Immuno-PET Imaging of Atherosclerotic Plaques with [(89)Zr]Zr-Anti-CD40 mAb—Proof of Concept

SIMPLE SUMMARY: Atherosclerosis is characterized by gradual plaque build-up in the middle and large arteries and is the major cause of cardiovascular disease. Determining which plaques are prone to rupture and cause potential lethal effects (e.g., myocardial infarction) could greatly reduce the potential bad outcomes. CD40 is a co-stimulatory molecule present in various cells in the plaque and has been shown to correlate with plaque vulnerability. In this manuscript, we have combined a murine monoclonal antibody against CD40 (a costimulatory molecule directly linked to plaque progression and present on many cells within the lesion) with Zirconium-89 to test its applicability to detect lesions in a mouse model of atherosclerosis using PET/CT. We show that this Zirconium-89 labeled antibody can detect CD40 in atherosclerotic lesions in a mouse model of atherosclerosis. In wild type mice without plaques, no signal was found. Our results suggest that CD40 could be a potential marker for PET imaging of plaque inflammation and vulnerability. ABSTRACT: Non-invasive imaging of atherosclerosis can help in the identification of vulnerable plaque lesions. CD40 is a co-stimulatory molecule present on various immune and non-immune cells in the plaques and is linked to inflammation and plaque instability. We hypothesize that a (89)Zr-labeled anti-CD40 monoclonal antibody (mAb) tracer has the potential to bind to cells present in atherosclerotic lesions and that CD40 Positron Emission Tomography (PET) can contribute to the detection of vulnerable atherosclerotic plaque lesions. To study this, wild-type (WT) and ApoE(−/−) mice were fed a high cholesterol diet for 14 weeks to develop atherosclerosis. Mice were injected with [(89)Zr]Zr-anti-CD40 mAb and the aortic uptake was evaluated and quantified using PET/Computed Tomography (CT) imaging. Ex vivo biodistribution was performed post-PET imaging and the uptake in the aorta was assessed with autoradiography and compared with Oil red O staining to determine the tracer potential to detect atherosclerotic plaques. On day 3 and 7 post injection, analysis of [(89)Zr]Zr-anti-CD40 mAb PET/CT scans showed a more pronounced aortic signal in ApoE(−/−) compared to WT mice with an increased aorta-to-blood uptake ratio. Autoradiography revealed [(89)Zr]Zr-anti-CD40 mAb uptake in atherosclerotic plaque areas in ApoE(−/−) mice, while no signal was found in WT mice. Clear overlap was observed between plaque areas as identified by Oil red O staining and autoradiography signal of [(89)Zr]Zr-anti-CD40 mAb in ApoE(−/−) mice. In this proof of concept study, we showed that PET/CT with [(89)Zr]Zr-anti-CD40 mAb can detect atherosclerotic plaques. As CD40 is associated with plaque vulnerability, [(89)Zr]Zr-anti-CD40 mAb has the potential to become a tracer to detect vulnerable atherosclerotic plaques.