Abstract 4959: Imaging of Atherosclerotic Lesions in Apoe−/− Mice by Targeting of CD40
We investigated whether in-vivo imaging of the CD40 receptor can be used for identification of inflammatory atherosclerotic lesions.
Methods: Monoclonal antibodies against CD40 (CD40ab), IgGA2 (Coab) and Fab2 fragments (CD40Fab) were coupled to the near-infrared fluorescence (NIRF) dyes Cy5′5 or Cy7. Apolipoprotein E deficient mice (apoE−/−) and apoE−/−CD40−/− deficient mice (apoE−/−CD40−/−) were fed with high cholesterol diet for 8 –12 month. Antibody conjugates were injected into the tail vein and mice were investigated 6 to 48 hours after injection by conventional NIRF imaging, fluorescence microscopy of the aortic arch and in-vivo 3D fluorescence molecular tomography (FMT). Sudan staining and histological analysis were performed after imaging procedures.
Results: Application of Cy5′5-labeled CD40ab resulted in accumulation of fluorescence signal intensity mainly in Sudan-positive areas of the thoracic aorta, which was also seen with the Coab at high concentrations. Fluorescence increased slightly after 24 hours compared to 6 hours after injection. In vivo imaging was not feasible using conjugated antibodies due to circulating intravasal fluorescence (>48h). Significant background fluorescence was detected after application of CD40ab and Coab in all mice, probably due to FC receptor-dependent binding. Application of Cy7-CD40Fab induced strong in-vivo fluorescence detected by FMT in projection to the aortic arch and carotid arteries of apoE−/− (n=4), but not in apoE−/−CD40−/− mice (n=4). No signal was observed in mice injected with unlabelled Cy7. Ex-vivo imaging of the thoracic aorta confirmed a Cy7 signal in atherosclerotic lesions of apoE−/−, but not in apoE−/−CD40−/− after application of CD40Fab. Data were also confirmed by microscopic fluorescence imaging of the aortic wall and atherosclerotic lesions.
Conclusion: In vivo detection of CD40 in atherosclerotic plaques was feasible using antibody fragments but not with full anti-CD40 antibodies. The approach is applicable for detection of vascular inflammation, since CD40 is strongly associated with inflammatory lesion formation.