The cytokine interleukin-8 (IL-8) binds with high affinity to the CXCR1 and CXCR2 receptors on neutrophils. In previous studies, we showed that (99m)Tc-IL-8 could rapidly and effectively delineate foci of infection and inflammation in rabbit models of intramuscular infection, colitis, and osteomyelitis. Here, the in vivo kinetics and pharmacodynamics of (99m)Tc-IL-8 are studied in detail. A derivative of hydrazinonicotinamide (HYNIC) was used as a bifunctional coupling agent to label the protein with (99m)Tc.
Methods: To address specificity of uptake of (99m)Tc-IL-8 in the abscess, uptake in turpentine-induced abscesses in neutropenic rabbits was compared with uptake in turpentine-induced abscesses in normal rabbits. The pharmacokinetics of (99m)Tc-IL-8 were studied in neutropenic rabbits and compared with those in normal rabbits. To investigate the interaction of (99m)Tc-IL-8 with blood cells in circulation in normal rabbits, the distribution of the radiolabel over circulating white and red blood cells and plasma was determined. The in vivo kinetics of (99m)Tc-IL-8 were studied by quantitative analysis of whole-body images acquired between 0 and 6 h after injection. The results of this analysis (in vivo biodistribution) were validated by ex vivo counting of radioactivity in dissected tissues.
Results: The abscess uptake (percentage of injected dose per gram of tissue [%ID/g] +/- SEM) in immunocompetent rabbits (0.41 +/- 0.05) was 10 times higher than that in neutropenic rabbits (0.038 +/- 0.014), demonstrating specificity of the target uptake of (99m)Tc-IL-8. Abscess-to-muscle ratios +/- SEM were also 10 times higher (110 +/- 10 vs. 10 +/- 5). Lung and spleen uptake in normal rabbits was 3 times higher than that in neutropenic rabbits. The blood clearance of the radiolabel in neutropenic rabbits was similar to that in normal rabbits. In circulation, most of (99m)Tc-IL-8 (70%) was found in the plasma fraction. Less than one third was associated with red blood cells, and only a very low percentage (<2.5%) was associated with white blood cells. Image analysis revealed a gradually increasing abscess uptake over time up to >15%ID, which was confirmed by ex vivo gamma-counting of the infected muscle. The highest increase in uptake in the abscess was observed after 2 h following injection, when most of (99m)Tc-IL-8 was cleared from the blood, suggesting specific neutrophil-mediated accumulation of (99m)Tc-IL-8 in the abscess. Furthermore, region-of-interest analysis revealed that gradual accumulation of (99m)Tc-IL-8 in the abscess was accompanied by a simultaneous clearance of activity from the lungs, suggesting that neutrophil-associated (99m)Tc-IL-8 that was initially trapped in the lungs migrates to the abscess at later time points, favoring neutrophil-bound transportation from the lungs to the abscess.
Conclusion: Substantial support is given for the hypothesis that (99m)Tc-IL-8 localizes in the abscess, mainly bound to peripheral neutrophils. Accumulation in the abscess is a highly specific, neutrophil-driven process. As assessed by in vivo and ex vivo analysis, the total fraction that accumulates in the inflamed tissue is extremely high (up to >15 %ID) compared with that of other agents used for imaging infection and inflammation.