle ratio was 9.13±1.68 at 24 h, and increased to 12.79±2.13 at 48 h. Liver and kidney showed marked uptake of the dual-labeled imaging agent.
  
  
  CONCLUSIONS
  [(111)In]-DTPA-trastuzumab-IRDye 800 is an effective diagnostic biomarker that can be used to validate dual-labeled, molecularly targeted imaging agents and can allow these agents to be translated into clinical practice for identifying HER2+ lesions.
authors.affiliation: 1 Department of Nuclear Medicine, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Peking University Cancer Hospital & Institute, Beijing 100142, China ; 2 Division of Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
authors.forename: Xuejuan
authors.initials: X
authors.lastname: Wang
authors.affiliation: 1 Department of Nuclear Medicine, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Peking University Cancer Hospital & Institute, Beijing 100142, China ; 2 Division of Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
authors.forename: Melissa B
authors.initials: MB
authors.lastname: Aldrich
authors.affiliation: 1 Department of Nuclear Medicine, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Peking University Cancer Hospital & Institute, Beijing 100142, China ; 2 Division of Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
authors.forename: Milton V
authors.initials: MV
authors.lastname: Marshall
authors.affiliation: 1 Department of Nuclear Medicine, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Peking University Cancer Hospital & Institute, Beijing 100142, China ; 2 Division of Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
authors.forename: Eva M
authors.initials: EM
authors.lastname: Sevick-Muraca
keywords: Dual-labeling; human epidermal growth factor receptor 2 (HER2); near-infrared optical imaging (NIR optical imaging); radioimmunoscintigraphy; trastuzumab