title: Imaging of HER2/neu expression in BT-474 human breast cancer xenografts in athymic mice using [(99m)Tc]-HYNIC-trastuzumab (Herceptin) Fab fragments.
abstract: OBJECTIVE
  To evaluate the ability of trastuzumab (Herceptin) Fab, labelled with (99m)Tc through introduced hydrazinenicotinamide (HYNIC) functionalities, to image HER2/neu-overexpressing human breast cancer xenografts in athymic mice.
  
  
  METHODS
  Fab fragments were produced by immobilized papain digestion of trastuzumab immunoglobulin G (IgG), followed by purification by ultrafiltration. The immunoreactivity of trastuzumab Fab was evaluated by receptor-binding assays against HER2/neu-positive SK-BR-3 human breast cancer cells. Trastuzumab Fab fragments were labelled with (99m)Tc following modification with HYNIC N-hydroxysuccinimide ester. Biodistribution and tumour imaging studies were performed in athymic mice bearing subcutaneous HER2/neu-overexpressing BT-474 human breast cancer xenografts following intravenous injection of 1.1 or 25 MBq of [(99m)Tc]-trastuzumab Fab (30 microg), respectively. The specificity of tumour uptake was assessed by comparison with that of [(99m)Tc]-labelled irrelevant anti-CD33 HuM195 Fab.
  
  
  RESULTS
  Trastuzumab Fab was pure and exhibited preserved immunoreactivity towards SK-BR-3 cells (K(d) = 1.6 x 10(-8) M). Modification with HYNIC diminished its receptor-binding affinity fourfold. [(99m)Tc]-trastuzumab Fab localized avidly and specifically in BT-474 xenografts, achieving a tumour uptake of 10.7% of the injected dose (ID) per gram and a tumour to blood (T/B) ratio of 3 : 1 at 24 h. The tumour uptake and T/B ratio for [(99m)Tc]-trastuzumab Fab were significantly higher than those for control [(99m)Tc]-HuM195 Fab (2.6% ID x g(-1) and 0.9 : 1, respectively; P<0.05). Tumours were imaged as early as 2 h post-injection of [(99m)Tc]-trastuzumab Fab, but were more clearly visualized at 6 and 24 h post-injection.
  
  
  CONCLUSIONS
  [(99m)Tc]-HYNIC-trastuzumab Fab localized specifically in HER2/neu-overexpressing human breast cancer xenografts in athymic mice, allowing imaging of the tumours within the useful lifetime of the radionuclide.
authors.affiliation: Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Ying
authors.initials: Y
authors.lastname: Tang
authors.forename: Deborah
authors.initials: D
authors.lastname: Scollard
authors.forename: Paul
authors.initials: P
authors.lastname: Chen
authors.forename: Judy
authors.initials: J
authors.lastname: Wang
authors.forename: Claire
authors.initials: C
authors.lastname: Holloway
authors.forename: Raymond M
authors.initials: RM
authors.lastname: Reilly
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents / Q000494:pharmacology; D001667:Binding, Competitive; D001943:Breast Neoplasms / Q000000981:diagnostic imaging / Q000473:pathology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D007128:Immunoglobulin Fragments / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry; D007700:Kinetics; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D015609:Organotechnetium Compounds / Q000494:pharmacology; D011859:Radiography; D019275:Radiopharmaceuticals; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis*; D013667:Technetium / Q000494:pharmacology; D013997:Time Factors; D014018:Tissue Distribution; D000068878:Trastuzumab
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D007074:Immunoglobulin G; D015609:Organotechnetium Compounds; D019275:Radiopharmaceuticals; D018719:Erb-b2 Receptor Tyrosine Kinases; D013667:Technetium; D000068878:Trastuzumab