title: Axl-targeted cancer imaging with humanized antibody h173.
abstract: OBJECTIVE
  The tyrosine kinase receptor Axl is overexpressed in various types of cancer and correlated with cancer malignancy. Selective Axl blockade reduces tumor growth and metastasis. The purpose of this study was to examine whether the humanized anti-Axl antibody humanized 173 (h173) labeled with near-infrared fluorescence (NIRF) dye Cy5.5 could be applied as a molecular imaging probe for NIRF imaging of Axl expression in tumor models.
  
  
  METHODS
  NIRF dye Cy5.5 was conjugated to h173 or human normal immunoglobulin G (hIgG) control through amino groups. The resulting probes were evaluated in both A549 (Axl positive) and NCI-H249 (Axl negative) lung cancer xenografts through in vivo NIRF imaging. Ex vivo imaging and probe distribution assay were also carried out to confirm the in vivo imaging results.
  
  
  RESULTS
  After conjugation, binding activity of h173-Cy5.5 was determined to be 97.75 % ± 2.09 % of the unmodified h173. In vitro fluorescence-activated cell sorting (FACS) and fluorescence microscopy analysis validated the specific binding of h173 toward Axl-positive A549 cells. h173-Cy5.5 was then applied to image Axl expression in vivo. In A549 (Axl positive) cancer xenografts, the tumor uptake of h173-Cy5.5 was significantly higher than that of the hIgG-Cy5.5 control (P < 0.05) at late time points (1, 2, 3, 4, and 7 days). On the contrary, in NCI-H249 (Axl negative) cancer xenografts, the tumor uptake of both hIgG-Cy5.5 and h173-Cy5.5 was low and showed no significant difference (P > 0.05) at all time points examined. Ex vivo imaging and immunofluorescence staining analysis further validated the in vivo imaging results.
  
  
  CONCLUSIONS
  Collectively, all in vitro, in vivo, and ex vivo data suggested that h173-Cy5.5 could serve as a valid probe for Axl-targeted cancer imaging, which could therefore aid in tumor diagnosis, prognosis, and treatment monitoring.
authors.affiliation: Molecular Imaging Center, Department of Radiology, University of Southern California, 2250 Alcazar St. CSC103, Los Angeles, CA, 90033, USA.
authors.forename: Dan
authors.initials: D
authors.lastname: Li
authors.forename: Shuanglong
authors.initials: S
authors.lastname: Liu
authors.forename: Ren
authors.initials: R
authors.lastname: Liu
authors.forename: Ryan
authors.initials: R
authors.lastname: Park
authors.forename: Haiyang
authors.initials: H
authors.lastname: Yu
authors.forename: Valery
authors.initials: V
authors.lastname: Krasnoperov
authors.forename: Parkash S
authors.initials: PS
authors.lastname: Gill
authors.forename: Zibo
authors.initials: Z
authors.lastname: Li
authors.forename: Hong
authors.initials: H
authors.lastname: Shan
authors.forename: Peter S
authors.initials: PS
authors.lastname: Conti
mesh_terms: D000818:Animals; D061067:Antibodies, Monoclonal, Humanized; D002232:Carbocyanines / Q000378:metabolism; D045744:Cell Line, Tumor; D003952:Diagnostic Imaging; D005452:Fluoresceins / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D051379:Mice; D009369:Neoplasms / Q000175:diagnosis* / Q000201:enzymology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D019265:Spectroscopy, Near-Infrared; D014018:Tissue Distribution; D000094963:Axl Receptor Tyrosine Kinase
chemical_list: D061067:Antibodies, Monoclonal, Humanized; C098793:CY5.5 cyanine dye; D002232:Carbocyanines; D005452:Fluoresceins; D007074:Immunoglobulin G; D011518:Proto-Oncogene Proteins; C024098:6-carboxyfluorescein; D020794:Receptor Protein-Tyrosine Kinases; D000094963:Axl Receptor Tyrosine Kinase