Delineation of Hypoxia in Canine Myocardium Using PET and Copper(II)-Diacetyl-bis(N4-Methylthiosemicarbazone)

Delineation of Hypoxia in Canine Myocardium Using PET and Copper(II)-Diacetyl-bis(N⁴-Methylthiosemicarbazone)

Jason S. Lewis, PhD¹, Pilar Herrero, MS¹, Terry L. Sharp¹, John A. Engelbach¹, Yasuhisa Fujibayashi, PhD, DMedSci², Richard Laforest, PhD¹, Attila Kovacs, MD¹, Robert J. Gropler, MD¹ and Michael J. Welch, PhD¹

¹ Division of Radiological Sciences, Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, Missouri
² Biomedical Imaging Research Center, Fukui Medical University, Matsuoka, Fukui, Japan

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FIGURE 1. Two-compartment model used to model biokinetics of *Cu-ATSM in myocardium.

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FIGURE 2. Phantom of Derenzo et al. (23) for ⁶⁴Cu (A) and ⁶⁰Cu (B). Unfavorable emissions associated with decay of ⁶⁰Cu have not reduced image quality in comparison with ⁶⁴Cu phantom in ECAT 962 HR⁺ scanner. Values shown are bore sizes, in millimeters.

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FIGURE 3. Time-activity curves generated from direct arterial sampling after arterial administration of 37 MBq of ⁶⁴Cu-ATSM to generate input function (n = 3 dogs). (A) Plot shows percentage of octanol-extractable ⁶⁴Cu-ATSM with respect to total ⁶⁴Cu-ATSM blood activity as function of time. (B) Plot shows total amount of radioactivity in arterial blood after injection of ⁶⁴Cu-ATSM as function of time and amount of radioactivity in form of ⁶⁴Cu-ATSM.

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FIGURE 4. Graphs show results for monoexponential analysis of *Cu-ATSM PET (protocol I) (A), kinetic modeling analysis of *Cu-ATSM (protocol I) (B), monoexponential analysis after injection of *Cu-ATSM (protocol II at 3 h) (C), and kinetic modeling analysis after injection of *Cu-ATSM (protocol II at 3 h) (D). Global hypoxia increased tracer retention (B). In each animal, LAD occlusion significantly decreased MBF and concomitantly increased tracer retention. Individual (joined points) and mean values for all animals (far right and far left) of myocardial retention of *Cu-ATSM are given. SD error bars associated with each animal are on individual points; if not seen, bars are within symbol.

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FIGURE 5. (A) Reconstructed midventricular functional PET images after LAD occlusion at 3 and 24 h (protocol II). Top 3 images were obtained after first *Cu-ATSM injection, 3 h after coronary occlusion, and bottom 3 were obtained after second injection, 24 h later. Horizontal long-axis view is shown, with septum on left, apex on top, and lateral free wall on right. Uptake of tracer is reduced in apical region at both 3 h and 24 h after LAD occlusion, indicating sustained hypoperfusion (perfusion image counts per pixel per minute). In contrast, retention of *Cu-ATSM (counts per pixel per minute) in apex is higher at 3 h after occlusion than at 24 h. This increase in retention at 3 h can clearly be seen after tracer retention is normalized to tracer uptake (*Cu-ATSM retention/perfusion [no units]), indicating hypoxic but viable tissue. No increase in retention is observed 24 h after normalization. Absence of *Cu-ATSM retention in this region 24 h after occlusion is consistent with necrosis, as was confirmed through postmortem TTC staining of heart. (B) Reconstructed midventricular short-axis functional PET images (counts per pixel per minute) show myocardial tracer activity after bolus injection of *Cu-ATSM (protocol III). Under resting conditions, tracer activity in anterior region is diminished, consistent with hypoperfused but normoxic myocardium. In contrast, during DOB, anterior myocardium shows increased tracer activity despite diminished perfusion to this region, suggesting hypoxia.

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FIGURE 6. (A) On right, TTC-stained 0.5-cm section of myocardium 24 h after occlusion shows significant necrosis. On left, electronic autoradiography shows absence of ⁶⁴Cu-ATSM retention in this region 24 h after occlusion, consistent with necrosis. Autoradiograph also shows area of increased ⁶⁴Cu-ATSM accumulation bordering necrotic regions, indicating hypoxic zone surrounding necrotic zone. (B–D) Graphs show MBF (B), 1/k_mono retention (C), and 1/k₄ retention (D) as ratios of LADR to NLR at 3 and 24 h. (C and D) *Cu-ATSM myocardial retention patterns reveal that tracer retention did not increase 3 h after LAD occlusion in dogs with small infarcts but increased significantly in dogs with decreased MBF in LADRs. At 24 h, MBF in LADRs was severely reduced in 3 of 4 animals (B). Retention values obtained from kinetic modeling (D) show that, in animals with severely reduced MBF at 24 h, retention in LADR decreased. Conversely, retention increased in the only animal (dog 11) in which MBF remained within 80% of normal. This pattern is consistent with necrotic tissue and with infarct size observed through postmortem staining. These data also demonstrate that retention and clearance of *Cu-ATSM are not related to perfusion.

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FIGURE 7. (A) In myocardial time-activity curves obtained after injection of ⁶⁴Cu-ATSM in dog 13 at rest (protocol III), normal septal/lateral region ( ${diamondsuit}$ ) and apical region ( ${blacksquare}$ ) show similar kinetics. Although data collection continued until 45 min, figure shows only first 12 min, for clarity. (B) In myocardial time-activity curves obtained after injection of ⁶⁴Cu-ATSM in dog 13 after DOB infusion (protocol III), normal septal/lateral region ( ${diamondsuit}$ ) shows rapid washout kinetics but apical region ( ${blacksquare}$ ) shows uptake and excellent tracer retention, indicating hypoxia. Although data collection continued until 45 min, figure shows only first 12 min, for clarity. (C-E) Graph of MBF (ratio of LADR to NLR [endocardium to epicardium]) (C) and graphs of 1/k_mono retention (D) and 1/k₄ retention (E) expressed as ratios between LADR and NLR at rest and stress show myocardial retention of ⁶⁴Cu-ATSM for each dog in protocol III. In all cases, DOB infusion significantly increased tracer retention in damaged apical regions (P < 0.005), despite increase in MBF (Table 3).