title: Design and Synthesis of Near-Infrared Peptide for in Vivo Molecular Imaging of HER2.
abstract: We report the development, characterization, and validation of a peptide specific for the extracellular domain of HER2. This probe chemistry was developed for molecular imaging by using a structural model to select an optimal combination of amino acids that maximize the likelihood for unique hydrophobic and hydrophilic interactions with HER2 domain 3. The sequence KSPNPRF was identified and conjugated with either FITC or Cy5.5 via a GGGSK linker using Fmoc-mediated solid-phase synthesis to demonstrate flexibility for this chemical structure to be labeled with different fluorophores. A scrambled sequence was developed for control by altering the conformationally rigid spacer and moving both hydrophobic and hydrophilic amino acids on the C-terminus. We validated peptide specificity for HER2 in knockdown and competition experiments using human colorectal cancer cells in vitro, and measured a binding affinity of kd = 21 nM and time constant of k = 0.14 min(-1) (7.14 min). We used this peptide with either topical or intravenous administration in a preclinical model of colorectal cancer to demonstrate specific uptake in spontaneous adenomas and to show feasibility for real time in vivo imaging with near-infrared fluorescence. We used this peptide in immunofluorescence studies of human proximal colon specimens to evaluate specificity for sessile serrated and sporadic adenomas. Improved visualization can be used endoscopically to guide tissue biopsy and detect premalignant lesions that would otherwise be missed. Our peptide design for specificity to HER2 is promising for clinical translation in molecular imaging methods for early cancer detection.
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Bishnu P
authors.initials: BP
authors.lastname: Joshi
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Juan
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Asha
authors.initials: A
authors.lastname: Pant
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Xiyu
authors.initials: X
authors.lastname: Duan
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Quan
authors.initials: Q
authors.lastname: Zhou
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Rork
authors.initials: R
authors.lastname: Kuick
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Scott R
authors.initials: SR
authors.lastname: Owens
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Henry
authors.initials: H
authors.lastname: Appelman
authors.affiliation: Department of Medicine, Division of Gastroenterology, ‡Department of Biomedical Engineering, §Department of Biostatistics, ∥Department of Pathology, and ⊥Department of Mechanical Engineering, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Thomas D
authors.initials: TD
authors.lastname: Wang
mesh_terms: D000818:Animals; D002232:Carbocyanines / Q000737:chemistry / Q000378:metabolism; D045744:Cell Line, Tumor; D003106:Colon / Q000378:metabolism / Q000473:pathology; D003110:Colonic Neoplasms / Q000378:metabolism / Q000473:pathology; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D051379:Mice; D008856:Microscopy, Fluorescence; D057054:Molecular Imaging / Q000379:methods*; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis* / Q000378:metabolism; D060327:Solid-Phase Synthesis Techniques
chemical_list: D002232:Carbocyanines; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D010455:Peptides; D018719:Erb-b2 Receptor Tyrosine Kinases; C098793:CY5.5 cyanine dye; C508053:ERBB2 protein, human; C524056:Erbb2 protein, mouse