title: Overexpressed Claudin-1 Can Be Visualized Endoscopically in Colonic Adenomas In Vivo.
abstract: OBJECTIVE
  Conventional white-light colonoscopy aims to reduce the incidence and mortality of colorectal cancer (CRC). CRC has been found to arise from missed polypoid and flat precancerous lesions. We aimed to establish proof-of-concept for real-time endoscopic imaging of colonic adenomas using a near-infrared peptide that is specific for claudin-1.
  
  
  METHODS
  We used gene expression profiles to identify claudin-1 as a promising early CRC target, and performed phage display against the extracellular loop of claudin-1 (amino acids 53-80) to identify the peptide RTSPSSR. With a Cy5.5 label, we characterized binding parameters and showed specific binding to human CRC cells. We collected in vivo near-infrared fluorescence images endoscopically in the CPC;Apc mouse, which develops colonic adenomas spontaneously. With immunofluorescence, we validated specific peptide binding to adenomas from the proximal human colon.
  
  
  RESULTS
  We found a 2.5-fold increase in gene expression for claudin-1 in human colonic adenomas compared with normal. We showed specific binding of RTSPSSR to claudin-1 in knockdown and competition studies, and measured an affinity of 42 nmol/L and a time constant of 1.2 minutes to SW620 cells. In the mouse, we found a significantly higher target-to-background ratio for both polypoid and flat adenomas compared with normal by in vivo images. On immunofluorescence, we found significantly greater intensity for human adenomas (mean ± SD, 25.5 ± 14.0) vs normal (mean ± SD, 9.1 ± 6.0) and hyperplastic polyps (mean ± SD, 3.1 ± 3.7;P= 10-5and 8 × 10-12, respectively), and for sessile serrated adenomas (mean ± SD, 20.1 ± 13.3) vs normal and hyperplastic polyps (P= .02 and 3 × 10-7, respectively).
  
  
  CONCLUSIONS
  Claudin-1 is overexpressed in premalignant colonic lesions, and can be detected endoscopically in vivo with a near-infrared, labeled peptide.
authors.affiliation: Department of Medicine, Division of Gastroenterology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Emily F
authors.initials: EF
authors.lastname: Rabinsky
authors.affiliation: Department of Medicine, Division of Gastroenterology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Bishnu P
authors.initials: BP
authors.lastname: Joshi
authors.affiliation: Department of Medicine, Division of Gastroenterology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Asha
authors.initials: A
authors.lastname: Pant
authors.affiliation: Department of Medicine, Division of Gastroenterology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Juan
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan.
authors.forename: Xiyu
authors.initials: X
authors.lastname: Duan
authors.affiliation: Department of Medicine, Division of Gastroenterology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Arlene
authors.initials: A
authors.lastname: Smith
authors.affiliation: Department of Biostatistics, University of Michigan, Ann Arbor, Michigan.
authors.forename: Rork
authors.initials: R
authors.lastname: Kuick
authors.affiliation: Department of Pathology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Shuling
authors.initials: S
authors.lastname: Fan
authors.affiliation: Department of Pathology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Asma
authors.initials: A
authors.lastname: Nusrat
authors.affiliation: Department of Pathology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Scott R
authors.initials: SR
authors.lastname: Owens
authors.affiliation: Department of Pathology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Henry D
authors.initials: HD
authors.lastname: Appelman
authors.affiliation: Department of Medicine, Division of Gastroenterology, University of Michigan, Ann Arbor, Michigan; Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan; Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan.
authors.forename: Thomas D
authors.initials: TD
authors.lastname: Wang
keywords: APC, adenomatous polyposis coli; BSA, bovine serum albumin; CLDN1, claudin-1; CRC, colorectal cancer; Colon Cancer; DAPI, 4′,6-diamidino-2-phenylindole; Early Detection; HRP, horseradish peroxidase; IF, immunofluorescence; IHC, immunohistochemistry; Molecular Imaging; PBS, phosphate-buffered saline; PBST, phosphate-buffered saline plus 0.1% Tween-20; PFA, paraformaldehyde; RT, room temperature; SSA, sessile serrated adenoma; T/B, target-to-background; TEER, transepithelial electrical resistance; TFA, trifluoroacetic acid; ZO-1, zonula occludens-1; siCL, control small interfering RNA; siRNA, small interfering RNA