title: Characterization of 99mTc-labeled cytokine ligands for inflammation imaging via TNF and IL-1 pathways.
abstract: BACKGROUND
  TNFR2-Fc and IL-1ra-Fc are recombinant cytokine ligands that target TNF and IL-1. TNFR2-Fc-IL-1ra, a dual-domain agent that incorporates both ligands, allows bifunctional binding of IL-1 receptors and TNF. This study was designed to characterize (99m)Tc-labeled forms of these ligands, (99m)Tc-IL-1ra-Fc (IF), (99m)Tc-TNFR2-Fc (TF), and (99m)Tc-TNFR2-Fc-IL-1ra (TFI), for inflammation imaging.
  
  
  METHODS
  The cytokine ligands were labeled with (99m)Tc by a direct approach via 2-iminothiolane (2-IT) reduction at various 2-IT/protein molar ratios. In vivo inflammation targeting studies were carried out in a mouse ear edema model created by topical application of 12-O-tetradecanoyl-phorbol-13-acetate (TPA) on the right ear of ICR mice.
  
  
  RESULTS
  Radiolabeling yields increased with increasing amounts of 2-IT. When the 2-IT/protein ratio reached 1000, the radiolabeling yield was greater than 90% without significant colloid production. TPA-treated ears showed high radioligand uptake, which was clearly detected by SPECT and autoradiographic imaging. The activities (%ID/g) in the inflamed and control ears at 3h after injection were 2.76 ± 0.20 vs. 0.69 ± 0.12 for IF, 5.86 ± 0.40 vs. 2.86 ± 0.61 for TF, and 7.61 ± 0.86 vs. 1.99 ± 0.31 for TFI (P<0.05 vs. controls). TFI showed significantly higher uptake in the inflamed ears compared to TF and IF (P<0.05). Blocking study results indicated specificity of radioligand binding with decreased radioactive uptake in the inflamed ears. Western blotting and ELISA analysis further confirmed a high expression of IL-1β and TNF-α in the inflamed ears.
  
  
  CONCLUSIONS
  (99m)Tc-labeled cytokine ligands are a promising approach for detecting and understanding the inflammatory process. TFI may be more useful than the single-domain ligands for noninvasive detection of inflammatory sites.
authors: Department of Medical Imaging, University of Arizona, Tucson, AZ, USA. zliu@radiology.arizona.edu
authors: Zhonglin
authors: Z
authors: Liu
authors: Leonie
authors: L
authors: Wyffels
authors: Christy
authors: C
authors: Barber
authors: Li
authors: L
authors: Wan
authors: Hua
authors: H
authors: Xu
authors: Mizhou M
authors: MM
authors: Hui
authors: Lars R
authors: LR
authors: Furenlid
authors: James M
authors: JM
authors: Woolfenden
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004355:Drug Stability; D004487:Edema / Q000139:chemically induced / Q000175:diagnosis / Q000378:metabolism; D005786:Gene Expression Regulation; D007249:Inflammation / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D053590:Interleukin 1 Receptor Antagonist Protein / Q000378:metabolism* / Q000493:pharmacokinetics; D007375:Interleukin-1 / Q000378:metabolism*; D053583:Interleukin-1beta / Q000378:metabolism; D007553:Isotope Labeling; D008024:Ligands; D008297:Male; D051379:Mice; D057054:Molecular Imaging / Q000379:methods*; D015609:Organotechnetium Compounds; D021381:Protein Transport; D051381:Rats; D047889:Receptors, Tumor Necrosis Factor, Type II / Q000378:metabolism* / Q000493:pharmacokinetics; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1; D053583:Interleukin-1beta; D008024:Ligands; D015609:Organotechnetium Compounds; D047889:Receptors, Tumor Necrosis Factor, Type II; D014409:Tumor Necrosis Factor-alpha; D013755:Tetradecanoylphorbol Acetate