Characterization of 99mTc-labeled cytokine ligands for inflammation imaging via TNF and IL-1 pathways

Zhonglin Liu, Leonie Wyffels, Christy Barber, Li Wan, Hua Xu, Mizhou M. Hui, Lars R Furenlid, James M. Woolfenden

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Introduction: 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 99mTc-labeled forms of these ligands, 99mTc-IL-1ra-Fc (IF), 99mTc-TNFR2-Fc (TF), and 99mTc-TNFR2-Fc-IL-1ra (TFI), for inflammation imaging. Methods: The cytokine ligands were labeled with 99mTc 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 3. h 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: 99mTc-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.

Original languageEnglish (US)
Pages (from-to)905-915
Number of pages11
JournalNuclear Medicine and Biology
Volume39
Issue number7
DOIs
StatePublished - Oct 2012

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Receptors, Tumor Necrosis Factor, Type II
Interleukin 1 Receptor Antagonist Protein
Interleukin-1
Cytokines
Ear
Ligands
Inflammation
Inbred ICR Mouse
Interleukin-1 Receptors
Colloids
Tetradecanoylphorbol Acetate
Single-Photon Emission-Computed Tomography
Edema
Proteins
Acetates
Western Blotting
Enzyme-Linked Immunosorbent Assay

Keywords

  • Tc
  • Cytokines
  • Imaging
  • Inflammation
  • Interleukin-1
  • Tumor necrosis factor

ASJC Scopus subject areas

  • Cancer Research
  • Molecular Medicine
  • Radiology Nuclear Medicine and imaging

Cite this

Characterization of 99mTc-labeled cytokine ligands for inflammation imaging via TNF and IL-1 pathways. / Liu, Zhonglin; Wyffels, Leonie; Barber, Christy; Wan, Li; Xu, Hua; Hui, Mizhou M.; Furenlid, Lars R; Woolfenden, James M.

In: Nuclear Medicine and Biology, Vol. 39, No. 7, 10.2012, p. 905-915.

Research output: Contribution to journalArticle

Liu, Zhonglin ; Wyffels, Leonie ; Barber, Christy ; Wan, Li ; Xu, Hua ; Hui, Mizhou M. ; Furenlid, Lars R ; Woolfenden, James M. / Characterization of 99mTc-labeled cytokine ligands for inflammation imaging via TNF and IL-1 pathways. In: Nuclear Medicine and Biology. 2012 ; Vol. 39, No. 7. pp. 905-915.
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abstract = "Introduction: 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 99mTc-labeled forms of these ligands, 99mTc-IL-1ra-Fc (IF), 99mTc-TNFR2-Fc (TF), and 99mTc-TNFR2-Fc-IL-1ra (TFI), for inflammation imaging. Methods: The cytokine ligands were labeled with 99mTc 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 3. h 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: 99mTc-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.",
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T1 - Characterization of 99mTc-labeled cytokine ligands for inflammation imaging via TNF and IL-1 pathways

AU - Liu, Zhonglin

AU - Wyffels, Leonie

AU - Barber, Christy

AU - Wan, Li

AU - Xu, Hua

AU - Hui, Mizhou M.

AU - Furenlid, Lars R

AU - Woolfenden, James M.

PY - 2012/10

Y1 - 2012/10

N2 - Introduction: 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 99mTc-labeled forms of these ligands, 99mTc-IL-1ra-Fc (IF), 99mTc-TNFR2-Fc (TF), and 99mTc-TNFR2-Fc-IL-1ra (TFI), for inflammation imaging. Methods: The cytokine ligands were labeled with 99mTc 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 3. h 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: 99mTc-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.

AB - Introduction: 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 99mTc-labeled forms of these ligands, 99mTc-IL-1ra-Fc (IF), 99mTc-TNFR2-Fc (TF), and 99mTc-TNFR2-Fc-IL-1ra (TFI), for inflammation imaging. Methods: The cytokine ligands were labeled with 99mTc 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 3. h 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: 99mTc-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.

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KW - Cytokines

KW - Imaging

KW - Inflammation

KW - Interleukin-1

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