title: MicroPET/CT imaging of patient-derived pancreatic cancer xenografts implanted subcutaneously or orthotopically in NOD-scid mice using (64)Cu-NOTA-panitumumab F(ab')2 fragments.
abstract: BACKGROUND
  Our objective was to study microPET/CT imaging of patient-derived pancreatic cancer xenografts in NOD-scid mice using F(ab')2 fragments of the fully-human anti-EGFR monoclonal antibody, panitumumab (Vectibix) labeled with (64)Cu. More than 90% of pancreatic cancers are EGFR-positive.
  
  
  METHODS
  F(ab')2 fragments were produced by proteolytic digestion of panitumumab IgG or non-specific human IgG, purified by ultrafiltration then modified with NOTA chelators for complexing (64)Cu. Panitumumab IgG and Fab fragments were similarly labeled with (64)Cu. EGFR immunoreactivity was determined in competition and direct (saturation) cell binding assays. The biodistribution of (64)Cu-labeled panitumumab IgG, F(ab')2 and Fab was compared in non-tumor-bearing Balb/c mice. MicroPET/CT and biodistribution studies were performed in NOD-scid mice engrafted subcutaneously (s.c.) or orthotopically with patient-derived OCIP23 pancreatic tumors, or in NOD-scid with s.c. PANC-1 human pancreatic cancer xenografts.
  
  
  RESULTS
  Panitumumab F(ab')2 fragments were produced in high purity (>90%), derivitized with 3.2±0.7 NOTA/F(ab')2, and labeled with (64)Cu (0.3-3.6MBq/μg). The binding of (64)Cu-NOTA-panitumumab F(ab')2 to OCIP23 or PANC-1 cells was decreased significantly by an excess of panitumumab IgG. The Kd for binding of (64)Cu-NOTA-panitumumab F(ab')2 to EGFR on PANC-1 cells was 0.14±0.05nmol/L. F(ab')2 fragments exhibited more suitable normal tissue distribution for tumor imaging with (64)Cu than panitumumab IgG or Fab. Tumor uptake at 48h post injection (p.i.) of (64)Cu-NOTA-panitumumab F(ab')2 was 12.0±0.9% injected dose/g (ID/g) in s.c. and 11.8±0.9% ID/g in orthotopic OCIP23 tumors vs. 6.1±1.1% ID/g in s.c. PANC-1 xenografts. Tumor/Blood (T/B) ratios were 5:1 to 9:1 for OCIP23 and 2.4:1 for PANC-1 tumors. Tumor uptake of (64)Cu-NOTA-non-specific F(ab')2 in OCIP23 xenografts was 5-fold lower than (64)Cu-panitumumab F(ab')2. All tumor xenografts were clearly imaged by microPET/CT at 24 or 48h p.i. of (64)Cu-NOTA-panitumumab F(ab')2.
  
  
  CONCLUSIONS
  (64)Cu-panitumumab F(ab')2 fragments bound with high affinity to EGFR on pancreatic cancer cells in vitro and localized specifically in patient-derived pancreatic cancer xenografts in mice in vivo, allowing tumor visualization by microPET/CT at 24 or 48h p.i.
authors.affiliation: Department of Pharmaceutical Sciences, University of Toronto, Toronto, ON, Canada.
authors.forename: Amanda J
authors.initials: AJ
authors.lastname: Boyle
authors.affiliation: Ontario Cancer Institute/Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.
authors.forename: Ping-Jiang
authors.initials: PJ
authors.lastname: Cao
authors.affiliation: Ontario Cancer Institute/Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.
authors.forename: David W
authors.initials: DW
authors.lastname: Hedley
authors.affiliation: Banting and Best Department of Medical Research, Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON, Canada.
authors.forename: Sachdev S
authors.initials: SS
authors.lastname: Sidhu
authors.affiliation: Department of Chemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Mitchell A
authors.initials: MA
authors.lastname: Winnik
authors.affiliation: Department of Pharmaceutical Sciences, University of Toronto, Toronto, ON, Canada; Department of Medical Imaging, University of Toronto, Toronto, ON, Canada; Toronto General Research Institute, University Health Network, Toronto, ON, Canada. Electronic address: raymond.reilly@utoronto.ca.
authors.forename: Raymond M
authors.initials: RM
authors.lastname: Reilly
mesh_terms: D000818:Animals; D001692:Biological Transport; D002471:Cell Transformation, Neoplastic; D003301:Copper Radioisotopes; D066246:ErbB Receptors / Q000378:metabolism; D006571:Heterocyclic Compounds / Q000737:chemistry*; D006573:Heterocyclic Compounds, 1-Ring; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D016688:Mice, Inbred NOD; D010190:Pancreatic Neoplasms / Q000000981:diagnostic imaging / Q000473:pathology*; D049268:Positron-Emission Tomography / Q000379:methods*; D011829:Radiation Dosage; D014018:Tissue Distribution; D055114:X-Ray Microtomography / Q000379:methods*
chemical_list: D003301:Copper Radioisotopes; D006571:Heterocyclic Compounds; D006573:Heterocyclic Compounds, 1-Ring; D007140:Immunoglobulin Fab Fragments; C048993:1,4,7-triazacyclononane-N,N',N''-triacetic acid; D066246:ErbB Receptors
keywords: Copper-64; Epidermal growth factor receptor (EGFR); Pancreatic cancer; Panitumumab; Positron emission tomography (PET); Tumor models